Anti-alpha Smooth Muscle Actin Research Articles

Histochemical analysis, smooth muscle immunolocalization and volumetric density of the elastic system fibres of the ostrich (Struthio camelus) phallus.

There are few scientific reports on the histology of the phallus of ratite birds. The aim of this study was to conduct a histochemical analysis to determine the distribution of smooth muscle cells and the volumetric density (Vv) of the fibres of the elastic system in the ostrich phallus. Adult ostriches, 14 months old, were used. The phalluses were fixed in Bouin's solution and then transferred to a buffered formalin solution. They were then processed using standard histological stains for paraffin and slices were obtained. The following techniques were performed: HE, Picrosirius red, Alcian Blue at pH 1.0 and 2.5. The Periodic acid-Schiff reagent and Weigert's Resorcin-Fuchsin with previous oxidation were performed. The M42 test system was used to quantify the elastic system fibres. For immunohistochemical analysis, an anti alpha smooth muscle actin monoclonal antibody was used. The surface of the phallus is covered by a non-keratinized stratified squamous epithelium, which becomes stratified cylindrical in the region of the spermatic sulcus. No glands associated with the connective tissue were observed. The Vv of the elastic system fibres was 4.75%. Smooth muscle cells were visualized only in the walls of blood vessels through immunostaining, with an absence in the lymphatic sinuses. Despite similarities with other birds, such as the presence of a fibrous external axis, a lymphatic core, and a spermatic groove, the ostrich phallus shows marked differences, including the absence of an elastic core, a non-keratinized lining epithelium, and the absence of glands throughout its extension.

Effects of Sprayable, Highly Adhesive Hydrophobized Gelatin Microparticles on Endoscopic Submucosal Dissection: A Swine Model

Introduction: Sprayable wound dressings containing hydrophobized microparticles (hMPs) are characterized by strong adhesiveness. We examined the effect of hMPs derived from Alaska pollock gelatin on endoscopic submucosal dissection (ESD) ulcers. Methods: (1) In an in vivo model of miniature swine gastric ESD, gastric ulcers were created by ESD and then sprayed with hMPs or untreated followed by microscopic examination. (2) In an ex vivo ESD model of resected stomach, a pinhole-shaped perforation was created on the ESD ulcer of resected stomach; hMPs were then sprayed on the perforation; and air leakage and intragastric pressure were measured. (3) In an in vivo duodenal ESD model of miniature swine, duodenal artificial ESD ulcers with pinhole-shaped perforation were examined; ulcers were classified into hMPs-sprayed and nonsprayed groups, and inflammation in the intrinsic muscle layer and serosa were compared between the groups. Results: (1) Histological observation of submucosal tissues showed a decreased number of invading inflammatory cells in hMP-sprayed tissues compared with the control in miniature swine gastric ESD (p < 0.05). In addition, the rates of anti-alpha smooth muscle actin and type I collagen positivity were significantly lower in the hMPs group than in the control group (p < 0.05). (2) Intragastric pressure could not be measured in the nonsprayed group, whereas no air leakage was observed in the sprayed group when pressurized up to 26 mm Hg in the resected stomach model. (3) The sprayed group showed suppressed inflammation of the intrinsic muscular layer and serosa in both cases compared with the nonsprayed group in miniature swine duodenal ESD (p < 0.05). Conclusions: Sprayable, tissue-adhesive hMPs are a promising medical material for intraoperative and postoperative treatment of ESD-induced wound via anti-inflammation and strong adhesiveness.

A Preliminary Study on the Promotion of Canine Adipose-Derived Stem Cell Differentiation by Perfusion-Decellularized Ureter Matrix

This study was to assess the possibility of the perfusing decellularized ureters (DUs) promoting the differentiation of the canine adipose stem cell (cASCs). cASCs were isolated and cultured in different induction media to determine their multidirectional differentiation potential. The perfusion system was used to prepare the DUs, and the prepared DUs were systematically evaluated. The DU coating was prepared by enzymatic digestion for cell culture. The cASCs were seeded on the coverslips covered with DU coating and samples were collected on days 3, 7, and 10. Immunofluorescence staining and molecular biology testing were used to examine the differentiation of cASCs seeded on the DU coating. The cASCs were isolated and identified by flow cytometry. The prepared DUs removed the nuclear materials, and the 3-dimensional structure and biological compositions of the ureter were well preserved. Immunofluorescence staining showed the expression of anti-alpha smooth muscle actin (α-SMA). Western blot results suggested that the content of α-SMA in the experimental group was significantly higher than that in the control group at 3 different time points, and the mRNA expressions of α-SMA in the experimental group gradually increased with extended the culture time, whereas there was no significant change in the control group. The cASCs seeded on the coverslips of DU coating could differentiate into smooth muscle cells, and the number of differentiated cASCs increased significantly with extended incubation time.

