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Chronic venous insufficiency (CVI) and venous ulcers in the lower areas are widespread problems with a prevalence of 1% - 2% in the western population. The pathophysiology of this pathological manifestation is not yet fully clarified and it can be assumed that a multifactorial genesis should be considered. Certainly the high pressure values ​​in the peripheral venous system, the defective valves in the capillary network, the increase in permeability and capillary neogenesis (discharge of liquids in the extracellular compartment - + edema), the discharge of fibrinogen and the subsequent formation of a pericapillary fibrin cuff are the most important mechanisms related to hypoxia and skin ischemia and represent the first step for the formation of skin ulcers of the lower areas. In addition, following ischemic events, white blood cells can cause damage to tissues through the release of free radicals, proteolytic enzymes and cytokines. Furthermore, several authors hypothesize a hypothesis of 'white cell entrapment' responsible for endothelial damage which translates into additional damage to microcirculation. NO plays a fundamental role in the circulatory system, albeit in different ways. In fact, under physiological conditions, O is produced enzymatically in small quantities and for a short period of time by endothelial cells (eNOS) and by nitroxyergic neurons that innervate the vessels (nNOS); under pathological conditions the production of NO is inducible in macrophages (iNOS) from bacterial LPS and / or from the cytokines of white blood cells (7-8) and is produced in large quantities and for a long period. The result of its action is a massive vasodilation, which leads to a microcirculatory stasis.

Medical histology Editor / s Nadir M. Maraldi, Carlo Tacchetti Author / s Saverio Cinti, Michelangelo Cordenonsi, Simona Corso, Ottavio Cremona, Massimo De Felici, Roberta Di Pietro, Nicoletta Gagliano, Silvia Giordano, Nadir M. Maraldi, Carla Martinelli, Alessandro Moretta, Sergio Morini, Beatrice Nico, Maria Prat, Domenico Puzzolo, Domenico Ribatti, Giovanni Francesco Spatola, Carlo Tacchetti ISBN 9788870513899 Medical histology Medical histology is a modern text more oriented in the "medical" sense, in order to provide students with the tools to frame the subject in the context of preclinical disciplines. In it are presented experimentally acquired data, framed in mechanistic models whose validity is widely confirmed and which contribute to an understanding of the pathophysiological processes. The goal was to provide a model of study of a complex subject, which is also applicable to other disciplines that the student will meet in the course of his medical studies. The volume has been divided into blocks of teaching material: • text and figures - essential subjects; • in-depth analysis with figures - supplementary topics; • areas of clinical histology - physiopathological correlations; • extra content in the Virtual Campus web site - additional material available in multimedia format. Each chapter contains a paragraph concerning the Prerequisites, that is the basic knowledge necessary for a correct classification of the topics covered, a paragraph concerning the Objectives that the study of the topic is aimed at and a Summary of the topic dealt with and concludes with Concepts key to the topics covered. The topics have been developed highlighting the differentiation mechanisms that allow, in both embryonic and adult life, the exchange and integration of stem cell elements in the context of functional specialized populations. Particular attention was paid to the biomechanical properties of tissues and to the properties of the different populations of adult stem cells, also in consideration of their increasing use, also in association with biomaterials, in regenerative medicine.

Tooth replantation, as a treatment concept, has been subject to controversies regarding the mechanism as well as the various parameters underlying this process. This work aimed to study time-related changes in the pulp of replanted mature human premolars through the changes in the levels of certain factors involved in the underlying mechanisms of pulpal tissue healing after replantation. Eleven experimental mature teeth were extracted, immediately replanted in the original socket and left without any other intervention for 1, 2, 3 and 12 weeks before re-extraction. Three premolars served as control. All specimens were subject to histological analysis and the levels of MMP-2, MMP-9, Annexin V, iNOS and BCL-2 (anti-apoptotic family) were analyzed employing immunohistochemistry. The results showed degradation of the extracellular matrix (ECM), inflammatory cell infiltrate, loss in pulpo-dentine interface and loss of odontoblasts in the dental pulp tissue. This was accompanied by increase over time of MMP-9, Annexin V, iNOS and a decrease of BCL-2 and MMP-2, suggesting that apoptosis increased throughout the experimental period.

