β1 integrin regulation

Integrins are heterodimeric adhesion receptors mediating adhesion to extracellular matrix proteins and to other cells. Integrins are important in embryonic development, structural integrity of connective tissue, blood thrombus formation, and immune defense system. Integrins are transmembrane proteins whose ligand binding capacity (activity) is regulated by large conformational changes. Extracellular ligand binding or intracellular effector binding to integrin cytoplasmic face regulate integrin activity. Integrins are thus able to mediate bi-directional signaling. Integrin function is also regulated by intracellular location. Integrins are constantly recycled from endocytic vesicles to plasma membrane, and this has been shown to be important for cell migration and invasion as well. Deregulation of integrin functionality can lead to deleterious illnesses, such as bleeding or inflammatory disorders. It is also evident that integrin deregulation is associated with cancer progression. In this study, a novel Beta1 integrin associating protein, Rab21, was characterized. Rab21 binding to integrin cytoplasmic tail was shown to be important for Beta1 integrin endo- and exocytosis – intracellular trafficking. It was furher shown that this interaction has an important role in cell adhesion, migration, as well as in the final step of cell division, cytokinesis. This work showed that abrogation of Rab21 function or β1 integrin endocytic traffic, can lead to defects in cell division and results in formation of multinucleated cells. Multinucleation and especially tetraploidy can be a transient pathway to aneuploidy and tumorigenesis. This work characterized chromosomal deletions in rab21 locus in ovarian and prostate cancer samples and showed that a cell line with rab21 deletion also had impairment in cell division, which could be rescued by Rab21 re-expression. The work demonstrates an important role for Rab21 and Beta1 integrin traffic regulation in cell adhesion and division, and suggests a probable associaton with tumorigenesis. In this study, Beta1 integrin activity regulation was also addressed. A novel cell array platform for genome-scale RNAi screenings was characterized here. More than 4500 genes were knocked-down in prostate cancer cells using siRNA-mediated silencing. The effects on Beta1 integrin activity were analyzed upon knock-downs. The screen identified more that 400 putative regulators of Beta1 integrin activity in prostate cancer. In conclusion, this work will help us to understand complex regulatory pathways involved in cancer cell adhesion and migration.

Airway surgery for obstructive sleep apnea and partial upper airway obstruction during sleep

This study analyzed the feasibility and efficacy of surgical therapies in patients with sleep-disordered breathing ranging from partial upper airway obstruction during sleep to severe obstructive sleep apnea syndrome. The surgical procedures evaluated were tracheostomy, laser-assisted uvulopalatoplasty (LUPP) and uvulopalatopharyngoplasty (UPPP) with laser or ultrasound scalpel. Obstructive sleep apnea and partial upper airway obstruction during sleep were measured with the static charge-sensitive bed (SCSB) and pulse oximeter. The patients with severe obstructive sleep apnea syndrome were treated with tracheostomy. Palatal surgery was performed only if the upper airway narrowing occurred exclusively at the soft palate level in patients with partial upper airway obstruction during sleep. The ultrasound scalpel technique was compared to laser-assisted UPPP. The efficacy of LUPP to reduce partial upper airway obstruction during sleep was assessed and histology of uvulopalatal specimen was compared to body fat distributional parameters and sleep study findings. Tracheostomy was effective therapy in severe obstructive sleep apnea. Partial upper airway obstruction and arterial oxyhemoglobin desaturation index during sleep decreased significantly after LUPP. The minimal retropalatal airway dimension increased and soft palate collapsibility decreased at the level where the velopharyngeal obstruction had occurred before the surgery. Ultrasound scalpel did not offer any significant benefits over the laser-assisted technique, except fewer postoperative haemorrhage events. The loose connective tissue as a manifestation of edema was the only histological finding showing correlation with partial upper airway obstruction parameters of SCSB. Tracheostomy remains a life-saving therapy and also long-term option when adherence to CPAP fails in patients with obstructive sleep apnea syndrome. LUPP effectively reduces partial upper airway obstruction during sleep provided that obstruction at the other levels than the soft palate and uvula were preoperatively excluded. Technically the ultrasound scalpel or laser surgeries are equal. In patients with partial upper airway obstruction the loose connective tissue is more important than fat accumulation in the soft palate. This supports the hypothesis that edema is a primary trigger for aggravation of upper airway narrowing during sleep at the soft palate level and evolution towards partial or complete upper airway obstruction during sleep.