Primary Splenic Pleomorphic Liposarcoma in a Bitch

Background: Liposarcoma is a malignant neoplasm of lipoblasts with low incidence in dogs, representing 1.7% of neoplasms diagnosed in the spleen. In veterinary medicine, this neoplasm is classified morphologically into the myxoid, well-differentiated, undifferentiated and pleomorphic subtypes, the latter being one of the most aggressive forms, mainly in cavity organs. This study reports a case of primary splenic pleomorphic liposarcoma in a female dog, addressing anatomopathological and immunohistochemical aspects.Case: A 14-year-old, 35 kg female mongrel canine with a history of absence of defecation, progressive weight loss, difficulty walking, sensitivity to abdominal palpation, prostration, pale mucous membranes, tachypnea and abdominal distention. The condition evolved to death and, on necroscopy, there was an increase in splenic volume with neoformation of whitish and reddish color, measuring 32 × 27 cm in its largest axes and weighing 8.9 kg. The neoformation exhibited areas of firm and soft consistency, and sectioning revealed the existence of focal areas of extensive necrosis and cavity collections of different diameters that allowed the flow of liquid serous contents with a brownish red color. Microscopy showed cells of neoplastic morphology infiltrating the splenic parenchyma, mostly with slightly acidophilic cytoplasm and few intracytoplasmic lipid vacuoles, which varied in size and distribution. The nuclei of the cells were large, eccentric and irregular, with round to oval morphology, grossly lacy chromatin and single or multiple evident nucleoli. These cells exhibited marked anisocytosis, anisokaryosis and pleomorphism, with more than one mitotic figure per high magnification field visible. Moderately inflammatory infiltrate, predominantly lymphocytic, permeated the neoplastic cells, and marked depletion of lymphoid follicles and atrophy of the red pulp were found in the remaining splenic parenchyma. Immunohistochemical tests revealed marked and discrete immunostaining for anti-vimentin and anti-S100 antibodies, respectively. No staining was observed for anti-pan cytokeratin, anti-desmin, anti-alpha smooth muscle actin or anti-CD20 antibodies. Based on anatomopathological and immunohistochemical aspects, it was concluded to be a splenic pleomorphic liposarcoma of primary origin.Discussion: the spleen is not a common anatomical site for the development of liposarcoma, a neoplasm whose origin remains unclear. Similar to what occurs in humans, liposarcoma is believed to develop from the adipose tissue of the splenic hilum. Thus, it should be considered as a differential diagnosis of invasive abdominal tumors. For the identification and classification of liposarcoma as a pleomorphic subtype, we considered mainly histological findings such as marked cell atypia and intracytoplasmic lipid vacuoles, which may or may not be present in neoplastic cells. Immunohistochemical examination favored the diagnosis of liposarcoma, regardless of the subtype, due to the marked immunostaining for the anti-vimentin antibody, unlike immunostaining for the anti-S100 antibody, for which it was variable. This fact is related to adipocyte differentiation, where lower amounts of intracytoplasmic lipids translate into lower immunostaining intensity for anti-S100. Histological and immunostaining aspects should be regarded with caution in the diagnosis of pleomorphic liposarcoma, as it is a distinct neoplastic entity, with a complex karyotype and without correlation with the other subtypes.

Immunohistological analysis on distribution of smooth muscle tissues in livers of various vertebrates with attention to different liver architectures

BackgroundThe liver architecture of vertebrates can be classified into two types, the portal triad type (having periportal bile ducts) and the non-portal triad type (having non-periportal bile ducts). The former is detectable from the hagfish, which is the most ancestral vertebrate, to tetrapod livers whereas many actinopterygian livers have the latter. The aim of the present study is to reveal the distribution of smooth muscle tissue in livers of various vertebrates with attention to their architectures. MethodsSmooth muscle was immunohistochemically compared in hepatic blood vessels and bile ducts of various vertebrates, using an anti-alpha-smooth muscle actin (ASMA) antibody. ResultsSmooth muscle was noted in the gallbladder and hepatic artery in all vertebrates, including the hagfish. Bile ducts having ASMA-positive smooth muscles were absent in the hagfish, but detected in the Chondrichthyes and conserved in actinopterygians with or without portal triads during the evolution of vertebrates. In tetrapods having portal triads, reptiles had a tendency to have strongly ASMA-positive biliary smooth muscle tissues whereas other tetrapods had bile ducts with poor smooth muscle tissues. Although the hagfish livers never had ASMA-positive smooth muscle tissue in the walls of portal and central veins, it was observed in discontinuous distributions or not observed in portal veins and central veins of chondrichthyans and actinopterygians. By contrast, in most tetrapods, ASMA-positive smooth muscle tissue was detectable in portal veins, which supported the adjacent endothelial cells as a circular layer. Central veins did not consistently have smooth muscle tissue in these groups. Discussion and conclusionThe hagfish liver may retain more ancestral characteristics than other vertebrates in terms of smooth muscle distribution in the vascular and biliary systems. Actinopterygians might have a different mechanism of bile transport from tetrapods from their smooth muscle distribution in intrahepatic bile ducts. The circular smooth muscle distribution in portal veins might be a characteristic acquired by tetrapods.

