O tecido ósseo tem como característica estar constantemente em plena absorção e recomposição celular, sendo controlado pela interação de RANKL e OPG. No caso da perda de sua continuidade, ou seja, quando ocorre algum defeito ósseo, sua remodelação leva à restauração e consequente integridade do esqueleto, sendo o seu metabolismo influenciado por fatores hormonais, locais, comportamentais, ambientais e nutricionais; e seu desequilíbrio é um dos maiores obstáculos para a eficácia da remodelação óssea, podendo interferir negativamente na consolidação de fraturas. A ingestão crônica de álcool pode contribuir para esse desequilíbrio, e embora correlações significativas venham sendo relatadas entre o consumo excessivo de álcool e a consolidação óssea, novos estudos devem ser desenvolvidos haja vista a grande disparidade entre o tempo de submissão e a quantidade de ingestão do álcool englobando os diferentes protocolos. Os objetivos desse estudo foram avaliar quantitativamente os efeitos do consumo crônico de álcool no peso, reparo ósseo e densidade óssea em ratos Wistar, além de observarmos qualitativamente as fibras colágenas e a expressão de OPG e RANKL. Para isso, separamos aleatoriamente 30 ratos Wistar em dois grupos , sendo (G1) 15 ratos consumindo solução de aguardente diluída em água por 100 dias com concentração progressiva e controlada (10ºGL, 15ºGL, 20ºGL, 25ºGL e 30ºGL) e 15 ratos não alcoólatras consumindo como dieta líquida somente água (G2). Após o 92º dia do período de indução do alcoolismo, ambos os grupos foram submetidos a um defeito ósseo realizado na tíbia com um motor rotacional contendo uma broca com tamanho de 3 mm de diâmetro. Após 8 dias após o procedimento cirúrgico os animais foram eutanasiados em câmara de C02, as tíbias foram removidas e aparadas nas proximidades do defeito ósseo para que fossem processadas através de secções histológicas descalcificadas. A porcentagem de osso neoformado e a densidade óssea foram avaliadas histometricamente. Através da imunohistoquimica observamos a expressão de OPG e RANKL e através de reação histoquímica pelo método Picro-sirius Red, estudamos o padrão de birrefringência das fibras colágenas. Como resultados, pudemos observar que o peso dos animais, a densidade e remodelação ósseas foram menores no grupo alcoólico. Encontramos também efeitos negativos em relação à qualidade e organização das fibras colágenas, bem como diferenciação na expressão de RANKL e OPG nos diferentes grupos. Nossos resultados demonstram que o protocolo proposto de ingestão de álcool exerce efeitos negativos no ganho de peso e qualidade óssea quando comparado ao grupo controle. / The osseous tissue features the continuous cellular absorption and recomposition regulated by the interaction of the Receptor activator of nuclear factor kappa- β ligand (RANKL) and Osteoprotegerin (OPG). In case its continuity is lost, that is, any kind of bone injury, its remodeling leads to restoration and consequent skeletal integrity and its metabolism is influenced by hormonal, local, behavioral, environmental and nutritional factors; its imbalance is one of the biggest obstacles to the efficiency of bone remodeling, and it might interfere negatively with fracture healing. The chronic consumption of alcohol may contribute to this imbalance, and, although significant correlations between the excessive alcohol intake and bone consolidation have been reported, new studies must be developed considering the great variety of protocols which approach the time of exposure and quantity of alcohol intake. The objective of this study was to verify quantitatively the effects of chronic alcohol consumption on body weight, bone healing and density in rats. It was also observed quantitatively the collagenous fibers and the expression of OPG and RANKL. For this purpose, 30 Wistar rats were randomly divided into two groups: G1 (group 1) consisted in 15 rats which had, for 100 days, a liquid diet of liquor diluted in water with a progressive and controlled concentration (10°GL, 15°GL, 20°GL, 25°GL and 30°GL); G2 (group 2) consisted in 15 rats on a liquid diet of only water and free of alcohol. After the 92nd day of the induction period of alcoholism, tibial defects of 3 mm in diameter were created in both groups and after 8 days from this surgery procedure the animals were euthanized in a CO2 chamber. The tibiae were removed and cut next to the bone defect in order to be processed through decalcified histological sections. The percentage of renovated bone and osseous density were submitted to histometric analysis. Through immunohistochemistry, the expression of OPG and RANKL were analyzed and using Picrosirius red staining method, the birefringence pattern of the collagen fibers was studied. As a result, it was verified that the body weight of the animals, the osseous density and bone remodeling were smaller in G1; negative effects regarding to the collagen fibers quality and organization and a differentiation in the expression of OPG and RANKL were also found in both groups. Conclusion: these results show that the proposed protocol of alcohol intake produces negative effects in the gain of body weight and bone quality when compared to the control group.

