The cutaneous microvascular architecture of human diabetic toe studied by corrosion casting and scanning electron microscopy analysis.

In this morphological study, we report on the three-dimensional microvascular architecture constituting the toes of a patient affected by diabetic microangiopathy. We applied corrosion casting (CC) technique to the toes of a patient affected by Type 2 diabetes, who underwent surgery for explantation of inferior left limb due to necrotic processes of soft tissues. The toes of a foot traumatically explanted in a motorcycle accident were kept as controls. According to technical protocols, toes were injected with a low-viscosity acrylic resin (Mercox) through the major digital artery, tissues were corroded in KOH solution (8%), and resulting casts processed for SEM observations. Already at low magnification, in diabetic toes, we found an impairment of the linear track-like disposition of the vessels of plantar side, with signs of vascular disruption and obliterations, stopped resin, and leakages. Capillaries under the nail and a lot of vascular villi in eponychium and nail borders were damaged, and vascular regression phenomena acting on them were clearly visible. Resin leakages and impairment of normal vascular architecture were also observed in the root of the nail. This preliminary report represents only the first step for further investigations regarding morphological three-dimensional appearance of diabetic microangiopathy. CC and scanning electron microscopy technique well documented these morphological modifications, highlighting on both structural and ultrastructural features of diabetic toes microvessels. In conclusion, our qualitative data try to better focus on the pathophysiological mechanisms involved in diabetic dermopathy and microangiopathy, proposing CC as useful method to investigate on them.

In vitro and in vivo characterization of A-940894: a potent histamine H4 receptor antagonist with anti-inflammatory properties.

BACKGROUND AND PURPOSE: The histamine H4 receptor is widely expressed in cells of immune origin and has been shown to play a role in a variety of inflammatory processes mediated by histamine. In this report, we describe the in vitro and in vivo anti-inflammatory properties of a potent histamine H4 receptor antagonist, A-940894 (4-piperazin-1-yl-6,7-dihydro-5H-benzo[6,7]cyclohepta[1,2-d]pyrimidin-2-ylamine). EXPERIMENTAL APPROACH: We have analysed the pharmacological profile of A-940894 at mouse native, rat recombinant and human recombinant and native, histamine H4 receptors by radioligand binding, calcium mobilization, mast cell shape change, eosinophil chemotaxis assays and in the mouse model of zymosan-induced peritonitis. KEY RESULTS: A-940894 potently binds to both human and rat histamine H4 receptors and exhibits considerably lower affinity for the human histamine H1, H2 or H3 receptors. It potently blocked histamine-evoked calcium mobilization in the fluorometric imaging plate reader assays and inhibited histamine-induced shape change of mouse bone marrow-derived mast cells and chemotaxis of human eosinophils in vitro. In a mouse mast cell-dependent model of zymosan-induced peritonitis, A-940894 significantly blocked neutrophil influx and reduced intraperitoneal prostaglandin D2 levels. Finally, A-940894 has good pharmacokinetic properties, including half-life and oral bioavailability in rats and mice. CONCLUSIONS AND IMPLICATIONS: These data suggest that A-940894 is a potent and selective histamine H4 receptor antagonist with pharmacokinetic properties suitable for long-term in vivo testing and could serve as a useful tool for the further characterization of histamine H4 receptor pharmacology.

Gait analysis of pre- and post-meniscectomy knee: a prospective study.

Degenerative changes in the knee joint after meniscectomy are well known. Although likely to be due to changed biomechanics, there is no evidence in the literature to identify the underlying biomechanical alterations. The aim of this study was to analyse lower limb gait biomechanics before and after meniscectomy. Ten patients who required partial medial meniscectomy for irreparable meniscal tear took part in motion analysis before surgery, then at 6 and 12 months post-operatively. A control group was also set up consisting of 10 healthy volunteers. Joint kinematics did not show significant alterations between pre-operative and 6 month post-operative evaluations. However flexion increases at the hip, knee and ankle joint were observed in late swing and early stance phase 12 months after surgery. Hip and knee flexion-extension moments were affected with knee moment altered both before and after surgery. Before surgery and at 6 months after, the changes occurred mainly at the point of push off, while at 12 months they occurred during the swing phase. Hip flexion-extension moment had also changed 12 months after surgery. When examining symmetry of gait patterns prior to surgery, there were differences between the flexion-extension moments of the healthy and of the injured knee at first impact and during late stance. After surgery, asymmetries were not more apparent at first impact, but in late stance phase a reduced knee extension moment in the injured limb was still present. Before surgery, the joint kinematics were not greatly altered and changes were mainly due to pain. After partial meniscectomy, the pain disappeared and new joint responses were observed. These could be caused by the altered mechanics and/or through proprioceptive mechanisms.