Measuring inflammation in patients with allergic rhinitis using different biomarkers

Allergic rhinitis is an inflammatory disease affecting the mucosal lining of the nose of individuals of all ages. The allergen in the air involves‏ attaching by Immunoglobulin E that causes the release of inflammatory chemicals from mast cells. This study aimed to measure the serum level of biomarkers and to determine the correlation between them in mediating activate cell in patients with AR. In this study, blood samples were taken from 88 AR patients and 88 healthy controls (HC) Symptom score was measured using the SNOT-22 questionnaire and blood sample was analyzed for eosinophil counts (B-Eos) using optical flow cytometry, IgE by electrochemiluminescence immunoassay, as well as eosinophilic cationic protein, anti-alpha smooth muscle actin (anti-SMA), cyclooxygenase- 2 (COX-2R), phosphatase and tensin (PTEN) homolog and Tryptase Alpha/Beta 1 (TPSABI) by ELISA. B-Eos, IgE, ECP, anti-SMA, and symptom scores were significantly higher in AR patients than in HC. As a result, we found that there was correlation between B-Eos and ECP (r = 0.66, p = 0.001). There was also a correlation between IgE and symptom score (r=0.64, p=0.01). COX-2 also correlated with symptoms score (r = 0.52, p = 0.04). There was a positive correlation between phosphatase and tensin homolog and TPSAB1 (r = 0.76, p&lt;0.001) and PTEN and ECP (r = 0.53, p = 0.03). In conclusion, we realized that in allergic rhinitis, the airway inflammation was characterized by high numbers of eosinophil, IgE, ECP and anti-SMA. B- Eos, IgE and COX-2 were related to symptom scores. The study highlights the importance of systemic inflammation in AR.

Advanced glycation end products are associated with immature angiogenesis and peritoneal dysfunction in patients on peritoneal dialysis.

Deposition of advanced glycation end products (AGEs) is frequently found in the peritoneum of patients on peritoneal dialysis (PD). Angiogenesis is also observed in the peritoneum. However, the clinical significance of AGEs and angiogenesis in the peritoneum is not fully understood. We evaluated the maturation of capillary vessels and investigated whether AGEs are associated with angiogenesis and peritoneal function in the peritoneal membrane. Peritoneum obtained when PD catheters were removed from 61 patients with PD was analyzed. The peritoneum was immunohistochemically stained with anti-CD34 (for endothelial cells), anti-alpha smooth muscle actin (αSMA) (for pericytes), and anti-AGE antibodies. We defined CD34-positive and αSMA-negative vessels as immature capillary vessels in peritoneal membranes using serial sections. We evaluated the associations between vessel density, peritoneal function (dialysate-to-plasma ratio for creatinine (D/P creatinine)), and the degree of AGE deposition. AGE accumulation in the interstitium was positively associated with the duration of PD (p < 0.01). AGE accumulation in the interstitium and vascular wall was positively correlated with the use of acidic solution (p < 0.05) and the maximum value of D/P creatinine (p < 0.05). AGE accumulation in the vascular wall was significantly associated with immature capillary density (CD34+/αSMA-) in the peritoneum (p < 0.01). Vessel density was not significantly correlated with the last measurement of D/P creatinine (p = 0.126, r = 0.202), However, immature capillary density was positively correlated with the last measurement of D/P creatinine (p < 0.05, r = 0.278). AGE accumulation is significantly associated with immature angiogenesis and peritoneal dysfunction in patients undergoing PD.

Establishment of tooth blood supply and innervation is developmentally regulated and takes place through differential patterning processes.

