Estudio del efecto de la hipoxia ambiental crónica sobre los tejidos orales de la rata

La hipoxia hipobárica (HX) puede definirse como un descenso de la presión barométrica con la consecuente disminución de la presión parcial de oxígeno en la atmósfera. La exposición a HX constituye un estímulo estresante para el organismo, por lo tanto, este debe desarrollar respuestas que le permitan hacer frente a las exigencias del ambiente, las cuales difieren entre las poblaciones que habitan permanentemente en zonas de gran altitud (hipoxia crónica continua, HCC) y aquellos individuos que trabajan en HX y retornan a zonas de baja altitud para descansar (hipoxia crónica intermitente, HCI). A pesar del hecho de que las respuestas fisiológicas que se desencadenan frente a la HX han sido ampliamente discutidas, las repercusiones de los bajos niveles de oxígeno a nivel de los tejidos orales aún no están claras. La literatura cuenta con estudios que analizan los tejidos periodontales y las glándulas salivales tanto en animales como en humanos expuestos a HX, pero en todos ellos las condiciones de exposición no son en las que habitualmente se desarrolla la vida. Por esta razón, este trabajo de tesis se centra en la evaluación del estado periodontal y glandular en ratas expuestas a 4200 metros de altura simulada, siendo nuestra hipótesis que la exposición a dicha altura causa una merma en la función biológica de los tejidos orales. Dichos tejidos serían capaces de aclimatarse pero esta respuesta podría variar dependiendo de si la exposición a HX es continua o intermitente.En nuestros modelos experimentales, se utilizaron ratas Wistar hembras (en crecimiento o adultas), las cuales se expusieron a HX durante 90 días con el fin de evaluar las propiedades biomecánicas mandibulares, la morfometría del hueso alveolar, la concentración de diversos biomarcadores del tejido gingival y de glándulas submaxilares (GSM), la histoarquitectura de GSM y la secreción salival. Además, hemos realizado experiencias en animales con inducción experimental de enfermedad periodontal (PE), con el fin de analizar el efecto de la HX en un hueso alveolar afectado por periodontitis. Los resultados del presente trabajo indican que la HX afecta negativamente las propiedades biomecánicas mandibulares en ratas en crecimiento, hecho probablemente vinculado a la disminución de la masa ósea de los animales y a la incapacidad de la mandíbula de adaptar su geometría durante la exposición a HX. El hueso alveolar también sufrió alteraciones, observándose reabsorción cortical lingual e interradicular, sin evidencia de desarrollo de una respuesta inflamatoria a nivel periodontal. La mencionada pérdida ósea podría ser considerada como un intento de aclimatación para aumentar la masa roja circulante, proceso fundamentalmente a cargo de la médula ósea de los huesos largos. El incremento de la actividad de la óxido nítrico sintasa inducible observado en el tejido gingival y en la GSM, da cuenta de la necesidad del organismo de aumentar el aporte de O2 en ambos sitios, lo que refuerza el concepto de intento de aclimatación a nivel periodontal y glandular.En ratas adultas, la exposición a HX no indujo cambios biomecánicos mandibulares, probablemente debido a que estos animales ya han completado su formación ósea. Es evidente que la exposición intermitente resultó más perjudicial en términos de aclimatación que la continua. Esto se vio reflejado en la menor secreción salival y en la mayor pérdida ósea interradicular en HCI, así como también en el aumento de la resorción ósea general. Los fenómenos homeostáticos desencadenados en respuesta a HX involucrarían la activación del factor inducible por hipoxia-1α y mecanismos adaptativos a nivel bioenergético mitocondrial, que parecen no ser del todo eficaces en HCI. Estos resultados conjuntamente con el análisis microscópico de las GSM sugieren una diferencia fenotípica durante la aclimatación en HCI y HCC, que involucraría cambios a nivel bioquímico, ultraestructural y molecular.Cuando se evaluó el efecto de la HX sobre ratas adultas sometidas a PE, nuestros resultados sugieren que la exposición a HX agrava las manifestaciones patológicas orales de la periodontitis, tanto a nivel óseo como glandular. Las vías de señalización que se activan a nivel celular durante la exposición a HCI y HCC podrían incrementar las señales de la cascada inflamatoria inducida por periodontitis, aumentando su efecto deletéreo sobre el periodonto y las GSM. En conclusión, la exposición a HX induce cambios en la fisiología del tejido periodontal y glandular de ratas en crecimiento y adultas, de forma diferencial cuando se las expone en forma continua como intermitente. Este trabajo de tesis constituye un antecedente valioso a la hora de evaluar la salud periodontal de individuos que habitan o trabajan en condiciones de HX y ayuda a una mejor comprensión de los procesos que regulan la homeostasis oral durante la aclimatación a la HX, pudiendo contribuir al desarrollo de estrategias para prevenir la hiposecreción salival y sus consecuencias en poblaciones expuestas a HX.Fil: Terrizzi, Antonela Romina. Universidad de Buenos Aires. Facultad de Odontología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin

Caracterización de mitocondrias y estado redox de la glándula submaxilar de ratas expuestas a hipoxia ambiental

Hypobaric hypoxia (HX) can be defined as a decrease in the partial pressure of oxygen (O2) leading to less levels of this gas in the tissues. HX evokes highly coordinated cellular responses in order to preserve cell viability and they differ from those populations that live permanently at high altitude (chronic continuous hypoxia, CCH) compared to those who work in HX conditions and then return to normoxic environments (chronic intermittent HX, CIH). Despite the fact that the effects of HX in the organism are well established, its role on oral health is still not clear. It has been reported salivary dysfunction in rats exposed to HX, but the mechanisms are still unknown, especially under these two types of HX conditions. In this sense, exposure to HX is known to increase cellular oxidative stress with consequent damage to lipids, proteins and DNA. As mitochondria are the most important organelle related with reactive oxygen species production, it is the aim of this master thesis to evaluate mitochondrial function in submandibular glands in a model of rats submitted to chronic hypobaric hypoxia during 3 months, in order to elucidate the mechanisms involved in glandular secretion at high altitude.Fil: Terrizzi, Antonela Romina. Universidad de Buenos Aires. Facultad de Farmacia y Bioquímica. Buenos Aires, Argentin

Altered production of reproductive neuropeptides in rats subjected to chronic intermittent hypoxia

Hypobaric hypoxia is a stressful condition known to decrease fertility both in humans and animals. However, the mechanism by which the hypothalamus-pituitary-gonad axis is altered remains unknown. The aim of the present study was to analyze the effects of chronic intermittent and continuous exposure to hypoxia on hypothalamic-pituitary-gonadal axis regulation in male rats. Thirty adult male Wistar rats were assigned to one of the following three groups: control group; chronic intermittent hypoxia: subjected to 600 mbar for 18 h/d five days a week; and chronic continuous hypoxia: subjected to 600 mbar for 23.5 hours/day seven days a week, for 30 days. Plasma luteinizing hormone and testosterone concentration, hypothalamic GnRh, Kiss1 and Rfrp3 mRNA levels and PGE2 content were determined. Levels of Rfrp3, a negative regulator of GnRH and LH release, were higher in intermittently exposed animals than in controls. Levels of Kiss1, a neuropeptide that stimulates the release of GnRH only increased in animals exposed to continuous hypoxia. Plasma luteinizing hormone and testosterone concentrations and body weight were lower in rats subjected to intermittent hypoxia as compared to the remaining groups. GnRh mRNA levels as well as PGE2 content remained unchanged in all groups. Taken together, results suggest that besides the well documented direct effects of hypoxia on the testes, infertility observed in male rats exposed to hypoxia may also be due to overexpression of negative regulators of GnRH and luteinizing hormone release. Intermittent, rather than continuous, to hypoxia exposure would seem to be more detrimental to fertility