Tumor estromal gastrointestinal de intestino delgado

Stromal tumors of the gastrointestinal tract (gists) represent relatively rare lesions that arise from connective tissue elements located along the entire length of the gut. They were initially identified by immunohistochemical investigation, proving their origin from nondifferentiated mesenchymal cells. Only a minority of this lesions, mainly those confined to the esophagus and rectum, have been shown to correspond to mature, well-differentiated types of neoplasms such as leiomyoma or leiomyosarcoma. The majority of gists corresponds to a heterogeneous group of lesions that have as their common denominator an immature proliferation of epithelioid or spindle cells arising from its muscle layers, or between them, showing partial or incomplete myoide, neural, ganglionic, or mixed features of differentiation. This case report intends to show a gist of small bowel in a male, 46 years old, with a two-year of evolution.

Closure of large wounds using rubber bands in rabbits

OBJECTIVE: to verify the effectiveness of the rubber elastic band in the treatment of large wounds of the body wall of rabbits by means of traction of its edges. METHODS: we studied 30 New Zealand rabbits, divided into three groups (n=10): Group 1- healing by secondary intention; Group 2- removal and eutopic repositioning of skin as full thickness skin graft; Group 3- Approximation of wound edges with elastic rubber band. In all animals, we removed a segment of the back skin and subcutaneous tissue down to the fascia, in accordance with an acrylic mold of 8cm long by 12cm wide. All animals were observed for 21 days. RESULTS: two animals of groups 1 and 2 had wound abscess. In Group 2, there was partial or total graft loss in 90% of animals. The complete closure of the wounds was observed in four animals of Group 1, six of Group 2 and eight of Group 3. There was no difference between the scar resistance values of groups 2 and 3, which were higher than those in Group 1. The scars of the three groups were characterized by the presence of mature connective tissue mixed with blood vessels and inflammatory infiltration, predominantly polymorphonuclear. CONCLUSION: the tensile strength of the wound edges with rubber elastic band is as efficient as the skin graft to treat rabbits' large body wounds.

Nerve endings of filliform, fungiform and vallate papillae of dorsal tongue mucosa of White-lipped peccary (Tayassu pecari): Neurohistological observations

The neurohistologic observations were performed using the specimens prepared by Winkelmann and Schmitt silver impregnation method. The tissues were fixed in 10% formalin solution and sections of 40µm thickness were obtained by Leica Cryostat at -30ºC. The sections of dorsal mucosa of White-lipped peccary tongue showed numerous filliform and fungiform papillae, and two vallate papillae on the caudal part. The epithelial layer revealed queratinized epithelial cells and the connective tissue papillae of different sizes and shapes. Thick nerve fiber bundles are noted into the subepithelial connective tissue of the papillae. The connective tissue of fungiform and vallate papillae contained numerous sensitive nerves fibers bundles forming a complex nerve plexus.

Morphology of the male agouti accessory genital glands (Dasyprocta prymnolopha Wagler, 1831)

The morphology of the accessory genital glands of the male agouti was studied in twenty-three animals that were raised in captivity. Twenty animals had their genital glands dissected in situ for macroscopic description. The samples of each gland were recovered, embedded in paraffin, sliced and stained by Hematoxylin-Eosin technique. It was founded four pairs of glands: the vesicular glands, the coagulating glands, the prostate and the bulbourethral glands. Histological characteristics of the vesicular, coagulating and prostate glands showed similar morphology, within the pseudostratified columnar epithelium. The tubulo-alveolar type of the bulbourethral glands showed a lack of connective tissue among the tubules, a small amount of red stained presented it the cytoplasm, and the presence of vacuoles in the tissue. This study concluded that the agouti showed to have similar morphological aspect described in the others species of rodents.

Nutritional fibrous osteodystrophy in goats

Seven out of 25 goats from a southern Brazilian flock developed nutritional fibrous osteodystrophy. Affected animals were younger than 1 year of age and were confined in stalls and fed a concentrate ration containing 1:6 calcium:phosphorus ratio. The remaining flock (35 goats) was managed at pasture and showed no disease. Clinical signs were characterized by mandibular and maxillary enlargements, varying degrees of mouth opening and protruding tongue, dyspnea, apart of abnormalities of prehension and mastication. Affected animals had increased seric levels of phosphorus and parathormone, as well as higher alkaline phosphatase activity. Postmortem examination on three succumbed goats revealed bilateral enlargement of the maxilla and mandibula, and loose teeth, apart of multiple incomplete rib fractures in one of them. Severe diffuse proliferation of loose connective tissue surrounded the osteoid trabeculae, many of which were partially or completely non-mineralized. Mineralized osteoid trabeculae showed osteoclasts in the Howship's lacunae.

