Glial cell line-derived neurotrophic factor added to a sciatic nerve fragment grafted in a spinal cord gap ameliorates motor impairments in rats and increases local axonal growth

Purpose: The aversive nature of regenerative milieu is the main problem related to the failure of neuronal restoration in the injured spinal cord which however might be addressed with an adequate repair intervention. We evaluated whether glial cell line-derived neurotrophic factor (GDNF) may increase the ability of sciatic nerve graft, placed in a gap promoted by complete transections of the spinal cord, to enhance motor recovery and local fiber growth. Methods: Rats received a 4 mm-long gap at low thoracic level and were repaired with a fragment of the sciatic nerve. GDNF was added (NERVE+GDNF) or not to the grafts (NERVE-GDNF). Motor behavior score (BBB) and sensorimotor tests-linked to the combined behavior score (CBS), which indicate the degree of the motor improvement and the percentage of functional deficit, respectively, and also the spontaneous motor behavior in an open field by means of an infrared motion sensor activity monitor were analyzed. At the end of the third month post surgery, the tissue composed by the graft and the adjacent regions of the spinal cord was removed and submitted to the immunohistochemistry of the neurofilament-200 (NF-200), growth associated protein-43 (GAP-43), microtubule associated protein-2 (MAP-2), 5-hidroxytryptamine (serotonin, 5-HT) and calcitonin gene related peptide (CGRP). The immunoreactive fibers were quantified at the epicenter of the graft by means of stereological procedures. Results: Higher BBB and lower CBS levels (p < 0.001) were found in NERVE+GDNF rats. GDNF added to the graft increased the levels of individual sensorimotor tests mainly at the third month. Analysis of the spontaneous motor behavior showed decreases in the time and number of small movement events by the third month without changes in time and number of large movement events in the NERVE+GDNF rats. Immunoreactive fibers were encountered inside the grafts and higher amounts of NF-200, GAP-43 and MAP-2 fibers were found in the epicenter of the graft when GDNF was added. A small amount of descending 5-HT fibers was seen reentering in the adjacent caudal levels of the spinal cords which were grafted in the presence of GDNF, event that has not occurred without the neurotrophic factor. GDNF in the graft also led to a large amount of MAP-2 perikarya and fibers in the caudal levels of the cord gray matter, as determined by the microdensitometric image analysis. Conclusions: GDNF added to the nerve graft favored the motor recovery, local neuronal fiber growth and neuroplasticity in the adjacent spinal cord.

Histologic Study of Perifascial Areolar Tissue Implanted in Rabbit Vocal Folds: An Experimental Study

Objectives: Perifascial areolar tissue (PAT) consists of loose areolar tissue with viscoelastic properties that are similar to those found in tissues in the superficial layer of the vocal fold. The aim of this study was to quantify the inflammatory process and the collagen content of the graft, as well as that of the host tissue, after placement of a strip of PAT into the rabbit vocal fold. Methods: Surgeries were performed on 30 rabbits. The grafts were implanted in pockets that were surgically created in the right vocal fold. The left vocal fold (control group) was subjected only to surgical manipulation. The animals were divided into 3 groups for evaluations at 15 days, 3 months, and 6 months, and their larynx tissues were subsequently reviewed by histology. Results: The grafts were characterized by disorganized and thick collagen bundles and were identified in all study groups. The collagen density stayed constant over time. There was an acute inflammatory response induced by the graft at 15 clays that did not exist in the specimens taken at 3 and 6 months. Deposition of collagen fibers in the lamina propria was observed starting at 15 days after the operation and was more intense in the experimental vocal fold than in the control vocal fold. Conclusions: Our findings indicated that PAT has a low tendency for promoting an inflammatory response. However, there was a loss of the original architecture of the graft tissue and a greater deposition of collagen in the implanted vocal folds than in the control group.