This research has the objective to investigate immunohistochemical expression of CB1 receptor and its probable changes in Gastroenteropancreatic system (GEP) of obese and lean Zucker rats and understand the endocannabinoid pathophysiological implications in the obesity. Male obese (fa/fa) and lean Zucker rats 6 weeks old were obtained from Harlan Italy Srl; the rats were sacrificed at 8, 12 and 16 weeks old. Normal rats also were sacrificed. Specimens of stomach, jejunum-ileum and pancreas were fixed in Bouin’s mixture and embedded in paraffin; obtained sections were processed with anti-CB1 (Biosource Europe SA) by Streptavidin-Biotin-Complex Method. The findings show that CB1 receptor is expressed not only in enteric neurons as documented by earlier studies up to now but more widely and with stronger intensity in obese animals compared with their lean counterparts by several structures of gastrointestinal tract (epithelium, glands, endocrine cells and immune cells of villi stroma). In obese Zucker rats pancreas unlike the normal rats where the CB1 receptor is essentially expressed by A-cells, the CB1 immunoreactivity even extends with higher intensity to B-cells. It is concluded that GEP System represents a new and wide peripheral target of EC action that have a direct autocrine, endocrine-paracrine and neurocrine control on many functions of GEP. In addition in GEP of obese Zucker rats in comparison with lean ones, the CB1 receptor is overexpressed and consequently the peripheral endocannabinoid system is upregulated and negatively modulated by leptin. It may contribute to increase hyperphagia, body weight and hyperglycaemia.

Objective. This study seeks to clarify the histological origin of apical lesions, through the expression of certain cytokeratins.Study design. In 3 years 30 patients were selected. After clinical and radiological exams were chosen patients who had severe apical lesions and tooth decay with exposure of the pulp chamber left untreated for very long periods (more the 12 months). Serial slides were prepared both for immunohistochemistry and Hematoxylin- Eosin. Material and methods. Twenty specimens coming from the 30 patients were used for our purpose. All periapical lesions were surgically extracted and fixed in Bouin mixture, and embedded in paraffin. Samples were processed immunohistochemically employing the instructions of the Envision Dako cytomation kit. Monoclonal antibobies against Cytokeratins (CKs) 1, 5 , 8, 10 and 14 were used. Slides were observed with Leica Laborlux-S microscope and the image were acquired with NIKON DSL2 System. Each sample has been analyzed with a “double-blind” system by two different operators. Moreover, the digital images acquired have been compared to an image analysis tools by Adobe photoshop CS5 Extended. Results. CKs 1, 5 and 8 are expressed in all samples at external epithelial layer with no statistically significant differences. CK 10 is weakly expressed in some epithelial cells lining the periapical lesion. CK 14 was negative in most samples, and little positive in some specimens. Conclusion. Our results suggest that most of the apical lesions we studied have an epithelial origin. The absence of CK14 positivity may be explained by the fact that during the extirpation of the lesion the basal epithelium remained attached to the surrounding bone., Italian Journal of Anatomy and Embryology, Vol 116, No 1 (Supplement) 2011