3D analysis of SEM images of corrosion casting using adaptive stereo matching.

The corrosion casting method represents one of the most widely used technique to study the 3D microvascularization of many tissues, both in their normal and pathological conditions. For a long time this technique was used only to perform a qualitative evaluation of the images obtained by scanning electron microscopy (SEM). A quantitative evaluation of vascular parameters (e.g., interbranching and intervascular distances, angle measurements, lengths and diameters) was lacking, mainly because of the difficulties found in the measurement performed on 2D SEM images. Then, some authors reported a quantitative method based on the analyses of stereo-pair images that allowed precise morphometric measurements. To visualize the specimens in 3D, it was necessary to use red-green glasses. In this article we describe a new approach by which we can automatically obtain a 3D reconstruction of vascular cast specimen's surface directly from stereo-images. Moreover, we developed a software that performed micrometric measurements on the 3D construct generated from the stereo-pictures. In conclusion, implementing together these two softwares and applying them to corrosion casting samples made it possible to render in 3D the surface of corrosion cast as well as make quantitative measurements on the corrosion casts.

The three-dimensional microvascular architecture of the human Kaposi sarcoma implanted in nude mice: a SEM corrosion casting study.

The human Kaposi sarcoma represents one of the most common skin lesions associated with AIDS. Its clinical presentation and anatomopathological structure seem to demonstrate a particularly rich vascularity. The latest therapies aim to limit its intrinsic angiogenic activity in an attempt to reduce vascular density and the formation of new vessels. For these reasons, we decided to study the microvascular architecture of Kaposi sarcoma in three dimensions. We used a corrosion casting technique applied to nude mice previously transplanted subcutaneously with human modified neoplastic Kaposi sarcoma cells. The cooption of host vessels made by the tumor was demonstrated by three-dimensional scanning electron microscopy (SEM) images. At high magnification several angiogenic patterns were observed in the form of potato-shaped vessels, sprouts, intussusceptions and mouse tailed end tipped capillaries along with some ultrastructural features such as intercellular extravasations and endothelial cell modifications. Our investigation allowed us to build a detailed map of tumor vasculature in human Kaposi sarcoma. Furthermore, this study want to shed light on the sharp morphological three-dimensional conformation of angiogenic sprouts so to be able to better understand their modifications occurred during time and after antiangiogenic experimental therapies, by now observed only by immunohistochemical or immunofluorescent assays.

In situ hybridization by scanning electron microscopy for painting, centromeric, and YAC localization.

The hybridization site of a DNA probe was detected using a scanning electron microscope (SEM), modifying the standard in situ hybridization (ISH) method. The experiments were performed on human metaphases obtained from lymphocyte cultures of human peripheral blood. The libraries and probes used were: 1-chromosome library for the painting of chromosome 1 (wcp 1), an alphoid centromere-specific probe of chromosome 8 (pZ8.4), and the yeast artificial chromosome (YAC) 964-C10 mapped at band p13 on chromosome 12. These probes were labeled by nick translation with biotin and displayed with a gold-conjugated anti biotin goat antibody. The gold signal was amplified by silver enhancement. The chromatides appeared as packages of thin filaments 120 nm high; some of them collapsed, probably due to ISH procedures. All the probes were clearly detected as small gold particles grouped on the surface of the target chromosomes and chromosome sites. Thus, this procedure is useful to clarify the positional relationship between the chromatin filaments and the probe.

Plexiform vascular structures in the human digital dermal layer: a SEM--corrosion casting morphological study.

This study aimed to describe the impressive diversity of vascular plexiform structures of the hypodermal layer of human skin. We chose the human body site with the highest concentration of dermal corpuscles, the human digit, and processed it with the corrosion casting technique and scanning electron microscopy analysis (SEM). This approach proved to be the best tool to study these microvascular architectures, free from any interference by surrounding tissues. We took high-definition pictures of the vascular network of sweat glands, thermoreceptorial and tactile corpuscles, the vessels constituting the glomic bodies and those feeding the hair follicles. We observed that the three-dimensional disposition of these vessels strictly depends on the shape of the corpuscles supplied. We could see the tubular vascularization of the excretory duct of sweat glands and the ovoid one feeding their bodies, sometimes made up of two lobes. In some cases, knowledge of these morphological data regarding the normal disposition in space and intrinsic vascularization structure of the dermal corpuscles can help to explain many of the physiopathological changes occurring during chronic microangiopathic diseases.

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.

The microvasculature of the lateral choroid plexus in the rat: a scanning electron microscopy study of vascular corrosion casts.

The three-dimensional microvascular structure of many organs can be adequately investigated only using the corrosion casting technique. We applied this method, consisting of an injection of low viscosity acrylic resin through the major vessels and the subsequent digestion of the organic component with strong alkali or acids, to the choroid plexus of the lateral ventricles in the rat, focusing on its structural vascular features. This approach allowed a qualitative morphological description of the choroid plexus of the lateral ventricles, revealing several aspects of their capillary architecture as well as the morphological details underlying its main functional activity, essential to cerebrospinal fluid turnover. Observation of the casts with scanning electron microscopy gave a detailed picture of the choroid plexus of the lateral ventricles of the rat and enabled us to distinguish four different regions, depending on the site that the lateral ventricles occupied: the anterior olfactory region, the main central region, the longer branch and the inferior horn. Each region mostly consisted of spiral capillaries and had specific characteristics. At high magnification, the casts revealed distinctive vascular specializations, such as numerous bulges regularly placed on the capillaries. This morphological investigation underpins a better comprehension of the pathological mechanisms involving the choroid plexus in the lateral ventricles.

A new method to make vascular and bronchial casts of voluminous organs.

Vascular and bronchial endocasts represent a useful instrument to study the ramification pattern of these structures. Casts have been made from different materials, such as waxes in ancient times and, more recently, silicon-like compounds or resins (see e.g. Mercox) to study the finest details. These techniques are valuable for small specimens, whereas they are inadequate for very large organs, where technical difficulties require the development of specific instrumentation. In this study we present a new simple injection technique, based on expanded polyurethane, which allows preparing vascular and bronchial trees for macroscopic and microscopic studies. The new injection technique is very easy to carry out, since the propulsion is provided by compressed air, and it does not require special instrumentation. To this aim, endocasts of the entire tracheal-bronchial tree and casts of vascular kidney from different animals were prepared. The specimens have a very low weight, show the finest ramifications, and are very stable and resistant to mechanical stress. To examine microscopically the details of the casts, specimens from the kidney cast were also analyzed by scanning electron microscopy, revealing good preservation of microcirculatory structures, functional sphincters and endothelial cell impressions. Therefore, the technique may be useful for macroscopic studies of large specimens, retaining sufficiently fine details.

[CT pulmonary density mapping: surgical utility]. / Mappe TAC di isodensità polmonare: significato chirurgico.

BACKGROUND: The present paper considers the technique of CT scan maps of pulmonary isodensity, examining lung density differences as a function of the type of disease and considering their significance for the purposes of refined, useful diagnosis in a surgical context. METHODS. The method is used to examine 3 groups of subjects selected on a clinical/anamnestic basis and a further group already admitted for surgery. For each patient we obtained 2 thoracic density scans during the phase of maximum inspiration and expiration. On each scan we constructed 50 isodensity maps, the equivalent of more than 2500 measurements: the preliminary standard was represented by 100 wide windows to produce total "illumination" of the pulmonary fields. The isodensity windows were then codified differently. Subsequently, the density scans were analysed with the technique of scalar decomposition. RESULTS: The CT scan maps of lung isodensity proved useful for certain lung diseases in which early diagnosis, topographic extent of the pathology and the refined definition of the pathological picture provide important solutions as regards the indication and planning of surgical treatment and for the evaluation of the operative risk and prognosis. CONCLUSIONS: We consider that the technique is rapidly performed, not complex and inexpensive and is able to supply detailed information on the lung parenchyma such as to be used not only as a routine technique but also in emergencies.

[Hemorrhoids: epidemiology and correlation with chronic constipation]. / Malattia emorroidaria: inquadramento epidemiologico e correlazione con la stipsi cronica.

Following a review of international literature, AA report main results and refer their opinion about the correlation of hemorrhoidal disease with constipation, considering some variants as well as age, sex, breed, social-economic condition and geographic distribution in USA, England and Wales. Epidemiologically ten millions of people, in USA, are affected by hemorrhoidal disease; the incidence rate is 4.4% with an age distribution that shows a prevalence between 45-65 years old subject while constipation has an exponential increase with aging. Hemorrhoidal disease is significantly influenced by sex and geographic distribution that is in white breed more than in black, in social high class and in men more than women. In black breed constipation and hemorrhoidal disease present especially in lower social classes. Based on these results hemorrhoidal disease shows on epidemiological pattern that differ from constipation's one. Many questions are still present about correlations between hemorrhoidal disease and chronic constipation regarding etiopathogenesis. Only future case-control studies will solve the problem.

[Hemorrhoid disease. Physiopathology, etiopathology and surgical approach]. / La malattia emorroidaria. Riflessioni fisiopatologiche ed etiopatologiche in funzione dell'approccio chirurgico.

The authors report a number of cases of hemorrhoid disease and describe the therapeutic iter followed with particular reference to the surgical approach used. After a description of the physiopathological aspects linked to the disease, bearing in mind the use of electromanometry and electromyography in diagnosis, the authors underline the contemporary presence of varices in the lower limbs and hemorrhoid disease, as well as the frequent finding of hemorrhoids in a syndrome of portal hypertension. They then affirm how it is impossible to establish the causes of this pathology with any certainty and how a single standardised treatment plan is untenable. The authors then indicate the guidelines used to choose the most appropriate form of surgery rather than pharmacological treatment, based on the ideal cases and conditions for surgery. The ultimate goals of surgery are also outlined. The study compares four possible surgical techniques, providing synthetic information regarding their adaptability to the various cases treated and the characteristics of their use. This means that, once decided, surgery must successfully resolve the patient's problems. In conclusion, once hemorrhoid disease has been diagnosed, it is important to intervene with appropriate medical treatment to control the evolution of the pathology; if this is not sufficient, surgery becomes an inevitable choice.

Evidence for nicotinic receptors potentially modulating nociceptive transmission at the level of the primary sensory neuron: studies with F11 cells.

F11 cells are a dorsal root ganglion (DRG) cell line used to model the function of authentic type C, peptidergic, nociceptive neurons. The cellular events underlying the antinociceptive effects of (+/-)-epibatidine, a nicotinic acetylcholine receptor (nAChR) ligand that is 200-fold more potent than morphine, is unknown. The present study investigated the ability of cholinergic channel activators (ChCAs) to effect nAChR-gated ion flux and modulate the release of substance P (SP), a neuropeptide identified to play a critical role in nociception. The prototypical agonists (-)-nicotine and (-)-cytisine, the ganglionic stimulant 1,1-dimethyl-4-phenylpiperazinium, the novel ChCA ABT-418 [(S)-3-methyl-5-(-1-methyl-2-pyrrolidinyl)isoxazole], and (+/-)-epibatidine evoked a concentration-dependent stimulation of rubidium (86Rb+) efflux with EC50 values of 14.2 +/- 1.6, 63.4 +/- 24, 3.8 +/- 2.0, 29.8 +/- 2.6, and 0.019 +/- 0.001 microM as well as maximal intrinsic activities of 100, 97, 69, 75, and 102%, respectively. The noncompetitive nAChR antagonist mecamylamine potently antagonized (-)-nicotine-evoked ion flux, whereas the competitive antagonist dihydro-beta-erythroidine was a weak antagonist, giving support to an alpha3beta4 nAChR subtype. In addition, concentrations of (+/-)-epibatidine, similar to those necessary to induce maximal 86Rb+ efflux, evoked spontaneous release of SP from these cells, which was blocked by mecamylamine. Furthermore, prolonged exposure to (+/-)-epibatidine desensitized the functional response of the nAChR in this cell line (IC50 = 12 +/- 9 nM). These findings in F11 cells provide a model to investigate the role nAChRs play in modulating DRG cell function, and may lead to insights into the role these receptors have in modulating nociceptive transmission.

Effect of apolipoprotein E on neurite outgrowth and beta-amyloid-induced toxicity in developing rat primary hippocampal cultures.

The correlation between the epsilon 4 allele of apolipoprotein E (apoE) and Alzheimer's disease is well established. However, the role of apoE in normal as well as pathological brain processes remains unclear. We evaluated the effect of apoE treatment on development and beta-amyloid (A beta)-induced toxicity using primary cultures of developing rat hippocampal neurons. The source of apoE was conditioned media from HEK cells stably transfected with human apoE3 or apoE4 cDNA, a preparation where apoE is lipid-associated. Morphological and biochemical changes in the cultures were assessed at 1 and 3 days following low- and high-density plating with either apoE3 or E4 with or without A beta. Both apoE isoforms were neurotrophic, as measured by increased neurite length. Aged A beta(1-42), a peptide preparation exhibiting extensive fibril and aggregate formation, is toxic to these cultures. Addition of apoE3 and E4 significantly and comparably attenuated the A beta-induced reduction in both neurite length and cell viability. The level of protection against this toxicity was proportional to the neurotrophic actions of the two apoE isoforms. Thus, apoE acts as a potent growth factor in both the absence and the presence of A beta, supporting a potentially important role for apoE in neurobiology.

The presence of the complement cascade does not lead to neuronal cell death in primary hippocampal cultures.

To investigate the consequences of complement activation on neuronal viability, the effects of serum treatment on neuron-rich and mixed neuronal/glial cultures were evaluated. The neurotoxicity observed following treatment with either human or rat serum was variable and did not appear to be mediated through a complement-mediated mechanism. Serum lots lacking CH50 activity induced neurotoxicity, and heat treatment of toxic lots of either human or rat sera did not abolish toxicity. In cases where serum treatment did not induce cell death, treatment with PIPLC to remove endogenous membrane-bound complement inhibitors prior to serum exposure, did not result in cell death.

Inhibition of age-induced beta-amyloid neurotoxicity in rat hippocampal cells.

The conditions under which beta-amyloid (Abeta) is toxic to primary rat hippocampal neurons were investigated. Synthetic Abeta(1-42) peptide was neurotoxic following "aging" for 7 to 14 days at 37 degrees C in modified Eagle's media. Neurotoxicity included decreases in neurite length, cell number, and metabolic state. In contrast, aging Abeta(1-42) in the presence of the media supplement B27 inhibited Abeta (1-42) induced neurotoxicity. Differences in the aggregation state of the two preparations did not account for differences in the biological activities elicited by each peptide. Since components of B27 include antioxidants as well as other agents that provide protection against oxidative damage, we suggest that free radicals may be responsible, in part, for the toxicity that occurs following the aging of the peptide.

[Traumatic tearing of the mesentery: immediate or deferred emergency surgery]. / Le lacerazioni traumatiche dei mesi: scelta chirurgica in urgenza immediata o differita.

We have considered a 1 year first-aid survey and focused a 6 checked mesentery vascular traumatic tearing cases. We have valued some hypotheses on pathogenic mechanism assumptions about these vascular injuries in abdomen trauma. Among these, we have focused our attention on the relationship between visceral stretching from alimentary bolus and abdominal vascular trauma. During diagnosis, we valued the importance of abdominal sounding, of echotomography and Computerized Tomography; the first one seems to be remarkable for reliability and quickness of accomplishment. In one case we have performed laparoscopic diagnostic-therapeutic approach. We suggested some standards for surgery choice related to injury amount. Monitoring is particularly important for those patients it can performed by invasive or image means seen that the unacknowledged endo-abdominal injury increases mortality by about 5-65%.

Characterization of cloned human dopamine D1 receptor-mediated calcium release in 293 cells.

Dopamine (DA) D1 receptors are generally known to couple only to Gs and cAMP production. Recently, D1 receptors expressed in mouse Ltk- cells have been shown to induce cAMP production, phosphoinositide (PI) hydrolysis, and calcium mobilization [Mol. Endocrinol. 6: 1815-1824 (1992)]. To further evaluate second messenger systems that could be activated by the D1 receptor, we examined the effects of DA, (R)-(+)-SKF-38393, and DA antagonists on cAMP production and calcium release in human embryonic kidney 293 cells stably expressing three different levels (Bmax = 0.12, 1.4, and 23 pmol/mg of protein) of the human D1 receptor. DA and (R)-(+)-SKF-38393 activated cAMP production and calcium release in all three D1-293 clones, and their potency was proportional to receptor density. The efficacy of SKF-38393 was also increased with receptor density in both cAMP and calcium studies. The effect of DA on calcium release consisted of a transient peak response (< 20 sec) that declined to an ethylene glycol bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid-sensitive plateau level above the base-line (>5 min). The effect of DA on cAMP and calcium release was selectively inhibited by SCH23390, a selective D1 antagonist, and not by spiperone, a selective D2 antagonist. DA did not induce PI hydrolysis in any of the three receptor-expressing clones. A 24-hr pretreatment with cholera toxin (2 micrograms/ml) greatly attenuated the effect of DA on cAMP formation and calcium release. To address how DA could activate calcium release without enhancing PI hydrolysis, the effects of forskolin, thapsigargin, and isoproterenol (Iso) were studied. Similarly to the effects of DA, forskolin and Iso stimulated cAMP production and calcium release from D1-293 cells. Cells that were stimulated with Iso or forskolin showed a reduced response to subsequent addition of DA. Pretreatment of D1-293 cells with thapsigargin, a selective Ca2+-ATPase inhibitor, elicited calcium release from the inositol-1, 4, 5-trisphosphate-sensitive calcium store and attenuated the response to subsequent addition of DA. Carbachol stimulated PI hydrolysis and calcium release but had little effect on cAMP production. Prestimulation with carbachol abolished the calcium response to DA, Iso, or forskolin. These studies indicate that D1 receptor-mediated calcium mobilization in 293 cells is dependent on cAMP production and the cAMP-dependent calcium store is part of the inositol-1,4,5-trisphosphate-sensitive calcium pool.

beta-Amyloid-induced toxicity in rat hippocampal cells: in vitro evidence for the involvement of free radicals.

The conditions under which amyloid is toxic to primary rat hippocampal neurons were investigated. Synthetic A beta (1-42) peptide elicited neurotoxic activity following "aging" for 7 to 14 days at 37 degrees C in Modified Eagles Media. Neurotoxicity included decreases in neurite length, cell number, and a loss in 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide reduction. In contrast, the addition of the media supplement B27, during the aging process, promoted the neurotrophic actions of aged A beta (1-42), as indicated by an increase in neurite length and the number of cells possessing neurites, and attenuated toxicity. The differences in the biological actions elicited by these two preparations of aged peptide were attributed to the presence of the B27 components. B27 consists of a mixture of agents that provide protection against oxidative damage. In support, aging A beta (1-42) in the presence of superoxide dismutase and catalase, two components of B27, significantly reduced the toxic actions of peptide; hence, suggesting that free radicals may be required for the toxicity that accumulates during the aging of the peptide. To determine the contribution of particular amino acid residues in amyloid toxicity, studies were carried out with an aged preparation of the A beta (1-42) analog, A beta (1-42)Nle35. Findings from these studies suggest that the methionine residue may play a part, but is not required, for amyloid toxicity to occur.