Estrogen effects on pilocarpine-induced temporal lobe epilepsy in rats

Objetive: To evaluate the effects of conjugated equine estrogens (CEE) on the pilocarpine-induced epilepsy in rats. Study design: 40 female rats were divided into: GPC (positive control) presented ""status epilepticus"" (SE) induced by pilocarpine; GOC(ovariectomized control) only castrated; GNC (negative control) received only saline solution; GPE received pilocarpine, presented SE, castrated and received 50 mu g/kg CEE treatment; GPV received pilocarpine, castrated and received propylene glycol (vehicle). The animals were monitored by a video system. At the end of observation, the brains removed for later histologic analysis using Neo-Timm and Nissl methods. Results: The GPE presented a reduction in number of seizures compared to GPV. The Neo-Timm analysis showed that GPV had greater sprouting of mossy fibers, with a denser band in the area of the dentate gyrus hilum compared to GPE. On Nissl staining, GPE showed evident neuronal loss in the CA3 area. GPV presented loss in CA1 and dentate gyrus. Conclusion: Estrogen may have a protecting effect on the central nervous system. (C) 2008 Elsevier Ireland Ltd. All rights reserved.

Immunophenotyping and remodeling process in small airways of idiopathic interstitial pneumonias: functional and prognostic significance

Background and Aims: To test whether different degrees of immunologic and fibrotic airway remodeling processes occur in idiopathic interstitial pneumonias (IIPs), with impact on functional tests and survival, we studied the collagen/elastic system and immune cell density in the bronchiolar interstitium of lungs with the major types of IIPs. Materials and Methods: Histochemistry, immunohistochemistry and morphometric analysis were used to evaluate collagen/elastic fibers and immune cells in the bronchiolar interstitium of open lung biopsies of patients with cryptogenic organizing pneumonia [COP/organizing pneumonia (OP) = 10], acute interstitial pneumonia [AIP/diffuse alveolar damage (DAD) = 20], nonspecific interstitial pneumonia (NSIP/NSIP = 20) and idiopathic pulmonary fibrosis/usual interstitial pneumonia (UIP) = 20. Results: OP lungs presented a significant increase in collagenous/elastic fibers and in the total density of immune cells in the bronchiolar interstitium compared to controls, DAD, NSIP and UIP. We observed a significant increase in CD4, CD8 and CD20 lymphocytes, as well as in neutrophils, macrophages and plasma cells in OP. The increased amount of elastic fibers in the bronchiolar interstitium from OP lungs has a direct association with forced vital capacity (FVC) (r(s) = 0.99, P = 0.03). The most important survival predictor was CD20+ lymphocytes in the bronchiolar interstitium. In decreasing order, patients with UIP [Odds Ratio (OR) = 35.01], high forced expiratory volume in 1 s (FEV1)/FVC FVC (OR = 7.01), increased CD20+ lymphocytes (OR = 4.44) and collagenous/elastic fiber densities (OR = 2.03 and OR = 1.49, respectively) in the bronchiolar interstitium were those who had the greatest risk of death, followed by those with AIP, NSIP and COP. Conclusion: Different degrees of immunologic and fibroelastotic airway remodeling processes occur in the major types of IIPs with impact on physiological tests and survival.

Effect of ultrasound therapy on the repair of Gastrocnemius muscle injury in rats

The aim of this study was to evaluate the effect of the pulsed ultrasound therapy (PUT) in stimulating myoregeneration and collagen deposition in an experimental model of lacerative gastrocnemius muscle lesion in 30 Wistar rats. Fifteen rats were treated (TG) daily with 1 MHz pulsed ultrasound (50%) at 0.57 W/cm(2) for 5 min, and 15 were control animals (CG). Muscle samples were analyzed on postoperative days 4, 7 and 14 through H&E, Picrosirius-polarization and immunohistochemistry for desmin. The lesions presented similar inflammatory responses in both treated and control groups. The areal fraction of fibrillar collagen was larger in the TG at 4 days post-operatively (17.53 +/- 6.2% vs 6.79 +/- 1.3%, p = 0.0491), 7 days (31.07 +/- 7.45% vs 12.57 +/- 3.6%, p = 0.0021) and 14 days (30.39 +/- 7.3% vs 19.13 +/- 3.51%, p = 0.0118); the areal fraction of myoblasts and myotubes was larger in the TG at 14 days after surgery (41.66 +/- 2.97% vs 34.83 +/- 3.08%, p = 0.025). Our data suggest that the PUT increases the differentiation of muscular lineage cells, what would favor tissue regeneration. On the other hand, it is also suggested that there is a larger deposition of collagenous fibers, what could mean worse functional performance. However, the percentage of fibers seems to have stabilized at day 7 in TG and kept increasing in CG. Furthermore, the collagen supramolecular organization achieved by the TG is also significant according to the Sirius red staining results. (C) 2008 Elsevier B.V. All rights reserved.