The Process of Acclimation to Chronic Hypoxia Leads to Submandibular Gland and Periodontal Alterations: An Insight on the Role of Inflammatory Mediators

The exposition to hypoxia is a stressful stimulus, and the organism develops acclimation mechanisms to ensure homeostasis, but if this fails, it leads to the development of pathological processes. Considering the large number of people under hypoxic conditions, it is of utmost importance to study the mechanisms implicated in hypoxic acclimation in oral tissues and the possible alteration of some important inflammatory markers that regulate salivary and periodontal function. It is the aim of the present study to analyze submandibular (SMG) and periodontal status of animals chronically exposed to continuous (CCH) or intermittent (CIH) hypoxia in order to elucidate the underlying molecular mechanisms that may lead to hypoxic acclimation. Adult Wistar rats were exposed to CCH or CIH simulating 4200 meters of altitude during 90 days. Salivary secretion was decreased in animals exposed to hypoxia, being lower in CIH, together with increased prostaglandin E2 (PGE2) content, TBARS concentration, and the presence of apoptotic nuclei and irregular secretion granules in SMG. AQP-5 mRNA levels decreased in both hypoxic groups. Only the CCH group showed higher HIF-1α staining, while CIH alone exhibited interradicular bone loss and increased concentration of the bone resorption marker CTX-I. In summary, animals exposed to CIH show a worse salivary secretion rate, which related with higher levels of PGE2, suggesting a negative role of this inflammatory mediator during hypoxia acclimation. We link the weak immunorreactivity of HIF-1α in CIH with improper hypoxia acclimation, which is necessary to sustaining SMG physiology under this environmental condition. The alveolar bone loss observed in CIH rats could be due mainly to a direct effect of PGE2, as suggested by its higher content in gingival tissue, but also to the indirect effect of hyposalivation. This study may eventually contribute to finding therapeutics to treat the decreased salivary flow, improving in that way oral health

Deleterious effect of chronic continuous hypoxia on oral health

Objective: To evaluate the effect of chronic continuous hypoxia (CCH) in alveolar bone and its correlation with the inflammatory markers which play a key role in the development of periodontitis. Material and methods: Wistar rats were exposed to CCH (600 mbar, 3 months). Macroscopic and histological analyses of alveolar bone were performed, together with measurement of oxidative stress and inflammatory parameters in gums and submandibular glands (SMG). Results: HCC induced cortical alveolar bone loss, decreased interradicular bone volume and increased the periodontal ligament height compared to control rats (p < 0.05). CCH enhanced iNOS activity in gums (from 2735,04 ± 662,96 nmol/min/mg proteins to 4289,58 ± 915,63 p < 0.05) and in SMG (from 56,71 ± 12,05 nmol/min/mg proteins to 90,15 ± 21,78 p < 0.05). PGE2 did not change in gums or in SMG by means of CCH, while TNFα decreased in gums (p < 0.05). Regarding oxidative stress, thiobarbituric acid reactive species concentration in CCH animals was higher both in gums as in SMG, and catalase activity was decreased in SMG. Conclusion: Higher iNOS activity both in gums and SMG under CCH could be associated with the alveolar bone loss observed. The increase in oxidative stress occurring in SMG and gums, together with a lower antioxidant capacity might indicate a deleterious effect of HX in oral health.Fil: Terrizzi, Antonela Romina. Universidad de Buenos Aires. Facultad de Odontología. Cátedra de Fisiología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Fernández Solari, José Javier. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad de Buenos Aires. Facultad de Odontología. Cátedra de Fisiología; ArgentinaFil: Lee, Ching Ming. Universidad de Buenos Aires. Facultad de Odontología. Cátedra de Fisiología; ArgentinaFil: Conti, Maria Ines. Universidad de Buenos Aires. Facultad de Odontología. Cátedra de Fisiología; ArgentinaFil: Martinez, Maria del Pilar. Universidad de Buenos Aires. Facultad de Odontología. Cátedra de Fisiología; Argentin