Teeth are richly supported by blood vessels and peripheral nerves. The aim of this study was to describe in detail the developmental time-course and localization of blood vessels during early tooth formation and to compare that to innervation, as well as to address the putative role of vascular endothelial growth factor (VEGF), which is an essential regulator of vasculature development, in this process. The localization of blood vessels and neurites was compared using double immunofluorescence staining on sections at consecutive stages of the embryonic (E) and postnatal (PN) mandibular first molar tooth germ (E11-PN7). Cellular mRNA expression domains of VEGF and its signaling receptor VEGFR2 were studied using sectional radioactive insitu hybridization. Expression of VEGF mRNA and the encoded protein were studied by RT-PCR and western blot analysis, respectively, in the cap and early bell stage tooth germs, respectively. VEGFR2 was immunolocalized on tooth tissue sections. Smooth muscle cells were investigated by anti-alpha smooth muscle actin (αSMA) antibodies. VEGF showed developmentally regulated epithelial and mesenchymal mRNA expression domains including the enamel knot signaling centers that correlated with the growth and navigation of the blood vessels expressing Vegfr2 and VEGFR2 to the dental papilla and enamel organ. Developing blood vessels were present in the jaw mesenchyme including the presumptive dental mesenchyme before the appearance of the epithelial dental placode and dental neurites. Similarly, formation of a blood vessel plexus around the bud stage tooth germ and ingrowth of vessels into dental papilla at E14 preceded ingrowth of neurites. Subsequently, pioneer blood vessels in the dental papilla started to receive smooth muscle coverage at the early embryonic bell stage. Establishment and patterning of the blood vessels and nerves during tooth formation are developmentally regulated, stepwise processes that likely involve differential patterning mechanisms. Development of tooth vascular supply is proposed to be regulated by local, tooth-specific regulation by epithelial-mesenchymal tissue interactions and involving tooth target expressed VEGF signaling. Further investigations on tooth vascular development by local VEGF signaling, as well as how tooth innervation and development of blood vessels are integrated with advancing tooth organ formation by local signaling mechanisms, are warranted.

Diagnostic Criteria of Pediatric Intestinal Myopathies.

The authors aim to identify criteria for the diagnosis of intestinal visceral myopathy (IVM); results were compared with ultrastructural studies. Six IVM patients and 7 pediatric control cases (without gastrointestinal diseases) were studied. One case was a typical megacystis-microcolon-intestinal hypoperistalsis syndrome. The diagnostic path included: rectal suction biopsy, one-trocar transumbilical laparoscopic intestinal full-thickness biopsy technique. Pathological analysis included anti-alpha smooth muscle actin staining, and US study of intestinal biopsies. IVM histological examination demonstrated thinning of longitudinal muscle layer. The ratio of circular/longitudinal thickness was evaluated in all samples; in cases, this ratio presented as a mean value of 2.91, and in controls, a mean value of 1.472 (P = 0.0002). Ultrastructural diagnosis revealed variable myofibrils density in smooth muscle cells, irregularity of sarcolemma membranes, interstitial fibrosis, and myofiber disarray. The authors concluded that in IVM, circular/longitudinal thickness ratio and alpha smooth muscle actin staining can be used as significant tools to address the diagnosis.

CAN OCIMUM BASILICUM (BASIL) RELIEF THE IMPACT OF CHRONIC UNPREDICTABLE MILD STRESS ON THE MICE SALIVARY GLANDS?

Background: Salivary glands are affected in acute and chronic stressful conditions. Ocimum basilicum (OB), basil,&#x0D; possesses anxiolytic and antidepressant like effect so this study aimed to evaluate the effect of the chronic unpredictable&#x0D; mild stress (CUMS) on the structure of salivary glands of mice and the efficacy of OB in relieving this effect.&#x0D; Material and Methods: Forty male mice were distributed equally into four groups; the control, CUMS (exposed to the&#x0D; CUMS for 4 weeks), CUMS+Fluoxetine (treated with Fluoxetine after exposure to CUMS), CUMS+OB (treated with OB&#x0D; after exposure to CUMS). Treatments continued for 2 weeks. Behavioral changes and serum corticosterone level were&#x0D; assessed at the end of the experiment. Submandibular and parotid glands were histopathologically examined and stained&#x0D; with anti-alpha smooth muscle actin (ASMA) and anti-brain derived neurotropic factor (BDNF) antibodies.&#x0D; Results: A depressive behavior was observed in mice exposed to the CUMS. Serum corticosterone level significantly&#x0D; increased in these mice compared to the control (130.9±8.8 versus 21.03±2.1, p

Therapeutic effect of Aloe vera and silver nanoparticles on acid-induced oral ulcer in gamma-irradiated mice.