Individual risk prediction of nodal and distant metastasis for patients with typical bronchial carcinoid tumors

Objective: Bronchial typical carcinoid tumors are tow-grade malignancies. However, metastases are diagnosed in some patients. Predicting the individual risk of these metastases to determine patients eligible for a radical lymphadenectomy and patients to be followed-up because of distant metastasis risk is relevant. Our objective was to screen for predictive criteria of bronchial typical carcinoid tumor aggressiveness based on a logistic regression model using clinical, pathological and biomolecular data. Methods: A multicenter retrospective cohort study, including 330 consecutive patients operated on for bronchial typical carcinoid tumors and followed-up during a period more than 10 years in two university hospitals was performed. Selected data to predict the individual risk for both nodal and distant metastasis were: age, gender, TNM staging, tumor diameter and location (central/peripheral), tumor immunostaining index of p53 and Ki67, Bcl2 and the extracellular density of neoformed microvessels and of collagen/elastic extracellular fibers. Results: Nodal and distant metastasis incidence was 11% and 5%, respectively. Univariate analysis identified all the studied biomarkers as related to nodal metastasis. Multivariate analysis identified a predictive variable for nodal metastasis: neo angiogenesis, quantified by the neoformed pathological microvessels density. Distant metastasis was related to mate gender. Discussion: Predictive models based on clinical and biomolecular data could be used to predict individual risk for metastasis. Patients under a high individual risk for lymph node metastasis should be considered as candidates to mediastinal lymphadenectomy. Those under a high risk of distant metastasis should be followed-up as having an aggressive disease. Conclusion: Individual risk prediction of bronchial typical carcinoid tumor metastasis for patients operated on can be calculated in function of biomolecular data. Prediction models can detect high-risk patients and help surgeons to identify patients requiring radical lymphadenectomy and help oncologists to identify those as having an aggressive disease requiring prolonged follow-up. (C) 2008 European Association for Cardio-Thoracic Surgery. Published by Elsevier B.V. All rights reserved.

Effects of amiodarone on lung tissue mechanics and parenchyma remodeling

We studied the results of chronic oral administration of amiodarone on in vitro lung tissue mechanics, light and electron microscopy. Fifteen Wistar male rats were divided into three groups. In control (CTRL) group animals received saline (0.5 mL/day). In amiodarone (AMIO) groups, amiodarone was administered by gavage at a dose of 175 mg/kg 5 days per week for 6 (6AMIO) or 12 weeks (12AMIO). Lung tissue strips were analyzed 24 h after the last drug administration. Tissue resistance and elastance were higher in 6AMIO and 12AMIO than in CTRL, while hysteresivity was similar in all groups. Total amount of collagen fibers in lung parenchyma increased progressively with the time course of the lesion. However, at 6 weeks there was an increase in the amount of type III collagen fibers, while in 12AMIO mainly type I collagen fibers were found. In our study amiodarone increased lung tissue impedance that was accompanied by matrix remodeling and lesion of type II pneumocytes. (C) 2008 Elsevier B.V. All rights reserved.

Effects of inducible nitric oxide synthase inhibition in bronchial vascular remodeling-induced by chronic allergic pulmonary inflammation

Vascular remodeling is an important feature in asthma pathophysiology. Although investigations suggested that nitric oxide (NO) is involved in lung remodeling, little evidence established the role of inducible NO synthase (iNOS) isoform in bronchial vascular remodeling. The authors investigated if iNOS contribute to bronchial vascular remodeling induced by chronic allergic pulmonary inflammation. Guinea pigs were submitted to ovalbumin exposures with increasing doses (1 similar to 5 mg/mL) for 4 weeks. Animals received 1400W (iNOS-specific inhibitor) treatment for 4 days beginning at 7th inhalation. Seventy-two hours after the 7th inhalation, animals were anesthetized, mechanical ventilated, exhaled NO was collected, and lungs were removed and submitted to picrosirius and resorcin-fuchsin stains and to immunohistochemistry for matrix metalloproteinase-9 (MMP-9), tissue inhibitor of metalloproteinase-1 (TIMP-1), and transforming growth factor-beta (TGF-beta). Collagen and elastic fiber deposition as well as MMP-9, TIMP-1, and TGF-beta expression were increase in bronchial vascular wall in ovalbumin-exposed animals. The iNOS inhibition reduced all parameters studied. In this model, iNOS inhibition reduced the bronchial vascular extracellular remodeling, particularly controlling the collagen and elastic fibers deposition in pulmonary vessels. This effect can be associated to a reduction on TGF-beta and on metalloproteinase-9/TIMP-1 vascular expression. It reveals new therapeutic strategies and some possible mechanism related to specific iNOS inhibition to control vascular remodeling.