Lead Intoxication under Environmental Hypoxia Impairs Oral Health

We have reported that chronic lead intoxication under hypoxic environment induces alveolar bone loss that can lead to periodontal damage with the subsequent loss of teeth. The aim of the present study was to assess the modification of oral inflammatory parameters involved in the pathogenesis of periodontitis in the same experimental model. In gingival tissue, hypoxia increased inducible nitric oxid synthase (iNOS) activity (p < .01) and meanwhile lead decreased prostaglandin E2 (PGE2) content (p < .05). In submandibular gland (SMG), iNOS activity was enhanced by lead and PGE2 content was increased by both lead and hypoxia (p < .01) and even more by combined treatments (p < .001). In the SMG, hypoxia stimulated angiogenesis (p < .01) with blood extravasation. Adrenal glands were 22% bigger in those animals exposed to lead under hypoxic conditions. Results suggest a wide participation of inflammatory markers that mediate alveolar bone loss induced by these environmental conditions. The lack of information regarding oral health in lead-contaminated populations that coexist with hypoxia induced us to evaluate the alteration of inflammatory parameters in rat oral tissues to elucidate the link between periodontal damage and these environmental conditions.Fil: Terrizzi, Antonela Romina. Universidad de Buenos Aires. Facultad de Odontología. Cátedra de Fisiología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay; ArgentinaFil: Fernández Solari, José Javier. Consejo Nacional de Investigaciones Científicas y Técnicas. Universidad de Buenos Aires; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay; ArgentinaFil: Lee, Ching Ming. Universidad de Buenos Aires. Facultad de Odontología. Cátedra de Fisiología; ArgentinaFil: Martinez, Maria del Pilar. Universidad de Buenos Aires. Facultad de Odontología. Cátedra de Fisiología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay; ArgentinaFil: Conti, María Inés. Universidad de Buenos Aires. Facultad de Odontología. Cátedra de Fisiología; Argentin

Effects of chronic lead exposure on bone mineral properties in femurs of growing rats

Lead exposure has been associated with several defective skeletal growth processes and bone mineral alterations. The aim of the present study is to make a more detailed description of the toxic effects of lead intoxication on bone intrinsic material properties as mineral composition, morphology and microstructural characteristics. For this purpose, Wistar rats were exposed (n = 12) to 1000 ppm lead acetate in drinking water for 90 days while control group (n = 8) were treated with sodium acetate. Femurs were examined using inductively coupled plasma optical emission spectrometry (ICP-OES), Attenuated Total Reflection Fourier transform infrared spectroscopy (ATR-FTIR), X-ray diffraction (XRD), and micro-Computed Tomography (µCT). Results showed that femur from the lead-exposed rats had higher carbonate content in bone mineral and (Ca2++Mg2++Na+)/P ratio values, although no variations were observed in crystal maturity and crystallite size. From morphological analyses, lead exposure rats showed a decreased in trabecular bone surface and distribution while trabecular thickness and cortical area increased. These overall effects indicate a similar mechanism of bone maturation normally associated to age-related processes. These responses are correlated with the adverse actions induced by lead on the processes regulating bone turnover mechanism. This information may explain the osteoporosis diseases associated to lead intoxication as well as the risk of fracture observed in populations exposed to this toxicant.Fil: Álvarez Lloret, Pedro. Universidad de Oviedo; EspañaFil: Lee, Ching Ming. Universidad de Buenos Aires. Facultad de Odontología. Cátedra de Fisiología; ArgentinaFil: Conti, Maria Ines. Universidad de Buenos Aires. Facultad de Odontología. Cátedra de Fisiología; ArgentinaFil: Terrizzi, Antonela Romina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Universidad de Buenos Aires. Facultad de Odontología. Cátedra de Fisiología; ArgentinaFil: González López, Santiago. Universidad de Granada; EspañaFil: Martinez, Maria del Pilar. Universidad de Buenos Aires. Facultad de Odontología. Cátedra de Fisiología; Argentin