Abstract Persistent organic pollutants (POPs) are ubiquitous and bioaccumulative man-made chemicals, resistant to chemical, biological and photolytic degradation and widely distributed to sediments, wildlife, and human. Many of these chemicals have adverse effects on a variety of targets, including the endocrine system, organogenesis and reproduction. Due to these effects and wide distribution, many of them are either banned or strictly controlled. However, because of persistency, they continue to interact with organisms globally. Despite the existing knowledge of the adverse effects of POPs, the effects of many chemicals on bone tissue are still poorly known. In the present study, we investigated the adverse effects of three common POPs, including tributyltin (TBT), 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and perfluorooctanoic acid (PFOA) on the skeletal system. In vitro models were used to study the effects of PFOA in mouse and in human, and the co-effects of TBT and TCDD on differentiating osteoblasts and osteoclasts of mice. An in vivo model for mice was used to study the developmental effects of maternal PFOA-exposure on pups among with morphometrical and biomechanical property analyses. Mass-spectrometry was used to study the presence of PFOA in bones both in mice and in human, the latter acquired from the bone bank held in the Oulu University Hospital, Finland. The bones were also analyzed with cone beam computer tomography and microcomputer tomography. The results show that PFOA exposure in utero and during lactation leads to the accumulation of PFOA in bone, traceable even 17 months after exposure. PFOA exposure decreased the mineral density of the tibias and increased the medullary area. Nearly all of the human samples contained PFAS, including PFOA. PFOA also disturbed the differentiation of osteoblasts and with lower doses, increased bone resorption of osteoclasts both in mouse and human, the phenomenon being slightly stronger in mice. Co-exposure to TBT and TCDD led to decreased differentiation of osteoblasts and osteoclast, and the co-effect was partially synergistic in osteoblasts. These results show disruption of bone development, bone cell differentiation, and PFAS accumulation in bone. Further studies are recommended to evaluate the co-effects of different POPs and the possible effects of long-term accumulation of POPs in bone and other tissues. / Tiivistelmä Pysyvät orgaaniset ympäristömyrkyt (POP-yhdisteet) ovat kemikaaleja, jotka ovat levinneet ihmisen toiminnan seurauksena laajalle ympäristöön, sen eliöihin ja ihmisiin. Monilla POP-yhdisteillä on haitallisia vaikutuksia esimerkiksi hormonaaliseen toimintaan, elinten muodostukseen ja hedelmällisyyteen. Toksisten vaikutusten ja niiden yleisyyden vuoksi monien POP-yhdisteiden käyttö on joko rajattua tai kielletty kokonaan. Laajan levinneisyytensä ja hitaan puoliintumisaikansa takia POP-yhdisteet ovat kuitenkin edelleen vuorovaikutuksessa ympäristön ja sen eliöiden kanssa. POP-yhdisteiden luustovaikutuksista tiedetään edelleen vähän. Tässä väitöskirjassa tutkittiin kolmen yleisen POP-yhdisteen, tributyylitinan (TBT), 2,3,7,8-tetraklooridibentso-p-dioksiinin (TCDD) ja perfluoro-oktaanihapon (PFOA), vaikutuksia luustoon. PFOA:n vaikutuksia hiiren ja ihmisen luustoon sekä TBT:n ja TCDD:n yhteisvaikutuksia hiiren erilaistuvien osteoblastien ja osteoklastien suhteen selvitettiin in vitro -malleilla. In vivo -mallilla tutkittiin hiiriemon PFOA-altistuksen vaikutusta syntyvien poikasten luuston kehitykseen ja remodelaatioon analysoimalla poikkileikekuvia sekä luiden biomekaanisia ominaisuuksia. Lisäksi luiden PFOA-pitoisuudet mitattiin massaspektrometrilla. Tutkimusta laajennettiin ihmiseen analysoimalla Oulun yliopistollisen sairaalan luupankkinäytteitä. Ihmisnäytteet analysoitiin myös kartiokeila-TT:n ja mikro-TT:n avulla. Tulosten mukaan PFOA kertyy luuhun; hiiriltä voitiin mitata PFOA-pitoisuuksia jopa 17 kuukautta altistumisen jälkeen. Lisäksi PFOA-altistus pienensi luun mineraalitiheyttä ja kasvatti luuydinontelon tilavuutta. Lähes kaikki ihmisluunäytteet sisälsivät PFOA:ta ja muita PFAS-yhdisteitä. Solukokeiden perusteella PFOA-altistus häiritsee osteoblastien erilaistumista ja pienillä pitoisuuksilla lisää osteoklastien luunhajotusta sekä hiirellä että ihmisellä. TBT:n ja TCDD:n yhteisaltistus vaikuttaa puolestaan vähentävän sekä osteoblastien että osteoklastien erilaistumista ja toimintaa; osteoblastien osalta yhteisvaikutus oli osaksi synergistinen. Väitöskirja antaa lisätietoa POP-yhdisteiden vaikutuksista luun kehitykseen ja luusolujen erilaistumiseen sekä PFAS-yhdisteiden kertymisestä luuhun. Väitöksessä myös suositellaan lisätutkimuksia yhdisteiden yhteisvaikutuksista sekä pitkän aikavälin ympäristökemikaalikertymän vaikutuksista luussa ja muissa kudoksissa.