Radiation combined injury, a life-threatening condition, has higher mortality than simple radiation injury. The aim of the present study was to analyze the efficiency of Aloe vera and silver nanoparticles in improving the healing of ulcerated oral mucosa after irradiation. Thirty male Albino mice were divided into five groups: control, radiation, Aloe vera (AV), silver nanoparticles (NS), and AV+NS. The mice were exposed to whole body 6Gy gamma-radiation. After one hour, 20% acetic acid was injected into the submucosal layer of the lower lip for ulcer induction. The animals received topical treatment with the assigned substances for 5 days. Lip specimens were subjected to hematoxylin and eosin and anti alpha-smooth muscle actin immunohistochemical staining. Results demonstrated occurance of ulcer three days post irradiation in all groups except in the AV+NS group where only epithelial detachment was developed. After seven days, data revealed persistent ulcer in radiation group, and almost normal epithelium in the AV+NS group. A significant reduction of epithelial thickness was detected in all groups at the third day as compared to control. At the seventh day, only the AV+NS group restored the epithelial thickness. Area percent of alpha-smooth muscle actin expression was significantly decreased in radiation group at the third day followed by significant increase at the seventh day. However, all treatment groups showed significant increase in alpha-smooth muscle actin at the third day, which decreased to normal level at the seventh day. Our study demonstrated the efficiency of Aloe vera and silver nanoparticles in enhancing ulcer healing after irradiation.

The 50/10 Oxygen-Induced Retinopathy Model Serves as a Hyperoxia and Hypoxia Model of Bronchopulmonary Dysplasia

BackgroundAnimal models of bronchopulmonary dysplasia (BPD) are mainly created by hyperoxia exposure. However, these models do not fully recapitulate BPD pathophysiology as observed in clinical practice. To find a better BPD model, we established a rat 50/10 oxygen-induced retinopathy (OIR) model and analyzed the pathologic features of the lungs. MethodsThe rat OIR model was established by exposing newborn rats (P0) to 50% and 10% oxygen (hyperoxia and hypoxia) on alternating days for 14 days. Lungs were harvested immediately on postnatal day 14 (P14) and on P18 after 4 days of normoxia exposure for hematoxylin and eosin staining, antialpha smooth muscle actin (α-SMA) immunohistochemistry and Picrosirius red staining of collagen. Retinas were obtained to confirm successful model establishment by isolectin B4 staining of retinal vasculature. ResultsOIR rats presented with fewer and enlarged alveoli, and the septal walls were thicker than those in age-matched controls. α-SMA immunohistochemistry indicated increased abundance of myofibroblasts in OIR rats. At P18, α-SMA-positive myofibroblasts were present at extremely low levels from the alveolar walls of control rats, while OIR rats showed myofibroblast persistence. The amount of collagen in OIR rats was also higher than that in control rats at both P14 and P18 as evidenced by Picrosirius red staining. ConclusionsAlveolar changes observed by hematoxylin and eosin staining, prolonged and stronger α-SMA expression and augmented collagen accumulation resemble the histopathology of BPD, suggesting that the rat 50/10 OIR model is suitable for use in BPD research.

PD49-07 ENGINEERING OF CORPORAL TISSUE CONSTRUCTS USING NON-HUMAN PRIMATE CORPUS CAVERNOSAL SMOOTH MUSCLE AND ENDOTHELIAL CELLS FOR CLINICAL APPLICATIONS

You have accessJournal of UrologySexual Function/Dysfunction: Penis/Testis/Urethra: Benign Disease & Malignant Disease I1 Apr 2017PD49-07 ENGINEERING OF CORPORAL TISSUE CONSTRUCTS USING NON-HUMAN PRIMATE CORPUS CAVERNOSAL SMOOTH MUSCLE AND ENDOTHELIAL CELLS FOR CLINICAL APPLICATIONS Joao Zambon, Young-Min Ju, Koudy Williams, Ryan Terlecki, SIta Somara, Alexander Baume, Ashley Dean, John Jackson, Julie Allickson, James Yoo, and Anthony Atala Joao ZambonJoao Zambon More articles by this author , Young-Min JuYoung-Min Ju More articles by this author , Koudy WilliamsKoudy Williams More articles by this author , Ryan TerleckiRyan Terlecki More articles by this author , SIta SomaraSIta Somara More articles by this author , Alexander BaumeAlexander Baume More articles by this author , Ashley DeanAshley Dean More articles by this author , John JacksonJohn Jackson More articles by this author , Julie AllicksonJulie Allickson More articles by this author , James YooJames Yoo More articles by this author , and Anthony AtalaAnthony Atala More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2017.02.2231AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Numerous conditions exist, both congenital and acquired, that threaten male sexual health through changes in form and/or function. Efforts in penile reconstruction are often limited by poor availability of functionally intact penile tissue. We have previously demonstrated the ability to reconstruct functional corporal tissue via autologous cell-seeded collagen matrix in a rabbit model. In this study, we investigated the feasibility of engineering corporal constructs using cavernosal smooth muscle (SMCs) and endothelial cells (ECs) from non-human primates (NHPs) seeded onto 3D acellular corporal collagen matrices. METHODS Corpora cavernosa were isolated from NHPs. These specimens were then subjected to an established decellularization process to create acellular corporal collagen matrices. Autologous corporal SMCs and ECs were isolated, expanded in vitro, and seeded onto matrices via a multistep static/dynamic procedure. RESULTS Histologic and immunohistochemical analyses were performed. The corporal construct treated with the TritronX-100 protocol was effectively decellularized based on results of both DAPI (4,6 diamidino 2 phenylindole) staining and DNA assay (< 50 ng dsDNA/mg dry sample weight). Scanning electron microscopy demonstrated highly porous 3D structure and structural integrity. Evenly distributed cellular attachment and phenotype of corporal ECs and SMCs before/after dynamic culture conditioning were evaluated by immunohistochemical staining with anti-von Willebrand factor and anti-alpha smooth muscle actin. CONCLUSIONS We have demonstrated the feasibility of engineering viable and well-organized corpora cavernosa using tissue from NHPs. This represents an important achievement toward clinical utility for humans. Such technology may have application for congenital anomalies, penile cancer, traumatic penile injury, and selected cases of erectile dysfunction and Peyronies disease. © 2017FiguresReferencesRelatedDetails Volume 197Issue 4SApril 2017Page: e976 Advertisement Copyright & Permissions© 2017MetricsAuthor Information Joao Zambon More articles by this author Young-Min Ju More articles by this author Koudy Williams More articles by this author Ryan Terlecki More articles by this author SIta Somara More articles by this author Alexander Baume More articles by this author Ashley Dean More articles by this author John Jackson More articles by this author Julie Allickson More articles by this author James Yoo More articles by this author Anthony Atala More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...

The influence of nicotine in healing of small bowel anastomoses in rats: angiogenesis and miofibroblasts.

to know the effect of nicotine on angiogenesis and myofibroblast formation in anastomoses of the small bowel of rats. we randomly divided 60 Wistar rats into the groups Nicotine (N) and control (C), according to the proposed treatment. Each group was subdivided into three subgroups according to the time interval used for the evaluation (7, 14 or 28 days). The N group with 30 animals received nicotine subcutaneously at a dose of 2mg/kg body weight, diluted in 0.3ml of 0.9% saline, twice daily for 28 days prior to the operation, and for more 7, 14 or 28 days, depending on the subgroup. The C group (also 30 animals) received only saline on the same conditions and time intervals. After 28 days we carried out an end-to-end anastomosis 10cm distal to the duodenojejunal flexure in each rat. After 7, 14 or 28 days after surgery, we euthanized ten animals of each group, sent specimens of the anastomosis areas, 1cm proximal to 1cm distal, to counting of blood vessels and myofibroblasts through immunohistochemical staining by the application of monoclonal anti-factor VIII antibodies and anti-smooth muscle alpha-actin. the administration of nicotine led to the decrease in the number of blood vessels measured on the 28th postoperative day and the number of myofibroblasts measured on the seventh day following completion of the anastomoses. administration of nicotine was deleterious on angiogenesis and myofibroblast formation in rats' small intestine anastomoses. conhecer o efeito da nicotina sobre a angiogênese e formação de miofibroblastos em anastomoses do intestino delgado de ratos. sessenta ratos Wistar foram divididos de maneira aleatória em grupos Nicotina(N) e Controle (C), conforme o tratamento proposto. Cada grupo foi subdividido em três subgrupos, de acordo com o intervalo de tempo utilizado para a avaliação (7, 14 ou 28 dias). O grupo N, com 30 animais, recebeu nicotina por via subcutânea, na dose de 2mg/Kg de peso, diluída em 0,3ml de solução salina a 0,9%, em duas aplicações diárias, durante 28 dias prévios à operação e por mais 7, 14 ou 28 dias, conforme o subgrupo. O grupo C (igualmente com 30 animais) recebeu somente a solução salina nas mesmas condições e intervalos de tempo. Após 28 dias efetuou-se, em cada rato, anastomose término-terminal a 10cm da flexura duodenojejunal. Após 7, 14 ou 28 dias da cirurgia, os dez animais de cada subgrupo foram eutanasiados, sendo que as áreas anastomosadas, 1cm proximal a 1cm distal, foram encaminhadas para contagem de vasos sanguíneos e miofibroblastos, através de coloração imuno-histoquímica por aplicação dos anticorpos monoclonais antifator VIII e anti-alfa-actina muscular lisa. a administração de nicotina levou à diminuição do número de vasos sanguíneos aferidos no 28o dia pós-operatório e do número de miofibroblastos aferidos no sétimo dia após a realização das anastomoses. a administração de nicotina foi deletéria sobre a angiogênese e formação de miofibroblastos em anastomoses do intestino delgado de ratos.

PTH-102 Stat2 is a key inflammatory molecule in human non-alcoholic fatty liver disease and murine liver injury

<h3>Introduction</h3> In patients with non-alcoholic fatty liver disease (NAFLD), liver inflammation is a key histological feature that distinguishes benign simple steatosis from progressive non-alcoholic steatohepatitis (NASH). The mechanisms that underlie this process are poorly understood. We recently showed that <i>Stat2</i>plays a pivotal role in acute inflammation, independent of its role as an interferon mediator. The aim of the current study is to test the hypothesis that STAT2 is associated with hepatic inflammation in patients with NASH and in a murine model of liver injury. <h3>Method</h3> Expression of (phosphorylated, p) STAT2 and STAT1 was assessed by immunohistochemistry in anonymised sections of archived liver tissue taken from consenting patients with NASH at The Royal London and St Mary’s Hospitals (London, UK). STAT2^-/-and WT mice were injected with incremental doses of carbon tetrachloride (CCl4) for 4 or 6 weeks. Serum and liver tissue were harvested 24 or 72hrs after the last injection. Immunohistochemistry for activated stellate cells (anti-alpha smooth muscle actin, aSMA) and macrophage/Kupfer cells (anti-F4/80) was performed. <h3>Results</h3> Tissue from 62 patients was examined. Hepatocyte nuclear expression in NASH liver tissue was significantly greater for STAT1 (74% vs 10%, p &lt; 0.0001) and STAT2 (98% vs 35%, p &lt; 0.0001) compared to normal liver. There was a significant correlation between STAT1 expression (r = 0.6, p = 0.02), but not STAT2 and the NAFLD activity score (NAS). Animal models confirm that Stat 2 plays a role in liver inflammation – <i>Stat2^-/-</i>mice were protected from CCl₄-mediated liver injury with reduced serum alanine aminotransferase (ALT, 2165 vs 1090 iU/ml, p = 0.03), inflammation and necrosis histologically and fibrosis (Picrosirius red staining). There were no significant differences in the number of aSMA or F4/80-positive cells between WT and <i>Stat2^-/-</i>mice. <h3>Conclusion</h3> These data identify a novel role for STAT2 in the development of liver disease, with expression of STAT1 and STAT2 relating to disease grade. The protection afforded by Stat2 loss in murine models is unrelated to macrophage and stellate cell activation, and we hypothesise that the mechanism of protection is therefore reduced production of Stat2-mediated cytokine production. <h3>Disclosure of interest</h3> None Declared.

AB0206 Altered Expression of Angiopoietins and Tie-2 in Ischemic Lesions of UCD-206 Chickens, an Animal Model for Systemic Sclerosis

BackgroundSystemic sclerosis (SSc) is an autoimmune connective tissue disease affecting skin and internal organs. It is characterized by vascular damage, inflammation, and fibrosis. Microvascular injury and insufficient angiogenesis lead to...

Development of biologic scaffolds for reconstructive surgery from decellularized human skeletal muscle

The development of engineered skeletal muscle tissues has recently been proposed as a potential solution for replacement of volumetric muscle losses. Experimental studies available in literature have mainly been performed in animals. The aim of the present work was to evaluate the possibility of obtaining homologous acellular scaffolds from decellularization of human skeletal muscle, also through a comparison with rats and rabbits, and evaluating the capability of muscle scaffolds to repair defects of the abdominal wall. Muscle samples of rats, rabbits and humans were treated with two different decellularization methods. Surgical tests were also performed in rabbits, implanting the decellularized scaffold in the abdominal wall of animal with surgical defects of the abdominal wall. Human skeletal muscles were sampled from body parts removed during surgery and given to the Body Donation Program of the Institute of Human Anatomy of the University of Padua. Histological stainings confirmed the effectiveness of decellularization, resulting in cell-free scaffolds with no residual cells in the matrix. The complex three-dimensional networks of collagen (azan-Mallory) and elastic fibers (Van Gieson) were maintained, in the absence of nuclear material (DAPI staining and DNA extraction/quantification). Ultrastructural analyses with transmission and scanning electron microscopy also showed the preservation of collagen and elastic fibres, together with proteoglycan components. The vascular structures in the tissue were also still visible, with preservation of collagen and elastic wall components and loss of endothelial (anti-CD31 and –CD34 immunohistochemistry) and smooth muscle (anti-alpha smooth muscle actin) cells. Scaffold implantations in rabbits gave good results in terms of integration with surrounding structures (in vivo US analyses and post mortem macro/ microscopic analyses), although recellularization by surrounding muscle cells was not satisfactory. In conclusion, not only animal (rat and rabbit) skeletal muscles but also human samples may be decellularized to obtain complex 3D scaffolds preserving tissue architecture potentially suitable for recellularization. In rabbits, these scaffolds have maintained some biomechanical properties once implanted, although spontaneous recellularization was not achieved. Further analyses will be necessary to verify the possibility of in vitro recolonization of scaffolds by homologous myoblasts and autologous satellite cells, before in vivo re-implantation.

Prevention of Capsular Contracture with Guardix-SG® After Silicone Implant Insertion

Capsular contracture is the most common side effect of breast implant insertion and the problem that breast surgeons seek to avoid the most. Previous animal studies have proved that an antiadhesive barrier solution (AABS) prevents peri-implant capsule formation. In this study, the authors sought to explore the effect that Guardix-SG(®), an AABS that can encapsulate implants in the form of a gel, can have on capsular contracture. This study used 12 female New Zealand white rabbits weighing 2.5-3 kg. Implants were inserted into the subpanniculus carnosus plane through an incision in the bilateral midback area. Once the implant was inserted, 3 g of Guardix-SG(®) and normal saline were instilled into the left and right sides, respectively. The rabbits were killed 6 months after the procedure. The intracapsular pressure was measured using tonometry with a 38.2-g circular glass piece, and capsular thickness was measured by dyeing the biopsy specimen with hematoxylin and eosin and Masson's trichrome stain. The myofibroblasts were quantitatively analyzed through monoclonal anti-alpha smooth muscle actin antibody immunohistochemistry staining. The intracapsular pressure in the control group (4.51 ± 0.98 mmHg) was significantly higher (p = 0.002) than in the study group (3.51 ± 0.4 mmHg). The average capsular thickness was significantly greater in the control group (0.33 ± 0.15 mm; p = 0.015). In the analysis, the interrelation between capsular thickness and intracapsular pressure was insignificant in both groups, as was the number of myofibroblasts in both groups (p = 0.582). Through this study, the authors were able to demonstrate that capsular contracture can be suppressed in the rabbit model by instilling Guardix-SG(®) after insertion of cohesive gel implants in the subpanniculus carnosus plane. This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266 .

Decellularized omentum as novel biologic scaffold for reconstructive surgery and regenerative medicine

Homologous tissues, such as adipose tissue, may be an interesting source of acellular scaffolds, maintaining a complex physiological three-dimensional (3D) structure, to be recellularized with autologous cells. The aim of the present work is to evaluate the possibility of obtaining homologous acellular scaffolds from decellularization of the omentum, which is known to have a complex vascular network. Adult rat and human omenta were treated with an adapted decellularization protocol involving mechanical rupture (freeze-thaw cycles), enzymatic digestion (trypsin, lipase, deoxyribonuclease, ribonuclease) and lipid extraction (2-propanol). Histological staining confirmed the effectiveness of decellularization, resulting in cell-free scaffolds with no residual cells in the matrix. The complex 3D networks of collagen (azan-Mallory), elastic fibers (Van Gieson), reticular fibers and glycosaminoglycans (PAS) were maintained, whereas Oil Red and Sudan stains showed the loss of lipids in the decellularized tissue. The vascular structures in the tissue were still visible, with preservation of collagen and elastic wall components and loss of endothelial (anti-CD31 and -CD34 immunohistochemistry) and smooth muscle (anti-alpha smooth muscle actin) cells. Fat-rich and well vascularized omental tissue may be decellularized to obtain complex 3D scaffolds preserving tissue architecture potentially suitable for recellularization. Further analyses are necessary to verify the possibility of recolonization of the scaffold by adipose-derived stem cells in vitro and then in vivo after re implantation, as already known for homologus implants in regenerative processes.

Tamoxifen decreases the myofibroblast count in the healing bile duct tissue of pigs

OBJECTIVE:The aim of this study was to evaluate the effect of oral tamoxifen treatment on the number of myofibroblasts present during the healing process after experimental bile duct injury.METHODS:The sample consisted of 16 pigs that were divided into two groups (the control and study groups). Incisions and suturing of the bile ducts were performed in the two groups. Tamoxifen (20 mg/day) was administered only to the study group. The animals were sacrificed after 30 days. Quantification of myofibroblasts in the biliary ducts was made through immunohistochemistry analysis using anti-alpha smooth muscle actin of the smooth muscle antibody. Immunohistochemical quantification was performed using a digital image system.RESULTS:In the animals treated with tamoxifen (20 mg/day), there was a significant reduction in immunostaining for alpha smooth muscle actin compared with the control group (0.1155 vs. 0.2021, p = 0.046).CONCLUSION:Tamoxifen reduced the expression of alpha smooth muscle actin in the healing tissue after bile duct injury, suggesting a decrease in myofibroblasts in the scarred area of the pig biliary tract. These data suggest that tamoxifen could be used in the prevention of biliary tract stenosis after bile duct surgeries.