Morphological and morphometric evaluation of prepubertal gilt ovaries, uterine tubes and uterus at different oestrus cycle stages

Studies are performed in developing techniques/procedures that provide greater reproductive performance in farm animals, including pigs. In this sense, the study of gilts reproductive organs at different oestrus cycle stages for assessing the presence of abnormalities and/or other parameters that may affect the future animal fertility is important. In order to evaluate the morphological, morphometric and histomorphometric features of ovaries, uterus and uterine tubes (UTs) characteristics of prepubertal gilts at different oestrus cycle stages, reproductive tracts from 48 animals immediately after slaughter were obtained. After, the structures were dissected and removed, and the ovaries were used for classification of oestrus cycle stage of each gilt in follicular phase (FP) and luteal phase (FL). Then, morphometric evaluations of ovaries, UTs, uterine horns and uterine body were performed. Besides that, medial segments of UTs and uterus were fixed in Bouin solution, processed and included in paraffin, when histological sections of 5.0 micrometers (µm) were obtained and stained with Hematoxylin and Eosin. Histomorphometric analyzes using image capture system and specific software were performed. Afterwards, data were submitted to Student's t test for assessment the statistical differences (P<0.05) between the two different oestrus cycle stages (FP × LP) and between the placement of reproductive structures (right × left antimer). Among the gilts evaluated, 35 were in the FP and 13 in LP. There was no difference (P>0.05) between morphometric parameters of ovaries, UTs and uterus of gilts in FP and LP. Likewise, in respect to the placement of reproductive structures, both in the oestrus cycle stages, as in the general average, there was no difference (P>0.05). Regarding the histomorphometric variables, gilts classified in FP presented a higher (P<0.05) height of glandular and UT epithelium compared to animals in LP. On the other hand, the diameter of endometrial glands was higher (P<0.05) in gilts at LP compared to FP. Furthermore, gilts in LP presented a higher (P<0.05) proportion of endometrium occupied by glands, whereas animals in FP had a higher (P<0.05) proportion of connective tissue and blood vessels. In conclusion, in prepubertal gilts, the histomorphometric parameters as endometrial glands diameter, the height of glandular epithelium and of UT epithelium and the proportion of endometrium occupied by connective tissue, besides the glands and blood vessels varies through the oestrus cycle, possibly under the influence of ovarian steroids.

Distribution of immune response cells in the pelvic urethra and the prepuce of rams

The pathogens of the reproductive system in the male can penetrate and establish by ascending route, from to the prepuce to the urethra, accessory glands, epididymis and testicles. The aim of this paper is determine the distribution and number of cells involved in the immune response in prepuce and pelvic urethra of rams, without apparent clinical alterations in testicle, epididymis and prepuce. The distribution of some of the cells involved in the immune response at the level of the prepuce and the pelvic urethra was quantified in four one-year-old rams seronegative for B. ovis and A. seminis and without apparent lesions in the testicles, the epididymis, and the prepuce. At the moment of slaughter, samples were taken from the preputial fornix and the pelvic urethra and placed in 10% formalin and under freezing conditions. CD4, CD8, WC1, CD45RO, CD14 and CD1b cells were demonstrated by immunohistochemistry, and immunoglobulin-containing cells (ICC) of the IgA, IgG and IgM classes were demonstrated by immunofluorescence. The labeled cells present in the mucosa of both organs were counted with an image analyzer. The total number of cells was compared between both tissues and differentially between the epithelium and the connective tissue of the mucosa. Significant differences were found in the total number of CD4, CD45RO, and WC1 lymphocytes, in CD14 macrophages, and CD1b dendritic cells, with mean values being greater in the fornix than in the urethra (p<0.05) in all cases. Only dendritic cells were found in the prepuce. No differences were found in the number of CD8 lymphocytes between both organs. The ratio between each cell type in the connective and the intraepithelial tissues and between organs was 10/1 for CD4 in the fornix (p<0.05), against 7/1 in the urethra (p<0.05), while CD8 had a 1/1 distribution in both mucosae. The WC1 ratio was 5/1 in both mucosae (p<0.05). CD45RO labeling was 19/1 in the prepuce (p<0.05) and 1/1 in the urethra. IgA-containing cells did not show differences in the total number of cells in both tissues. In the urethra, no IgG-containing cells were observed and IgM-containing cells were scarce; in contrast, both cell types were present in the prepuce, in amounts greater than in the urethra (p<0.05). IgA-, IgG-, and IgM-containing cells were located in both organs in the mucosal connective tissue. The presence of antigen-presenting cells, macrophages, and dendritic cells, as well as of lymphocytes CD4, CD8 TCR γδ (WC1), IgA-, IgG and IgM positive cells, and CD45RO cells suggests that both mucosae may behave as inductive and effector sites for the mucosal immune response.