Oral tolerance attenuates changes in in vitro lung tissue mechanics and extracellular matrix remodeling induced by chronic allergic inflammation in guinea pigs

Oral tolerance attenuates changes in in vitro lung tissue mechanics and extracellular matrix remodeling induced by chronic allergic inflammation in guinea pigs. J Appl Physiol 104: 1778-1785, 2008. First published April 3, 2008; doi:10.1152/japplphysiol.00830.2007.-Recent studies emphasize the presence of alveolar tissue inflammation in asthma. Immunotherapy has been considered a possible therapeutic strategy for asthma, and its effect on lung tissue had not been previously investigated. Measurements of lung tissue resistance and elastance were obtained before and after both ovalbumin and acetylcholine challenges. Using morphometry, we assessed eosinophil and smooth muscle cell density, as well as collagen and elastic fiber content, in lung tissue from guinea pigs with chronic pulmonary allergic inflammation. Animals received seven inhalations of ovalbumin (1-5 mg/ml; OVA group) or saline (SAL group) during 4 wk. Oral tolerance (OT) was induced by offering ad libitum ovalbumin 2% in sterile drinking water starting with the 1st inhalation (OT1 group) or after the 4th (OT2 group). The ovalbumin-exposed animals presented an increase in baseline and in postchallenge resistance and elastance related to baseline, eosinophil density, and collagen and elastic fiber content in lung tissue compared with controls. Baseline and post-ovalbumin and acetylcholine elastance and resistance, eosinophil density, and collagen and elastic fiber content were attenuated in OT1 and OT2 groups compared with the OVA group. Our results show that inducing oral tolerance attenuates lung tissue mechanics, as well as eosinophilic inflammation and extracellular matrix remodeling induced by chronic inflammation.

Collagen Fiber and Versican Distribution Within the Lamina Propria of Fetal Vocal Folds

Objectives: To analyze the presence and distribution of collagen fibers and versican in human vocal fold lamina propria of fetal larynges. Study Design: Cross sectional analysis of cadaveric vocal folds of human fetuses. Methods: Seven fetal larynges obtained from 28- to 36-week-old fetuses were analyzed with the Picrosirius-polarization method, immunohistochemistry, and image analysis. Results: Collagen fibers within the lamina propria exhibited a monolaminar distribution pattern and spatial arrangement in ""wicker basket."" Versican distribution was larger in the superficial and intermediate layers when compared to the deep layer. Conclusion: Our findings suggest that collagen and versican distribution and arrangement within the lamina propria in the developing fetus are important for vocalization at birth.

Cholinergic Hyperresponsiveness of Peripheral Lung Parenchyma in Chronic Obstructive Pulmonary Disease

Background: Up to 60% of chronic obstructive pulmonary disease ( COPD) patients can present airway hyperresponsiveness. However, it is not known whether the peripheral lung tissue also shows an exaggerated response to agonists in COPD. Objectives: To investigate the in vitro mechanical behavior and the structural and inflammatory changes of peripheral lung tissue in COPD patients and compare to nonsmoking controls. Methods: We measured resistance and elastance at baseline and after acetylcholine (ACh) challenge of lung strips obtained from 10 COPD patients and 10 control subjects. We also assessed the alveolar tissue density of neutrophils, eosinophils, macrophages, mast cells and CD8+ and CD4+ cells, as well as the content of alpha-smooth muscle actin-positive cells and elastic and collagen fibers. We further investigated whether changes in in vitro parenchymal mechanics correlated to structural and inflammatory parameters and to in vivo pulmonary function. Results: Values of resistance after ACh treatment and the percent increase in tissue resistance (%R) were higher in the COPD group (p <= 0.03). There was a higher density of macrophages and CD8+ cells (p < 0.05) and a lower elastic content (p = 0.003) in the COPD group. We observed a positive correlation between %R and eosinophil and CD8+ cell density (r = 0.608, p = 0.002, and r = 0.581, p = 0.001, respectively) and a negative correlation between %R and the ratio of forced expiratory volume in 1 s to forced vital capacity (r = -0.451, p < 0.05). Conclusions: The cholinergic responsiveness of parenchymal lung strips is increased in COPD patients and seems to be related to alveolar tissue eosinophilic and CD8 lymphocytic inflammation and to the degree of airway obstruction on the pulmonary function test. Copyright (C) 2011 S. Karger AG, Basel

Low-Intensity Swimming Training Partially Inhibits Lipopolysaccharide-Induced Acute Lung Injury

RAMOS, D. S. C. R. OLIVO. F. D. QUIRINO SANTOS LOPES, A. C. TOLEDO, M. A. MARTINS, R. A. LAZO OSORIO. M. DOLHNIKOFF, W. RIBEIRO, and R. R VIEIRA. Low-Intensity Swimming Training Partially Inhibits Lipopolysaccharide-Induced Acute Lung Injury. Med. Sci. Sports Exerc.. Vol. 42, No. 1, pp. 113-119, 2010. Background: Aerobic exercise-decreases pulmonary inflammation and remodeling in experimental models of allergic asthma. However, the effects of aerobic exercise oil pulmonary inflammation of nonallergic Origin, such as in experimental models of acute long injury induced by lipopolysaccharide (LPS), have not been evaluated. Objective: The present study evaluated file effects of aerobic exercise in a model of LPS-induced acute lung injury. Methods: BALB/c mice were divided into four groups: Control, Aerobic Exercise, LPS, and Aerobic Exercise + LPS. Swimming tests were conducted at baseline and at 3 and 6 wk. Low-Intensity swimming training was performed for 6 wk, four times per week, 60 min per session. Intranasal LPS (1 mg.kg(-1) (60 mu g per mouse)) was instilled 24 It after the last swimming physical test in the LPS and Aerobic Exercise + LPS mice, and the animals were studied 24 It after LPS instillation. Exhaled nitric oxide, respiratory mechanics, total and differential cell Counts in bronchoalveolar lavage, and lung parenchymal inflammation and remodeling were evaluated. Results: LPS instillation resulted in increased levels of exhaled nitric oxide (P < 0.001), higher numbers of neutrophils in file bronchoalveolar lavage (P < 0.001) and in the lung parenchyma (P < 0.001), and decreased lung tissue resistance (P < 0.05) and volume proportion of elastic fibers (P < 0.01) compared with the Control group. Swim training in LPS-instilled animals resulted in significantly lower exhaled nitric oxide levels (P < 0.001) and fewer nelltrophils in the bronchoalveolar lavage (P < 0.001) and the lung parenchyma (P < 0.01) compared with the LPS group. Conclusions: These results Suggest that low-intensity swimming training inhibits lung neutrophilic inflammation, but not remodeling and impaired lung mechanics, in a model of LPS-induced acute lung injury.

The presence of a vocal ligament in fetuses: a histochemical and ultrastructural study

Although it is currently believed that the vocal ligament of humans undergoes considerable development postnatally, there is no consensus as to the age at which it first emerges. In the newborn infant, the lamina propria has been described as containing a sparse collection of relatively unorganized fibres. In this study we obtained larynges from autopsy of human fetuses aged 7-9 months and used light and electron microscopy to study the collagenous and elastic system fibres in the lamina propria of the vocal fold. Collagen fibres were viewed using the Picrosirius polarization method and elastic system fibres were stained using Weigert`s resorcin-fuchsin after oxidation with oxone. The histochemical and electron microscopic observations were consistent, showing collagen populations with an asymmetric distribution across different compartments of the lamina propria. In the central region, the collagen appeared as thin, weakly birefringent, greenish fibres when viewed using the Picrosirius polarization method, whereas the superficial and deep regions contained thick collagen fibres that displayed a strong red or yellow birefringence. These findings suggest that the thin fibres in the central region consist mainly of type III collagen, whereas type I collagen predominates in the superficial and deep regions, as has been reported in studies of adult vocal folds. Similarly, elastic system fibres showed a differential distribution throughout the lamina propria. Their distribution pattern was complementary to that of collagen fibres, with a much greater density of elastic fibres apparent in the central region than in the superficial and deep regions. This distribution of collagen and elastic fibres in the fetal vocal fold mirrors that classically described for the adult vocal ligament, suggesting that a vocal ligament has already begun to develop by the time of birth. The apparently high level of organization of connective tissue components in the newborn is in contrast to current hypotheses that argue that the mechanical stimuli of phonation are essential to the determination of the layered structure of the lamina propria and suggests that genetic factors may play a more significant role in the development of the vocal ligament than previously believed.

Direct measurement of instantaneous source speed for a HDR brachytherapy unit using an optical fiber based detector

Purpose: Several attempts to determine the transit time of a high dose rate (HDR) brachytherapy unit have been reported in the literature with controversial results. The determination of the source speed is necessary to accurately calculate the transient dose in brachytherapy treatments. In these studies, only the average speed of the source was measured as a parameter for transit dose calculation, which does not account for the realistic movement of the source, and is therefore inaccurate for numerical simulations. The purpose of this work is to report the implementation and technical design of an optical fiber based detector to directly measure the instantaneous speed profile of a (192)Ir source in a Nucletron HDR brachytherapy unit. Methods: To accomplish this task, we have developed a setup that uses the Cerenkov light induced in optical fibers as a detection signal for the radiation source moving inside the HDR catheter. As the (192)Ir source travels between two optical fibers with known distance, the threshold of the induced signals are used to extract the transit time and thus the velocity. The high resolution of the detector enables the measurement of the transit time at short separation distance of the fibers, providing the instantaneous speed. Results: Accurate and high resolution speed profiles of the 192Ir radiation source traveling from the safe to the end of the catheter and between dwell positions are presented. The maximum and minimum velocities of the source were found to be 52.0 +/- 1.0 and 17.3 +/- 1:2 cm/s. The authors demonstrate that the radiation source follows a uniformly accelerated linear motion with acceleration of vertical bar a vertical bar = 113 cm/s(2). In addition, the authors compare the average speed measured using the optical fiber detector to those obtained in the literature, showing deviation up to 265%. Conclusions: To the best of the authors` knowledge, the authors directly measured for the first time the instantaneous speed profile of a radiation source in a HDR brachytherapy unit traveling from the unit safe to the end of the catheter and between interdwell distances. The method is feasible and accurate to implement on quality assurance tests and provides a unique database for efficient computational simulations of the transient dose. (C) 2010 American Association of Physicists in Medicine. [DOI: 10.1118/1.3483780]

Sonographic appearance of gestational trophoblastic disease evolving into epithelioid trophoblastic tumor

Epithelioid trophoblastic tumor is a distinctive but rare trophoblastic tumor. It derives from intermediate trophoblastic cells of the chorion laeve and is usually associated with a previous gestational event. We report the case of a patient who had undergone dilatation and curettage for a missed miscarriage. Three months later gestational trophoblastic disease was suspected because of persistent vaginal bleeding and high levels of beta-human chorionic gonadotropin (beta-hCG). Transvaginal ultrasound revealed irregular echolucent lacunae within the myometrium, some of them filled with low-resistance, turbulent blood flow on Doppler examination, emphasizing the diagnosis of gestational trophoblastic disease. The patient was treated with 12 courses of multiagent chemotherapy. After a 2-year remission, a low rise in serum beta-hCG was observed. Transvaginal ultrasound revealed a well-circumscribed echogenic lesion with a diameter of 1.8 cm in the uterine fundus, with no detectable blood flow on Doppler imaging. A diagnosis of tumor of intermediate trophoblastic cells was suspected and total hysterectomy was performed. On pathological examination, the histological and immunohistochemical features were characteristic of epithelioid trophoblastic tumor. Most reported cases of epithelioid trophoblastic tumor have solitary nodules with sharp margins, which is consistent with our ultrasound findings. Ultrasound may be helpful in differentiating epithelioid trophoblastic tumor from placental-site trophoblastic tumor, another tumor of intermediate trophoblastic cells, which shows infiltrative growth insinuating between muscle fibers. Copyright (C) 2010 ISUOG. Published by John Wiley & Sons, Ltd.