No description available.

Mathematical modeling is an important tool in improving the understanding of complex biological processes. However, mathematical models are often faced with challenges that arise due to the limited knowledge of the underlying biological processes and the high number of parameters for which exact values are unknown. The method of generalized modeling is an alternative modeling approach that aims to address these challenges by extracting information about stability and bifurcations of classes of models while making only minimal assumptions on the specific functional forms of the model. This is achieved by a direct parameterization of the Jacobian in the steady state, introducing a set of generalized parameters which have a biological interpretation. In this thesis, the method of generalized modeling is extended and applied to different problems from cell biology. In the first part, we extend the method to include also the higher derivatives at the steady state. This allows an analysis of the normal form of bifurcations and thereby a more specific description of the nearby dynamics. In models of gene-regulatory networks, it is shown that the extended method can be applied to better characterize oscillatory systems and to detect bistable dynamics. In the second part, we investigate mathematical models of bone remodeling, a process that renews the human skeleton constantly. We investigate the connection between structural properties of mathematical models and the stability of steady states in different models. We find that the dynamical system operates from a stable steady state that is situated in the vicinity of bifurcations where stability can be lost, potentially leading to diseases of bone. In the third part of this thesis, models of the MAPK signal transduction pathway are analyzed. Since mathematical models for this system include a high number of parameters, statistical methods are employed to analyze stability and bifurcations. Thereby, the parameters with a strong influence on the stability of steady states are identified. By an analysis of the bifurcation structure of the MAPK cascade, it is found that a combination of multiple layers in a cascade-like way allows for additional types of dynamic behavior such as oscillations and chaos. In summary, this thesis shows that generalized modeling is a fruitful alternative modeling approach for various types of systems in cell biology. / Mathematische Modelle stellen ein wichtiges Hilfmittel zur Verbesserung des Verständnisses komplexer biologischer Prozesse dar. Sie stehen jedoch vor Schwierigkeiten, wenn wenig über die zugrundeliegende biologischen Vorgänge bekannt ist und es eine große Anzahl von Parametern gibt, deren exakten Werte unbekannt sind. Die Methode des Verallgemeinerten Modellierens ist ein alternativer Modellierungsansatz mit dem Ziel, diese Schwierigkeiten dadurch anzugehen, dass dynamische Informationen über Stabilität und Bifurkationen aus Klassen von Modellen extrahiert werden, wobei nur minimale Annahmen über die spezifischen funktionalen Formen getätigt werden. Dies wird erreicht durch eine direkte Parametrisierung der Jacobimatrix im Gleichgewichtszustand, bei der neue, verallgemeinerte Parameter eingeführt werden, die eine biologische Interpretation besitzen. In dieser Arbeit wird die Methode des Verallgemeinerten Modellierens erweitert und auf verschiedene zellbiologische Probleme angewandt. Im ersten Teil wird eine Erweiterung der Methode vorgestellt, bei der die Analyse höherer Ableitungen im Gleichgewichtszustand integriert wird. Dies erlaubt die Bestimmung der Normalform von Bifurkationen und hierdurch eine spezifischere Beschreibung der Dynamik in deren Umgebung. In Modellen für genregulatorische Netzwerke wird gezeigt, dass die so erweiterte Methode zu einer besseren Charakterisierung oszillierender Systeme sowie zur Erkennung von Bistabilität verwendet werden kann. Im zweiten Teil werden mathematische Modelle zur Knochenremodellierung untersucht, einem Prozess der das menschliche Skelett kontinuierlich erneuert. Wir untersuchen den Zusammenhang zwischen strukturellen Eigenschaften verschiedener Modelle und der Stabilität von Gleichgewichtszuständen. Wir finden, dass das dynamische System von einem stabilen Zustand operiert, in dessen Nähe Bifurkationen existieren, welche das System destabilisieren und so potentiell Knochenkranheiten verursachen können. Im dritten Teil werden Modelle für den MAPK Signaltransduktionsweg analysiert. Da mathematische Modelle für dieses System eine hohe Anzahl von Parametern beinhalten, werden statistische Methoden angewandt zur Analyse von Stabilität und Bifurkationen. Zunächst werden Parameter mit einem starken Einfluss auf die Stabilität von Gleichgewichtszuständen identifizert. Durch eine Analyse der Bifurkationsstruktur wird gezeigt, dass eine kaskadenartige Kombination mehrerer Ebenen zu zusätzliche Typen von Dynamik wie Oszillationen und Chaos führt. Zusammengefasst zeigt diese Arbeit, dass Verallgemeinertes Modellieren ein fruchtbarer alternativer Modellierungsansatz für verschiedene zellbiologische Probleme ist.

The purpose of this study is to further quantify adult ovine ovariectomized bone for new remodeling characteristics to obtain a better understanding of how remodeling is occurring and the effectiveness of this animal model for the study of postmenopausal osteoporosis. Postmenopausal osteoporosis is a major health concern and animal models to test new treatment options are needed. The ovine model is a good option because the ewes undergo Haversian remodeling, are a large sized animal, and have a similar hormone profile to humans. Ewes, however, do not undergo a natural menopause, so an ovariectomy surgery was conducted in the sheep to simulate the decreased levels in estrogen. Columbia-Rambouillet sheep were used in this study: some that have been ovariectomized as a model for postmenopausal osteoporosis and some that underwent a sham surgery to serve as a control. The sheep were sacrificed 12 months post operatively in the month of August, so the seasonal effects of remodeling were accounted for. The left radius was then processed into microradiographs of 6 regional cortical beams, where the cranial (tensile side) and caudal (compressive side) anatomical sections were analyzed in this study to determine regional differences in remodeling. Previous students’ theses have analyzed the similar samples for basic bone remodeling histology measurements, resulting in some significant seasonal, anatomical, and treatment differences. However, most of the results showed no particular increase in the amount of remodeled area for the ovariectomized sheep compared to the sham sheep, even though an ovariectomy is believed to cause a burst of remodeling in bone due to the decreased levels in estrogen. In this study, a new repeatable method was developed that further examines secondary bone by quantifying the extent to which secondary osteons encroach on previously-existing secondary osteons. Encroached and unencroached secondary osteons were quantified using two different methods: a point count method that measured the percentage of the area the encroached and unencroached secondary osteons inhabited and an osteon count method that measured the number of encroached and unencroached secondary osteons per area. These raw measurements were calculated into 18 parameters and 2-way repeated measures ANOVAs were run to determine the effects of surgery and anatomical region on each of the bone remodeling parameters. The results found significant effects from estrogen deletion which were different depending on if the bone region was predominately in compression or tension. The ovariectomy surgery caused an increase in remodeling, which was mostly confined on the compressive side to areas that have been previously remodeled, but on the tensile side, bone remodeling expanded into areas that used to be primary bone. The new secondary osteons, as a result of the ovariectomy surgery, were larger than in the control animals. There however, was not an increase in porosity from the ovariectomy surgery, which is one of the main characteristics of osteoporosis. The model could be further studied to determine what sheep are doing that prevents them from losing bone and that knowledge could be greatly beneficial for human treatment plans of postmenopausal osteoporosis.

No description available.

No description available.

BACKGROUND: Bone remodeling relies on the tightly regulated interplay between bone forming osteoblasts and bone digesting osteoclasts. Several studies have now described the molecular mechanisms by which osteoblasts control osteoclastogenesis and bone degradation. It is currently unclear whether osteoclasts can influence bone rebuilding. METHODOLOGY/PRINCIPAL FINDINGS: Using in vitro cell systems, we show here that mature osteoclasts, but not their precursors, secrete chemotactic factors recognized by both mature osteoblasts and their precursors. Several growth factors whose expression is upregulated during osteoclastogenesis were identified by DNA microarrays as candidates mediating osteoblast chemotaxis. Our subsequent functional analyses demonstrate that mature osteoclasts, whose platelet-derived growth factor bb (PDGF-bb) expression is reduced by siRNAs, exhibit a reduced capability of attracting osteoblasts. Conversely, osteoblasts whose platelet-derived growth factor receptor beta (PDGFR-beta) expression is reduced by siRNAs exhibit a lower capability of responding to chemotactic factors secreted by osteoclasts. CONCLUSIONS/SIGNIFICANCE: We conclude that, in vitro mature osteoclasts control osteoblast chemotaxis via PDGF-bb/PDGFR-beta signaling. This may provide one key mechanism by which osteoclasts control bone formation in vivo.

Introdução: Diabetes mellitus (DM) é uma desordem metabólica associada a diversas alterações sistêmicas e uma das características é afetar o metabolismo ósseo. As modificações teciduais induzidas pela força ortodôntica estão relacionadas à sua remodelação por ativação da reabsorção óssea alveolar no lado de pressão e conseqüente aposição óssea no lado de tração. Objetivo: avaliar o processo de remodelação do periodonto de sustentação através da análise imunohistoquímica e histológica após a aplicação da expansão rápida da maxila (ERM) em ratos sob estado diabético. Material e Métodos: Noventa ratos Wistar, machos, foram distribuídos aleatoriamente em seis grupos de estudo, contendo cada um, 15 animais: grupo C: animais não-diabéticos e sem expansão rápida da maxila (ERM); grupo DM: animais com diabetes mellitus (DM) induzidos por Streptozotocina (STZ); grupo ERM: animais com ERM; grupo DM+ERM: animais com DM + ERM; grupo DM+INS: animais com diabetes mellitus e tratados com insulina (INS); grupo DM+INS+ERM: animais com diabetes mellitus, tratados com insulina e realizada a ERM. Os animais foram submetidos à eutanásia 3, 7 ou 10 dias após a ERM. Foram realizadas análises histológicas qualitativas e imunoistoquímicas para avaliar a expressão da tríade protêica (TRAP, RANKL e OPG). Para as reações imunoistoquímicas foram realizadas análise qualitativa e semi quantitativa através da atribuição de scores para a intensidade da expressão das proteínas analisadas. Nos dados não paramétricos, foi utilizado o teste de Kruskal-Wallis e pós-teste de Dunn nas comparações (p0,05). Resultado: Em relação aos aspectos histológicos houve alteração na disposição das fibras colágenas, morfologia dos fibroblastos, quantidade de vasos sanguíneos e quantidade de tecido ósseo neo-formado caracterizado pelas linhas incrementais nos grupos ERM, DM+ERM e DM+INS+ERM comparado com seus respectivos grupos controle. Qualitativamente, a neoformação do osso alveolar dos ratos diabéticos foi menor quando comparado com os normais e, a alteração sistêmica diabetes melittus promoveu redução significativa na taxa de formação e maturação óssea. O grupo ERM mostrou os maiores valores para indicadores de osteoclastogênese (TRAP) nos períodos iniciais sendo que nos grupos diabéticos (DM+ERM e DM+INS+ERM), essa expressão foi maior nos períodos finais do experimento. O mesmo comportamento foi verificado com as proteínas de remodelação e neo-formação óssea (RANKL e OPG) óssea. Conclusão: A diabetes mellitus atrasou o processo de reparo do periodonto de sustentação e alterou o turnover ósseo. A insulina utilizada para o controle do diabetes melhoraram as respostas, mas não restabeleceram completamente os valores iniciais do grupo controle. Porém, apesar das diferenças qualitativas nas respostas teciduais do periodonto, a condição diabética em estado inicial mostrou pouco impacto clínico sobre o procedimento de movimentação dentária induzida / Introduction: Diabetes mellitus (DM) is a metabolic disorder associated with several systemic changes and one of the characteristics is to affect bone metabolism. Tissue modifications induced by orthodontic force are related to its remodeling by activation of alveolar bone resorption on the pressure side and consequent bone apposition on the traction side. Objective: To evaluate the remodeling process of the support periodontium through immunohistochemical and histological analysis after the application of rapid maxillary expansion (RME) in rats on diabetic state. Material and Methods: Ninety male Wistar rats were randomly assigned to six study groups, each containing 15 animals: GC: non-diabetic animals without rapid maxillary expansion (RME); GDM: animals with diabetes mellitus (DM) induced by Streptozotocin (STZ); GRME: animals with RME; GDM+RME: animals with DM + RME; GDM+INS: animals with DM, treated whit insulin; GDM+INS+RME: animals with DM, treated with insulin and RME. The animals were submitted to euthanasia 3, 7 or 10 days after RME. Qualitative and immunohistochemical histological analyzes were performed to evaluate the expression of the protein triad (TRAP, RANKL and OPG). For the immunohistochemical reactions, qualitative and semi quantitative analysis were performed through the assignment of scores for the expression intensity of the analyzed proteins. The non-parametric data were used the Kruskal-Wallis test and Dunn post-test in the comparisons (p<0.05). Results: In relation to the histological aspects, there was alteration in the collagen fibers, fibroblast morphology, number of blood vessels and quantity of neoformed bone tissue characterized by the incremental lines in the RME, DM+RME and DM+INS+RME groups, compared to their respective control groups. Qualitatively, the neoformation of the alveolar bone of the diabetic rats was smaller when compared to the normal ones, and the systemic alteration diabetes melittus promoted a significant reduction in the formation rate and bone maturation. The RME group showed the highest values for indicators of osteoclastogenesis (TRAP) in the initial periods, and this expression was higher in the diabetic groups (DM+RME and DM+INS+RME) in the final periods of the experiment. The same behavior was verified with bone remodeling and neoformation (RANKL and OPG) proteins. Conclusion: Diabetes mellitus delayed the periodontal repair process and altered bone turnover. Insulin used to control diabetes improved responses but did not completely restore control group initial values. However, in spite of the qualitative differences in the tissue responses of the periodontium, the diabetic condition in initial state showed little clinical impact on the induced tooth movement procedure

The Keap1-Nrf2 pathway regulates a wide range of cytoprotective genes, and has been found to serve a protective and beneficial role in many body systems. There is limited information available, however, about its role in bone homeostasis. While Nrf2 activation has been suggested as an effective method of increasing bone mass and quality, there have been conflicting reports which associate Keap1 deficiency with detrimental phenotypes. As Keap1 deletion is a common method of Nrf2 activation, further study should address the impacts of various methods of regulating Nrf2 expression. Also, little research has been conducted on the specific pathways by which Nrf2 activation improves bone quality. In this study, the effects of alterations to Nrf2 activation levels were explored in two specific and varied scenarios. In the first experiment, moderate Nrf2 activation was achieved via partial deletion of its sequestering protein, Keap1, in an aging mouse model. The hypothesis tested here is that moderate Nrf2 activation improves bone quality by affecting bone metabolism and response to mechanical loading. The results of this first experiment suggest a subtle, sex-specific effect of moderate Nrf2 activation in aging mice which improves specific indices of bone quality to varying degrees, but does not affect loading-induced bone formation. It is likely that the overwhelming phenotypic impacts associated with aging or the systemic effects of global Keap1 deficiency may increase the difficulty in parsing out significant effects that can be attributed solely to Nrf2 activation. In the second experiment, a cell-specific knockout of Nrf2 in the osteocytes was achieved using a Cre/Lox breeding system. The hypothesis tested here is that osteocyte-specific deletion of Nrf2 impairs bone quality by affecting bone metabolism and response to mechanical loading. The results of this experiment suggest an important role of Nrf2 in osteocyte function which improves certain indices of bone quality, which impacts male and female bones in different 7 ways, but did not significantly impact loading-induced bone formation. Further studies should modify the method of Nrf2 activation in an effort to refine the animal model, allowing the effects of Nrf2 to be isolated from the potential systemic effects of Keap1 deletion. Future studies should also utilize other conditional knockout models to elucidate the effects of Nrf2 in other specific cell types.