The endocannabinoid system plays a key role in energy homeostasis, with agonists and antagonists of CB1 receptors acting centrally to stimulate and inhibit food intake, respectively. The currently available literature, about islets cannabinoid receptors expression and function, is confusing and often contradictory. Aim of our study is compare the expression of CB1 receptors in normal and obese Zucker (ZDF) rats.12 Male ZDF (6 lean and 6 fa/fa) aged 8 weeks were obtained from Harlan Italy S.r.l. Each rat was fed with standard diet and unlimited water. Specimens were taken at the age of 8-12-16 weeks after birth. Pancreas samples were fixed in formaline and embedded in paraffin. Section were processed immunohistochemically using DakoCytomation EnVision kit for anti CB1antibody (Biosource Europe S.A.). Leica microscope, with a DSL2 Nikon was used to observe in double blind. Images were submitted at the image analysis tools of the Photoshop CS5Ex. At the same time pancreas specimens were frozen in liquid nitrogen and stored at -80°C until use. A mRNA was extracted for all samples and a RT PCR was performed. The pancreatic islets cells in fa/fa show deep apoptotic alterations. Our observations show a CB1 expression extended to most of the cell population (eighth week), up to a structural disorganization in the sixteenth week; the expression of CB1 receptor is upregulated in fa/fa compared to lean. These results according to the involvement of endocannabinoid system in the disregulation of the food intake mechanism and a prevalence of this system in the obesity., Italian Journal of Anatomy and Embryology, Vol 116, No 1 (Supplement) 2011

Recent studies document that CB1 cannabinoid receptors, considered since its first identification as “brain specific”, could be expressed by peripheral tissues targets (adipose organs, enteric nervous system, striated muscle, epatocytes). This receptors were found in the GI tract of different species, including mice, rats, guinea pigs, pigs and humans. Endocannabinoids play a role in the pathophysiology of the gastrointestinal tract. They play a role in emesis, acid-related disorders, motility-related disorders, irritable bowel syndrome and diarrhea, and in inflammatory bowel disease. Endocannabinoid system might be involved in cancer differentiation, growth and cell migration. Distribution of receptors in different colonic segment is not known. We studied CB1 receptors distribution in 10 patients colonic segments; the present report shows preliminary results. Fragments of sigma, transverse and ascending human colon were obtained during endoscopic screening procedure. Materials and Methods The specimens, for immunohistochemistry (IHC), were fixed in buffered formalin and embedded in paraffin; obtained sections were processed with anti CB1 (Biosource Europe S.A.) by EnVision+System HRP (AEC) and have been studied with photomicroscope Leica DMLB. The specimens for the Reverse Transcription Polymerase Chain Reaction (RT-PCR) were frozen in liquid nitrogen and stored at -80° until use. Total RNA extraction was performed using the “illustra RNA spin Mini Kit”. RT reaction was performed using the “Enhanced avian HS RT PCR kit” The PCR was performed using the “PCR enzyme Selection Kit-High specificity”. Results and Discussion Following the endoscopic procedure was performed anatomopathological examination of specimens. The results have revealed that four of the 10 patients had a polyp the others were not suffering from any disease of the colon. Our immunohistochemical evaluation has revealed the presence of CB1 receptors expressed by regional differences in patients who had a polyp. This reactivity appeared to be lower than that shown in non-pathological patients. The PCR results instead of doubtful interpretation. In fact, despite being detected in some cases also marked a positive control, in two cases, the PCR is a totally negative compared with a positive IHC. Since the number of cases studied is not statistically significant, it is our intention to continue to evaluate additional patients both normal and suffering from polyposis in order to clarify the difference beetween immunohistochemical and molecular biology results.

The oral cavity is exposed to chronic or recurrent, physical and chemical trauma that could lead to mucosal reactions (e.g. hyperplasia, dysplasia and tumors). The objective of this study is to investigate the expression and the possible changes of the two matrix metalloproteinases MMP-2 and MMP-9 in normal and pathological human oral mucosa samples. Normal oral mucosa samples and three different types of pathological conditions (hyperplasia, dysplasia and carcinoma) were used for this study. Immunohistochemical analysis was used to evaluate protein expression for the two enzymes, while Reverse Transcription ? Polymerase Chain Reaction (RT-PCR) was used to investigate gene expression. Image analysis was used to give a quantitative evaluation of the immunohistochemical data. In control samples we identified a weak expression of both MMP-2 and MMP-9 in the epithelial layers. In hyperplasia samples MMPs expression is limited to epithelial layers but the immunoreactivity is more intense than in the control. In dysplasia and carcinoma samples the two matrix metalloproteases are expressed not only in epithelium but also in some cells of the connective tissue and in the vessel walls. Qualitative RT-PCR and image analysis confirmed the immunohistochemical data. The results obtained in this study suggest the existence of a possible relationship between the entity of morphological disorganization of the oral mucosa in different pathologies and the increase of MMP-2 and MMP-9 expression