Hematological and histopathological evaluation of wildlife green turtles (Chelonia mydas) with and without fibropapilloma from the north coast of São Paulo State, Brazil

Blood profiles were determined in 47 juvenile green turtles, Chelonia mydas, from São Paulo northern coast, Brazil. Twenty-nine were affected by fibropapillomas and 18 were tumor free. Complete gross and histopathologic examinations of the fibropapillo were performed in 21 green turtles. Biometrical data, size, location and amount of tumors were recorded. The papillomas varied in morphology, location, size, color and texture. We found hyperplastic stroma, rich in blood vessels and connective tissue with increase in thickness of the dermis. The tumors w0ere classified as papillomas or fibropapillomas according to their epithelial and/or stromal proliferation. The lowest Mean Corpuscular Hemoglobin (HCM) values were observed in affected turtles.

Morphoquantitative description of bovine digital cushion

Abstract The digital cushion is characterized as a modified subcutaneous tissue that absorbs the shock during gait, assists venous return of the hoof and supports a considerable part of body weight. Digital cushions have particular importance in the pathogenesis of the hoof, since they need to properly work in order to prevent compression and traumas in soft tissues. This study aimed to measure and determine how is the arrangement of these structures, and for this it was established the proportions of connective, adipose, vascular tissues and collagen fibers and collagen types found in palmar and plantar digital cushion of bovine using fore and hindlimbs of twelve adult zebu cattle of both sexes, 11 male and one female, with 269kg average carcass weight and without limb disorders. Fragments of cushions were subjected to conventional histology, cut to a thickness of 4µm and stained with Red Picrosirius. With digital optical microscope, the quantification of the connective tissue and differentiation of types of collagen used the Image Pro Plus® software, and of adipose and vascular tissue, the test point system. The mean and standard error were estimated with the GraphPad Prism 5.0 software, and then data were subjected to Kolmogorov-Smirnov normality test and Student's t-test with significance level set at 5% for determining the amount of different tissues between fore and hindlimbs of studied animals. In forelimbs the mean and standard error of the connective tissue proportion was 50.10%+1.54, of the adipose tissue was 21.34%+1.44, and of vascular tissue was 3.43%+0.28. Hindlimbs presented a proportion of connective tissue of 61.61%+1.47, 20.66%+1.53 of adipose tissue, and 3.06%+0.20 of vascular tissue. A significant difference (p<0.001) was detected in the connective tissue proportion between fore and hindlimbs. Types I and II collagen fibers have presented, respectively, a proportion of 31.89% and 3.9% in forelimbs and 34.05% and 1.78% in hindlimbs. According to the used methodology, digital cushions had a clear differentiation relative to adipose tissue between fore and hindlimbs.

Prevalence study of SNP c.421G>T in the ADAMTS2 gene responsible for dermatosparaxis in White Dorper sheep in Brazil

Abstract: Dermatosparaxis is an autosomal recessive disorder of connective tissue; the disorder is clinically characterized by skin fragility and hyperextensibility. Dermatosparaxis in White Dorper sheep is caused by a single nucleotide polymorphism (SNP) (c.421G>T) in the ADAM metalloproteinase with thrombospondin type 1 motif, 2 (ADAMTS2) gene. The aim of this study was to investigate the prevalence of this SNP in a White Dorper herd in São Paulo state, Brazil. In this study, we collected blood DNA samples from 303 White Dorper sheep and performed polymerase chain reaction to amplify the SNP region. The samples were sequenced to determine the presence of the SNP in the ADAMTS2 gene. The SNP prevalence in the studied population was 15.5%; this finding indicates that more effective control measures should be used to prevent the inheritance of SNP c.421G>T in the ADAMTS2 gene in Brazilian White Dorper herds.

The Effects of Fibroblast Growth Factor 8b on Reproductive Organs and Prostate Tumorigenesis

