Thesis (M.S.)--University of Nevada, Reno, 2008. / "December, 2008." Includes bibliographical references (leaves 60-64). Online version available on the World Wide Web.

O anestésico local é o medicamento mais utilizado na Odontologia e sua associação com vasoconstrictores aumenta a duração da anestesia, diminuindo seus efeitos sistêmicos. A hipertensão e o tabagismo são freqüentes na população, sendo responsáveis por complicações sistêmicas. A felipressina, por não interferir com receptores simpáticos, poderia ser um vasoconstrictor indicado para pacientes hipertensos. O objetivo deste trabalho foi estudar a reatividade cardiovascular de animais simultaneamente hipertensos e fumantes passivos aos agentes vasoconstritores associados aos anestésicos locais, verificando também o efeito do tratamento com atenolol. Foram utilizados ratos Wistar machos, divididos em 5 grupos: 1) normotensos não fumantes; 2) normotensos fumantes passivos; 3) hipertensos não fumantes; 4) hipertensos fumantes passivos; 5) hipertensos fumantes passivos tratados com atenolol. A hipertensão renal foi induzida pela remoção do rim direito e instalação de clip de prata (abertura 0,25mm) na artéria renal esquerda, após anestesia com quetamina e xilazina. Os ratos fumantes passivos foram colocados diariamente por 10 minutos, durante 28 dias, em caixa de madeira de 30cmX25cmX15cm dividida em dois compartimentos. Em um deles, eram acesos 10 cigarros e no outro ficavam os animais. A tampa da caixa era fechada e um sistema de ventilação lançava fumaça dos cigarros para o compartimento dos ratos, num fluxo de 10l/min. Após medida indireta da pressão arterial, 14 dias após a cirurgia, o grupo tratado com atenolol foi medicado durante 14 dias seguintes (90 mg/Kg) por gavage. No 28o dia, todos receberam catéter de polietileno na artéria carótida esquerda (para medida de pressão) e outro na veia jugular direita (para injeção de drogas). Para os 5 grupos foram utilizadas: adrenalina (80, 160, 320, 640 e 1280ng) e felipressina (0,125, 0,25, 0,5, 1, 2 e 3 x 10-3UI). O catéter arterial era conectado a transdutor de pressão e o registro realizado por software específico. Foram analisadas: menor resposta hipotensora, maior resposta hipertensora e duração de resposta para cada dose. Os dados foram analisados por análise de variância de medidas repetidas, seguida do teste de Tuckey ou Holm-Sidack, com nível de significância de 5%. Os resultados mostraram que o fumo passivo reduziu significativamente a resposta vasodilatadora produzida pela adrenalina, em animais normotensos e hipertensos, potencializou suas respostas hipertensoras e aumentou a duração das respostas à adrenalina, ampliadas ainda mais pelo tratamento com atenolol. O tratamento com atenolol promoveu aumento adicional das respostas hipertensoras à adrenalina nos hipertensos-fumantes. A felipressina não apresentou ações vasodilatadoras e suas ações hipertensoras foram potencializadas pelo fumo passivo, em amplitude e duração. O atenolol não promoveu aumento adicional da amplitude das respostas à felipressina. Nos animais hipertensos, o tratamento com atenolol associado ao fumo passivo teve efeito expressivo, aumentando significativamente a duração total das respostas à felipressina. A felipressina, quando comparada à adrenalina, não apresentou efeitos hipotensores diretos, a resposta hipertensora máxima foi nitidamente inferior e a duração das respostas à felipressina foi o dobro da adrenalina. Dessa forma, a felipressina se torna uma droga interessante na hipertensão, devido a sua capacidade de promover vasoconstrição prolongada, sem potencializar a atividade simpática sistêmica. / The local anesthetic is the most common drug in dentistry and the associated vasoconstrictors increase the duration of anesthesia, decreasing its systemic effects. Hypertension and smoking are problems commonly found in the general population, being responsible for systemic complications. Felypressin, a vasoconstrictor that does not interact with sympathetic receptors, could be indicated to hypertensive patients. This study investigated the cardiovascular reactivity of hypertensive and passive smoker animals under atenolol treatment to epinephrine and felypressin. Male wistar rats were divided into five groups: 1) normotensive and non-smokers, 2) normotensive and passive smokers, 3) hypertensive and non-smokers, 4) hypertensive and passive smokers; 5) hypertensive, passive smokers and treated with atenolol. Renal hypertension was induced by removal of the right kidney and installation of a silver clip (with 0.25-mm opening) in the left renal artery, after anesthesia with ketamine and xylazine. The passive smoker rats were placed, 10 minutes per day, during 28 days in a 30cmX25cmX15cm wood box divided into two compartments. Ten cigarettes were lit in one compartment, and the rats were placed in the other. The box lid was closed and a ventilation system threw the cigarette smoke to the rat compartment. After indirect measurement of blood pressure, 14 days after the surgery, the group of rats treated with atenolol was medicated during the following fourteen days (90 mg/kg) by gavage. On the 28th day, a polyethylene catheter was inserted into the left carotid artery (for direct blood pressure measurements) and into the right jugular vein (for drug injection). The groups received epinephrine (80, 160, 320, 640 and 1280ng) or felypressin (0.125, 0.25, 0.5, 1, 2 and 3 x 10-3UI). The arterial catheter was connected to a pressure transducer and recording was made by a specific computer software. The following parameters were analyzed for all groups: lower hypotensive response, higher hypertensive response and duration of response for each dose. Data were statistically analyzed by repeated measures analysis of variance, followed by Tukey test or Holm-Sidack test, at a significance level of 5%. The results showed that passive smoking significantly decreased the vasodilator response produced by epinephrine in normotensive and hypertensive animals, increasing their hypertensive responses and increased the duration of response to epinephrine, that was further increased by atenolol treatment. Atenolol treatment increased the hypertensive responses in hypertensive-smokers rats. The felypressin did not show vasodilator responses and its hypertensive responses were increased by passive smoking. The atenolol did not cause additional increase in felypressin responses. In hypertensive animals, the atenolol treatment associated with passive smoking had expressive effects, significantly increasing the total duration of response to felypressin. Felypressin, when compared with epinephrine, did not show direct hypotensive effects, the higher hypertensive responses were smaller and the duration of response to felypressin was twice the epinephrine time. Then, felypressin becomes an interesting drug to hypertensive patients, due to its capacity to promote prolonged vasoconstrictor effect without increasing the sympathetic nerve activity.

Indiana University-Purdue University Indianapolis (IUPUI) / Cigarette smoking (CS) continues to be a significant modifiable factor contributing to a variety of diseases including cardiovascular, pulmonary and renal pathologies. It was suggested that smoking have detrimental effect of the body’s progenitor cells of bone marrow and peripheral organs. Since the concept of cell therapy that utilizes adipose stem/stromal cells (ASC) is gaining momentum it becomes critical to assess the therapeutic activities of the progenitors isolated from smokers. This study has revealed that CS negatively impacts the vasculogenic potential of ASC, in vitro, as well as weakening their therapeutic activity in vivo when tested in mouse model of hindlimb ischemia. We hypothesized that the decrease in vasculogenic activity of ASC is attributed to a higher level of expression of an angiostatic factor Activin A by ASC from CS donors. These findings clearly suggest that smokers should be evaluated for potential exclusion from early clinical trials of autologous cell therapies, or assessed as a separate cohort. The donor’s health status should be considered when choosing between autologous vs allogeneic cell therapies. We then examined the effect of CS on development of kidney pathology in mice. CS exposure led to decrease in kidney weights, capillary rarefaction, and cortical blood perfusion, and in parallel led to increase in kidney fibrosis and iron deposition. Interestingly, infusion of healthy ASC to the mice following CSexposure reversed CS-induced damages. This strongly support the notion that ASC-based therapy may provide rejuvenation effect. In the other subset of studies, we hypothesized that CS-induced lung emphysematous changes are preceded by suppression of bone marrow (BM) hematopoietic progenitor cells (HPC). We have revealed that intermittent BM mobilization with AMD3100 may mitigate the CS-induced myelo-suppression and deterioration of lung function and morphology. We observed that treatment of mice with AMD3100, while exposed to CS, preserves HPC at the levels of healthy control mice. Furthermore, AMD3100 treatment preserved lung parenchyma from pathological changes. These data suggest that while CS has a myelo-suppressive effect, administration of AMD3100 preserved BM-HPC and ameliorated lung damage.

No description available.

Pulmonary tuberculosis (TB) remains one of the most infectious causes of death worldwide. Mycobacterium tuberculosis (M.tb), the causative agent of TB is transmitted via infectious aerosols, and in the majority of cases the bacteria is effectively controlled, by the host, resulting in a chronic latent infection. Currently, the only available vaccine is the Bacillus Calmette-Guérin (BCG), which despite being successful in preventing childhood disseminated forms of TB, has failed to control the adult pulmonary TB epidemic. One of the major contributing factors in the failure of the BCG is that although antigen-specific T cells are present at the time of M.tb infection, the recruitment of such T cells into the site of infection is significantly delayed. This delay, while reduced compared to non-vaccinated hosts, allows the bacteria to replicate unchecked within the lung and establish a “foothold” prior to the arrival of protective T cells and subsequent immune control. Thus, novel initiatives seek to close this “immunological gap” through increasing the level of protective T cell responses within the airway mucosa immediately following M.tb infection. We therefore investigated the impact of deliberate modulation of T cell geography following BCG vaccination on the outcome of pulmonary M.tb infection. In addition, a number of environmental factors are also thought to affect the site of M.tb infection: the respiratory mucosa. However, little is currently known about the effects of environmental exposure to allergens and other substances such as cigarette smoke on the outcome of pulmonary TB. Throughout this thesis we have investigated the mechanisms of immune protection and failure of protection against pulmonary M.tb infection within the respiratory mucosa. / Thesis / Doctor of Philosophy (Medical Science)

S100 calcium-binding protein A9 (S100A9) is elevated in plasma and bronchoalveolar lavage fluid (BALF) of patients with chronic obstructive pulmonary disease (COPD), and aging enhances S100A9 expression in several tissues. Currently, the direct impact of S100A9- mediated signaling on lung function and within the aging lung is unknown. Here, we observed that elevated S100A9 levels in human BALF correlated with age. Elevated lung levels of S100A9 were higher in older mice compared with in young animals and coincided with pulmonary function changes. Both acute and chronic exposure to cigarette smoke enhanced S100A9 levels in age-matched mice. To examine the direct role of S100A9 on the development of COPD, S100a9 -/- mice or mice administered paquinimod were exposed to chronic cigarette smoke. S100A9 depletion and inhibition attenuated the loss of lung function, pressure-volume loops, airway inflammation, lung compliance, and forced expiratory volume in 0.05 s/forced vital capacity, compared with age-matched wild-type or vehicleadministered animals. Loss of S100a9 signaling reduced cigarette smoke-induced airspace enlargement, alveolar remodeling, lung destruction, ERK and c-RAF phosphorylation, matrix metalloproteinase- 3 (MMP-3), matrix metalloproteinase-9 (MMP-9), monocyte chemoattractant protein-1 (MCP-1), interleukin-6 (IL-6), and keratinocyte- derived chemokine (KC) release into the airways. Paquinimod administered to nonsmoked, aged animals reduced age-associated loss of lung function. Since fibroblasts play a major role in the production and maintenance of extracellular matrix in emphysema, primary lung fibroblasts were treated with the ERK inhibitor LY3214996 or the c-RAF inhibitor GW5074, resulting in less S100A9-induced MMP-3, MMP-9, MCP-1, IL-6, and IL-8. Silencing Toll-like receptor 4 (TLR4), receptor for advanced glycation endproducts (RAGE), or extracellular matrix metalloproteinase inducer (EMMPRIN) prevented S100A9-induced phosphorylation of ERK and c-RAF. Our data suggest that S100A9 signaling contributes to the progression of smoke-induced and age-related COPD.

Indiana University-Purdue University Indianapolis (IUPUI) / The complex pathogenesis of emphysema involves disappearance of alveolar structures, in part attributed to alveolar cell apoptosis. The mechanism by which cigarette smoke (CS) induces alveolar cell apoptosis is not known. We hypothesized that ceramides are induced by CS via specific enzymatic pathways that can be manipulated to reduce lung cell apoptosis. CS increased ceramides in the whole lung and in cultured primary structural lung cells. Exposure to CS activated within minutes the acid sphingomyelinase, and within weeks the de novo- ceramide synthesis pathways. Pharmacological inhibition of acid sphingomyelinase significantly attenuated CS-induced apoptosis. To understand the mechanisms by which ceramides induce apoptosis, we investigated the cell types affected and the involvement of RTP801, a CS-induced pro-apoptotic and pro-inflammatory protein. Direct lung augmentation of ceramide caused apoptosis of both endothelial and epithelial type II cells. Ceramide upregulated RTP801 and the transgenic loss of RTP801 inhibited only epithelial, but not endothelial cell apoptosis induced by ceramide. In conclusion, CS induces acid sphingomyelinase-mediated ceramide upregulation and apoptosis in a cell-specific manner, which in epithelial cells involves induction of stress response proteins that may further amplify lung injury. Molecular targeting of amplification pathways may provide therapeutic opportunities to halt emphysema progression.

Cigarette smoking is a common and lethal worldwide habit, with considerable mortality stemming from its deleterious effects on heart function. While current theories posit altered blood lipids and fibrinogen metabolism as likely mediators, none have explored the role of the sphingolipid ceramide in exacerbating heart function with smoke exposure. Ceramide production is a consequence of cigarette smoke in the lung, and considering ceramide's harmful effects on mitochondrial function, we sought to elucidate the role of ceramide in mediating smoke-induced altered heart mitochondrial respiration. Lung cells were exposed to cigarette smoke extract and heart cells were exposed to the lung-cell conditioned medium. Adult male mice were exposed sidestream cigarette smoke for 8 weeks with dietary intervention and ceramide inhibition. Ceramides and heart cell or myocardial mitochondrial respiration were determined. Lung cell cultures revealed a robust response to cigarette smoke extract in both production and secretion of ceramides. Heart cells incubated with lung-cell conditioned medium revealed a pronounced inhibition of myocardial mitochondrial respiration, though this effect was mitigated with ceramide inhibition via myriocin. In vivo, heart ceramides increased roughly 600% in adult mice with long-term sidestream cigarette smoke exposure. This resulted in a significant ceramide-dependent reduction in left myocardial mitochondrial respiration, as heart mitochondria from the mice exposed to both smoke and myriocin injections respired normally. These results suggest ceramide to be an important mediator of altered myocardial mitochondrial function with cigarette smoke exposure. Thus, anti-ceramide therapies might be considered in the future to protect heart mitochondrial function with smoke exposure.

No description available.

No description available.