The use of methods of image analysis has allowed us to make more reliable and reproducible results obtained by immunohistochemistry (IHC). Wider use and simplification of software allowing a colorimetric study has meant that these methods are available to everyone and made it possible to standardize the technique by reliable systems score. Moreover, the introduction in recent years of multispectral image acquisition systems methods has further refined these techniques allowing to avoid artifacts and release the evaluation of the data by the observer.

Introduction. The oral cavity contains many structures compressed in a small area. It is exposed to chronic or recurrent, physical and chemical trauma leading to mucosal reactions (e.g. hyperplasia, dysplasia and tumors). For a long time, our group has been studying the expression of some metalloproteinases (MMP-2, MMP-9) in healthy and pathological human oral mucosa through immunohistochemistry and RT-PCR to investigate the entity of the remodelling process in the different cases. Matherials and Methods. In this paper we investigate a sample of normal oral mucosa and four pathological samples (mucocele, fibroma, dysplasia and carcinoma). We identified the localization of MMP-2 and MMP-9 through immunohistochemistry and gene expression through RT-PCR analysis. Results. In control sample we identified a weak expression of both metalloproteinases (MMP-2, MMP-9) in the upper epithelial layers. The reactivity of the two enzymes is emphasized with higher intensity in pathological samples where a strong inflammatory activity is present (e.g. hyperplasia, dysplasia) or where there is a distruption of the architecture (as in tumors) while in localized lesions a functional silence is detected (mucocele). Particularly, in lesions like hyperplasia and dysplasia where (though it is present a inflammatory condition) the bio-architecture is preserved, the immunoreactivity is expressed only in the upper epithelial cell layers; while in more serious lesions (like tumors), where the architecture is fully disorganizated, the immunoreactivity is expressed also in deeper epithelial layers, in few connnective cells and in vessels wall. Qualitative RT-PCR analysis confirmed the expression of MMP-2 and MMP-9 in both normal and pathological oral mucosa samples. Conclusions. Our data suggest that there is a relationship between the increase of MMP-2 and MMP-9 expression and the entity of morphological disorganization according with the well known role of these enzymes.

We have used 12 Epulis taken during surgical operation and as control, some normal gingival specimen. The aim was to clarify the relationship between the degree of inflammation and NO production in several cases of Epulis, exhibiting different degree of inflammatory process and verifying the presence of cells involved in NO production. From the immuhistochemical data in our possession we can conclude that NO is one of the factor which module the pathogenecity degree of Epulis.

Aortic aneurysms (AA) is a degenerative vascular disease characterized by localized dilatation of the aortic wall as a result of altered matrix composition (elastin and collagen degradation). Howewer the pathogenesis of the changes is elusive and unclear. Some experimental evidences suggest that iNOS (who synthesize a large amount of NO in inflammatory processes) and the metalloproteinases (MMP) are implicated in the pathogenesis of AA but the relationship between NO and MMP to aneurismal disease is currently unknown. The aim of this study is to investigate the immunohistochemical expression of iNOS and MMP2 and MMP9 in human aneurysmal tissues.

The aim of present study is to investigate the immunohistochemical identification of cNOS-containing prostatic structures in various species and in hyperplastic human prostate. We used specimens of rat, guinea pig and hamster. Human prostatic specimens were removed from patients suffering of prostatic hyperplasia. The study was processed by ABC method. Immunohistochemical staining with anti nNOS antibody revealed the existence of strong reactivity in numerous ganglionic cell bodies and varicose nerve fibers in the fibromuscular stroma, in smooth cells around acini and in blood vessels endothelium. Surprisingly nNOS immunoreactivity appears in the glandular epithelium. In the hyperplastic prostate the nitroxidergic innervation of stroma and glandular epithelium is almost absent. These data provide a morphological evidence for a segmental differentiation of NOS-containing prostatic structures and consequently suggest that in the prostate NO plays selective neural and paraneural roles in the control of blood flow, smooth-muscle tone (relaxation) and secretory function. Further immunohistochemical studies are carrying out to elucidate the prostatic role in aging, after hormonal perturbation and pathological processes.

Heat shock proteins (Hsps) are an important class of molecules with various functions. Their classic role is to assist other proteins in folding and re-folding and, when proteins are defective or irreversibly misfolded, to drive their degradation. For this reason, some Hsps are also named molecular chaperones. During evolution, this class of proteins has also acquired extrachaperoning roles such as participation in immune system regulation, cell differentiation, programmed cell death and carcinogenesis. Hsp10 is a partner of Hsp60 in the Hsp60/10 folding machine, but numerous scientific studies have shown that Hsp10 may also play other roles. In fact, Hsp10 seems to have an immunomodulatory activity and a role in tumor progression. Hsp90 regulates late-stage maturation, activation and stability of a range of client proteins, such as HER2, EGFR and BRAF, some of which are involved in signal transduction and other key pathways important for malignancy in several cancers, including large bowel carcinomas. The aim of the present study was to evaluate levels and expression of Hsp10 and Hsp90 in a series of samples of large bowel mucosa obtained from healthy controls and patients with adenocarcinomas.

IntroductionThe World Health Organization declared the coronavirus disease 2019 (COVID-19) pandemic on March 11, 2020. Two vaccine types were developed using two different technologies: viral vectors and mRNA. Thrombosis is one of the most severe and atypical adverse effects of vaccines. This study aimed to analyze published cases of thrombosis after COVID-19 vaccinations to identify patients’ features, potential pathophysiological mechanisms, timing of appearance of the adverse events, and other critical issues.Materials and MethodsWe performed a systematic electronic search of scientific articles regarding COVID-19 vaccine-related thrombosis and its complications on the PubMed (MEDLINE) database and through manual searches. We selected 10 out of 50 articles from February 1 to May 5, 2021 and performed a descriptive analysis of the adverse events caused by the mRNA-based Pfizer and Moderna vaccines and the adenovirus-based AstraZeneca vaccine.ResultsIn the articles on the Pfizer and Moderna vaccines, the sample consisted of three male patients with age heterogeneity. The time from vaccination to admission was ≤3 days in all cases; all patients presented signs of petechiae/purpura at admission, with a low platelet count. In the studies on the AstraZeneca vaccine, the sample consisted of 58 individuals with a high age heterogeneity and a high female prevalence. Symptoms appeared around the ninth day, and headache was the most common symptom. The platelet count was below the lower limit of the normal range. All patients except one were positive for PF4 antibodies. The cerebral venous sinus was the most affected site. Death was the most prevalent outcome in all studies, except for one study in which most of the patients remained alive.DiscussionVaccine-induced thrombotic thrombocytopenia (VITT) is an unknown nosological phenomenon secondary to inoculation with the COVID-19 vaccine. Several hypotheses have been formulated regarding its physiopathological mechanism. Recent studies have assumed a mechanism that is assimilable to heparin-induced thrombocytopenia, with protagonist antibodies against the PF4–polyanion complex. Viral DNA has a negative charge and can bind to PF4, causing VITT. New experimental studies have assumed that thrombosis is related to a soluble adenoviral protein spike variant, originating from splicing events, which cause important endothelial inflammatory events, and binding to endothelial cells expressing ACE2.ConclusionFurther studies are needed to better identify VITT’s pathophysiological mechanisms and genetic, demographic, or clinical predisposition of high-risk patients, to investigate the correlation of VITT with the different vaccine types, and to test the significance of the findings.