Fibroblast growth factors (FGFs) are involved in the development and homeostasis of the prostate and other reproductive organs. FGF signaling is altered in prostate cancer. Fibroblast growth factor 8 (FGF8) is a mitogenic growth factor and its expression is elevated in prostate cancer and in premalignant prostatic intraepithelial neoplasia (PIN) lesions. FGF8b is the most transforming isoform of FGF8. Experimental models show that FGF8b promotes several phases of prostate tumorigenesis - including cancer initiation, tumor growth, angiogenesis, invasion and development of bone metastasis. The mechanisms activated by FGF8b in the prostate are unclear. In the present study, to examine the tumorigenic effects of FGF8b on the prostate and other FGF8b expressing organs, an FGF8b transgenic (TG) mouse model was generated. The effect of estrogen receptor beta (ERβ) deficiency on FGF8binduced prostate tumorigenesis was studied by breeding FGF8b-TG mice with ERβ knockout mice (BERKOFVB). Overexpression of FGF8b caused progressive histological and morphological changes in the prostate, epididymis and testis of FGF8b-TG-mice. In the prostate, hyperplastic, preneoplastic and neoplastic changes, including mouse PIN (mPIN) lesions, adenocarcinomas, sarcomas and carcinosarcomas were present in the epithelium and stroma. In the epididymis, a highly cancer-resistant tissue, the epithelium contained dysplasias and the stroma had neoplasias and hyperplasias with atypical cells. Besides similar histological changes in the prostate and epididymis, overexpression of FGF8b induced similar changes in the expression of genes such as osteopontin (Spp1), connective tissue growth factor (Ctgf) and FGF receptors (Fgfrs) in these two tissues. In the testes of the FGF8b-TG mice, the seminiferous epithelium was frequently degenerative and the number of spermatids was decreased. A portion of the FGF8b-TG male mice was infertile. Deficiency of ERβ did not accelerate prostate tumorigenesis in the FGF8b-TG mice, but increased significantly the frequency of mucinous metaplasia and slightly the frequency of inflammation in the prostate. This suggests putative differentiation promoting and anti-inflammatory roles for ERβ. In summary, these results underscore the importance of FGF signaling in male reproductive organs and provide novel evidence for a role of FGF8b in stromal activation and prostate tumorigenesis.

Unidirectional fiber-reinforced composite as an oral implant abutment material: Experimental studies of E-glass fiber/BisGMA-TEGDMA polymer in vitro

Fiber-reinforced composites (FRCs) are a new group of non-metallic biomaterials showing a growing popularity in many dental and medical applications. As an oral implant material, FRC is biocompatible in bone tissue environment. Soft tissue integration to FRC polymer material is unclear. This series of in vitro studies aimed at evaluating unidirectional E-glass FRC polymer in terms of mechanical, chemical, and biological properties in an attempt to develop a new non-metallic oral implant abutment alternative. Two different types of substrates were investigated: (a) Plain polymer (BisGMA 50%–TEGDMA 50%) and (b) Unidirectional FRC. The mechanical behavior of high fiber-density FRCs was assessed using a three-point bending test. Surface characterization was performed using scanning electron and spinning disk confocal microscopes. The surface wettability/energy was determined using sessile drop method. The blood response, including blood-clotting ability and platelet morphology was evaluated. Human gingival fibroblast cell responses - adhesion kinetics, adhesion strength, and proliferation activity - were studied in cell culture environment using routine test conditions. A novel tissue culture method was developed and used to evaluate porcine gingival tissue graft attachment and growth on the experimental composite implants. The analysis of the mechanical properties showed that there is a direct proportionality in the relationship between E-glass fiber volume fraction and toughness, modulus of elasticity, and load bearing capacity; however, flexural strength did not show significant improvement when high fiber-density FRC is used. FRCs showed moderate hydrophilic properties owing to the presence of exposed glass fibers on the polymer surface. Blood-clotting time was shorter on FRC substrates than on plain polymer. The FRC substrates also showed higher platelet activation state than plain polymer substrates. Fibroblast cell adhesion strength and proliferation rate were highly pronounced on FRCs. A tissue culture study revealed that gingival epithelium and connective tissue established an immediate close contact with both plain polymer and FRC implants. However, FRC seemed to guide epithelial migration outwards from the tissue/implant interface. Due to the anisotropic and hydrophilic nature of FRC, it can be concluded that this material enhances biological events related with soft tissue integration on oral implant surface.

Repair of the anophthalmic cavity of rats with synthetic hydroxyapatite

We placed spheres of synthetic hydroxyapatite (calcium chloride combined with sodium phosphate) in the eviscerated or enucleated orbital cavity of rats in order to evaluate the biocompatibility of this material with the orbital cavity. The study was conducted on 50 albino rats, 25 of which were submitted to enucleation and 25 to evisceration of one eye. The animals were sacrificed 7, 15, 21, 30 and 60 days after surgery and the orbital content was submitted to histopathological examination. A reaction of the young granulation tissue type was observed first. The hydroxyapatite was gradually surrounded by a granulomatous macrophage inflammatory response and covered with dense connective tissue that formed a sort of" mesh" septating and supporting progressively smaller blocks of the substance. The same type of reaction was observed in the enucleated and eviscerated cavities. We conclude that synthetic hydroxyapatite is an inert nonallergenic material which is appropriate for volume replacement in the anophthalmic cavity