Chronic Lead Exposure Alters Mineral Properties in Alveolar Bone

The objective of the present study was to investigate the effects of chronic lead exposure on the mineral properties of alveolar bone. For this purpose, female Wistar rats (n = 8) were exposed to 1000 ppm lead acetate in drinking water for 90 days, while the control group (n = 5) was treated with sodium acetate. The alveolar bone structure and chemical composition of the dissected man‐ dibles were examined using micro‐computed tomography (micro‐CT), scanning electron micros‐ copy (SEM), inductively coupled plasma optical emission spectrometry (ICP‐OES), attenuated total reflection Fourier transform infrared spectroscopy (ATR‐FTIR), and X‐ray diffraction (XRD) tech‐ niques to determine possible alterations in alveolar bone due to lead exposure. In addition, changes in bone mechanical properties were analysed using a three‐point bending test. Exposure to lead induced notable changes in bone mineralization and properties, specifically a reduction of the tra‐ becular thickness and bone mineral density. Furthermore, there was a reduction in carbonate con‐ tent and an increase in bone mineral crystallinity. These changes in bone mineralization could be explained by an alteration in bone turnover due to lead exposure. Three‐point bending showed a trend of decreased displacement at failure in the mandibles of lead‐exposed rats, which could com‐ promise the mechanical stability and normal development of the dentition.MICINN‐CGL2011‐25906 (Spain)UBACYT‐ 20020150100006BA (Argentina

Alveolar bone loss associated to periodontal disease in lead intoxicated rats under environmental hypoxia

Previously reported studies from this laboratory revealed that rats chronically intoxicated with lead (Pb) under hypoxic conditions (HX) impaired growth parameters and induced damages on femoral and mandibular bones predisposing to fractures. We also described periodontal inflammatory processes under such experimental conditions. Periodontitis is characterised by inflammation of supporting tissues of the teeth that result in alveolar bone loss. The existence of populations living at high altitudes and exposed to lead contamination aimed us to establish the macroscopic, biochemical and histological parameters consistent with a periodontal disease in the same rat model with or without experimental periodontitis (EP). Sixty female rats were divided into: Control; Pb (1000 ppm of lead acetate in drinking water); HX (506 mbar) and PbHX (both treatments simultaneously). EP was induced by placing ligatures around the molars of half of the rats during the 14 days previous to the autopsy. Hemi-mandibles were extracted to evaluate bone loss by histomorphometrical techniques. TNFα plasmatic concentration was greater (p < 0.01) in Pb and HX animals. TBA-RS content was significantly higher in gums of rats with or without EP only by means of Pb. The SMG PGE2 content increased by Pb or HX was higher in PbHX rats (p < 0.01). Pb and HX increased EP induced alveolar bone loss, while Pb showed spontaneous bone loss also. In conclusion, these results show that lead intoxication under hypoxic environment enhanced not only alveolar bone loss but also systemic and oral tissues inflammatory parameters, which could aggravate the physiopathological alterations produced by periodontal disease.Fil: Terrizzi, Antonela Romina. Universidad de Buenos Aires. Facultad de Odontología. Cátedra de Fisiología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Fernández Solari, José Javier. Universidad de Buenos Aires. Facultad de Odontología. Cátedra de Fisiología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Lee, Ching Ming. Universidad de Buenos Aires. Facultad de Odontología. Cátedra de Fisiología; ArgentinaFil: Bozzini, Clarisa. Universidad de Buenos Aires. Facultad de Odontología. Cátedra de Fisiología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Mandalunis, Patricia Mónica. Universidad de Buenos Aires. Facultad de Odontología. Cátedra de Histología y Embriología; ArgentinaFil: Elverdín, Juan Carlos. Universidad de Buenos Aires. Facultad de Odontología. Cátedra de Fisiología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Conti, Maria Ines. Universidad de Buenos Aires. Facultad de Odontología. Cátedra de Fisiología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Martinez, Maria del Pilar. Universidad de Buenos Aires. Facultad de Odontología. Cátedra de Fisiología; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentin