INTRODUÇÃO: O consumo mundial de açúcar triplicou nos últimos 50 anos e a sua ingesta abusiva é responsável pela resistência periférica à insulina, que origina a síndrome metabólica - obesidade, diabetes melito, hipertensão arterial e doenças coronarianas . Motivados pelo elevado número de pacientes que nos procuram com queixas vestibulares associadas aos distúrbios de metabolismo da glicose (DMG) resolvemos avaliar de forma objetiva, a influência dos DMG nas disfunções labirínticas e o efeito da dieta restritiva de carboidratos como forma de tratamento. OBJETIVO: Observar o impacto da dieta fracionada e restritiva de carboidratos na qualidade de vida dos pacientes portadores de distúrbios do equilíbrio corporal e DMG por meio da posturografia dinâmica computadorizada (PDC), do disability index (DI) e da escala análogo-visual (EAV). CASUÍSTICA E METODOLOGIA: Este estudo foi desenhado como um ensaio clínico prospectivo controlado randomizado realizado no Hospital das Clínicas da Faculdade de Medicina da Universidade de São Paulo. A amostra foi constituída de 51 pacientes divididos em dois grupos: Grupo Dieta (GD): indivíduos tratados com comprimidos de placebo e dieta fracionada com restrição de glicose, Grupo Controle (GC): receberam apenas placebo. Os pacientes realizaram PDC, DI e EAV no primeiro e trigésimo dias do estudo. RESULTADOS: A amostra mostrou-se homogêna quando comparados os grupos e observou-se melhora, estatísticamente comprovada nas condições posturográficas avaliadas quando comparados GD e GC. Observou-se ainda melhora clínica do GD na análise da EAV. CONCLUSÃO: A dieta fracionada e restritiva de carboidratos mostrou-se eficaz no tratamento da nossa amostra de pacientes portadores de disfunções vestibulares associadas a DMG. / INTRODUCTION: World sugar consumption has tripled in the last 50 years and its abusive ingestion is responsible for peripheral insulin resistance, which leads to metabolic syndrome - obesity, diabetes mellitus, hypertension and coronary heart disease. Because of the high number of patients with vestibular complaints and with glucose metabolism disorders (GMD) we decided to objectively evaluate the effect of glucose restrictive and fractionated diet as a option of treatment in these patients. OBJECTIVE: To evaluate the impact of the glucose restrictive and fractionated diet on the Computerized Dynamic Posturography (CDP), disability index (DI) and the visual analogue scale (VAS) in patients with balance disorders and disorders of glucose metabolism. SAMPLES AND METHODOLOGY: Randomized controlled trial. Sample of 51 patients divided into two groups: Diet Group (DG) treated with placebo pills and glucose restrictive and fractionated diet and Control Group (CG) with only placebo. The individuals performed CDP, DI and VAS at first and thirtieth days of study. RESULTS: The sample groups were homogeneous before the study. There were significant improvement of DG on CDP conditions 4, 5, 6 and composite score. There was, also, significant improvement of VAS analysis on DG after intervention. CONCLUSION: The glucose restrictive and fractionated diet was effective in the treatment of patients with vestibular dysfunction associated with glucose metabolism disorders

Indiana University-Purdue University Indianapolis (IUPUI) / Glucose homeostasis is maintained through the coordinated actions of insulin secretion from pancreatic beta cells and insulin action in peripheral tissues. Dysfunction of insulin action yields insulin resistance, and when coupled with altered insulin secretion, results in type 2 diabetes (T2D). Exocytosis of intracellular vesicles, such as insulin granules and glucose transporter (GLUT4) vesicles is carried out by similar SNARE (soluble NSF attachment receptor) protein isoforms and Munc18 proteins. An additional regulatory protein, Doc2b, was implicated in the regulation of these particular exocytosis events in clonal cell lines, but relevance of Doc2b in the maintenance of whole body glucose homeostasis in vivo remained unknown. The objective of my doctoral work was to delineate the mechanisms underlying regulation of insulin secretion and glucose uptake by Doc2b in effort to identify new therapeutic targets within these processes for the prevention and/or treatment of T2D. Towards this, mice deficient in Doc2b (Doc2b-/- knockout mice) were assessed for in vivo alterations in glucose homeostasis. Doc2b knockout mice were highly susceptible to preclinical T2D, exhibiting significant whole-body glucose intolerance related to insulin secretion insufficiency as well as peripheral insulin resistance. These phenotypic defects were accounted for by defects in assembly of SNARE complexes. Having determined that Doc2b was required in the control over whole body glycemia in vivo, whether Doc2b is also limiting for these mechanisms in vivo was examined. To study this, novel Doc2b transgenic (Tg) mice were engineered to express ~3 fold more Doc2b exclusively in pancreas, skeletal muscle and fat tissues. Compared to normal littermate mice, Doc2b Tg mice had improved glucose tolerance, related to concurrent enhancements in insulin mumsecretion from beta cells and insulin-stimulated glucose uptake in the skeletal muscle. At the molecular level, Doc2b overexpression promoted SNARE complex assembly, increasing exocytotic capacities in both cellular processes. These results unveiled the concept that intentional elevation of Doc2b could provide a means of mitigating two primary aberrations underlying T2D development.

Indiana University-Purdue University Indianapolis (IUPUI) / Diabetes mellitus is a group of related metabolic diseases that share a common pathological mechanism: insufficient insulin signaling. Insulin is a hormone secreted from pancreatic β cells that promotes energy storage and consequently lowers blood glucose. In contrast, the hormone glucagon, released by pancreatic α cells, plays a critical complementary role in metabolic homeostasis by releasing energy stores and increasing blood glucose. Restoration of β cell mass in diabetic patients via β cell regeneration is a conceptually proven approach to finally curing diabetes. Moreover, in situ regeneration of β cells from endogenous sources would circumvent many of the obstacles encountered by surgical restoration of β cell mass via islet transplantation. Regeneration may occur both by β cell self-duplication and by neogenesis from non-β cell sources. Although the mechanisms regulating the β cell replication pathway have been highly investigated, the signals that regulate β cell neogenesis are relatively unknown. In this dissertation, I have used zebrafish as a genetic model system to investigate the process of β cell neogenesis following insulin signaling depletion by various modes. Specifically, I have found that after their ablation, β cells primarily regenerate from two discrete cellular sources: differentiation from uncommitted pancreatic progenitors and transdifferentiation from α cells. Importantly, I have found that insulin and glucagon play crucial roles in controlling β cell regeneration from both sources. As with metabolic regulation, insulin and glucagon play counter-balancing roles in directing endocrine cell fate specification. These studies have revealed that glucagon signaling promotes β cell formation by increasing differentiation of pancreas progenitors and by destabilizing α cell identity to promote α to β cell transdifferentiation. In contrast, insulin signaling maintains pancreatic progenitors in an undifferentiated state and stabilizes α cell identity. Finally, I have shown that insulin also regulates pancreatic exocrine cell development. Insufficient insulin signaling destabilized acinar cell fate and impairs exocrine pancreas development. By understanding the roles of pancreatic hormones during pancreas development and regeneration can provide new therapeutic targets for in vivo β cell regeneration to remediate the devastating consequences of diabetes.

Indiana University-Purdue University Indianapolis (IUPUI) / Signal Transducers and Activators of Transcription 3 (STAT3), a transcription factor expressed in many cell types, including osteoblasts and osteoclasts, is emerging as a key regulator of bone mass and strength. STAT3 mutations cause a rare human immunodeficiency disease characterized by extremely elevated levels of IgE in serum that have associated craniofacial and skeletal features, such as reduced bone mineral density and recurrent pathological fractures. Our microarray data and immunohistochemical staining using a normal rat model have shown that STAT3 mRNA and protein levels markedly increase in response to mechanical loading. In addition, as indicated by STAT3 phosphorylation in MC3T3-E1 osteoblastic cells, STAT3 activity significantly increases in response to 30 to 90 minutes fluid shear stress. In order to further study the role that STAT3 plays in bone responsiveness to loading, tissue-selective STAT3 knockout (KO) mice, in which inactivation of STAT3 occurs in osteoblasts, were generated by breeding the transgenic mice in which Cre recombinase cDNA was cloned downstream of a 3.6 or 2.3 kb fragment of the rat Col1a1 promoter (Col3.6-Cre and Col2.3-Cre, respectively) with a strain of floxed mice in which the two loxP sites flank exons 18-20 of the STAT3 gene were used. Mice engineered with bone selective inactivation of STAT3 in osteoblasts exhibited significantly lower bone mineral density (7-12%, p<0.05) and reduced ultimate force (21-34%, p<0.01) compared to their age-matched littermate controls. The right ulnae of 16-week-old bone specific STAT3 KO mice and the age-matched control mice were loaded with peak forces of 2.5 N and 2.75 N for female and male mice, respectively, at 2 Hz, 120 cycles/day for 3 consecutive days. Mice with inactivation of STAT3 specific in bone were significantly less responsive to mechanical loading than the control mice as indicated by decreased relative mineralizing surface (rMS/BS, 47-59%, p<0.05) and relative bone formation rate (rBFR/BS, 64-75%, p<0.001). Bone responsiveness was equally decreased in mice in which STAT3 is inactivated either in early osteoblasts (Col3.6-Cre) or in mature osteoblasts (Col2.3-Cre). Accumulating evidence indicates that bone metabolism is significantly affected by activities in mitochondria. For instance, although STAT3 is reported to be involved in bone formation and resorption through regulation of nuclear genes, inactivation of STAT3 is shown to disrupt mitochondrial activities and result in an increased level of reactive oxygen species (ROS). Inactivation of STAT3 suppressed load-driven mitochondrial activity, which led to an elevated level of ROS in cultured primary osteoblasts. Oxidative stress induced by administration of buthionine sulfoximine (BSO) significantly inhibits load-induced bone formation in wild type mice. Taken together, the results support the notion that the loss-of-function mutation of STAT3 in osteoblasts and osteocytes diminishes load-driven bone formation and impairs the regulation of oxidative stress in mitochondria.

INTRODUÇÃO: A esteato-hepatite não alcóolica (NASH) é considerada problema de saúde pública, dada sua crescente incidência e seu possível papel na carcinogênese hepato-celular. Terapias atuais envolvem alterações de dieta e estilo de vida, mas têm seu resultado prejudicado pela baixa aderência dos pacientes. Abordagens farmacológicas ainda são precárias. Uma grande dificuldade no manejo de NASH reside no limitado entendimento de sua fisiopatologia, que parece envolver complexas alterações metabólicas e inflamatórias. Ácidos graxos poli-insaturados ômega-3 (AGPIs n-3) são reconhecidos por suas propriedades moduladoras do metabolismo lipídico e da inflamação, e estão diminuídos em pacientes com NASH. O uso clínico de AGPIs n-3 tem mostrado benefício no controle da esteatose e na produção de marcadores da resposta metabólica e inflamatória em NASH, embora com algumas observações contraditórias. A compreensão de mecanismos moleculares modulados por AGPIs n-3 em NASH podem ser úteis para identificar alvos moleculares que auxiliem no desenho de intervenção farmacológica efetiva. Nesse sentido, ciências ômicas são particularmente úteis para a compreensão de mecanismos moleculares com alto valor translacional para a prática clínica e podem contribuir para a identificação desses alvos. OBJETIVO: O presente estudo avaliou a resposta proteômica hepática e lipidômica plasmática de pacientes com NASH perante o tratamento com AGPIs n-3. MÉTODO: As avaliações proteômicas e lipidômicas foram desenvolvidas por espectometria de massas e/ou cromatografia gasosa em amostras de biópsias hepáticas e plasma coletadas de pacientes envolvidos em estudo preliminar, realizado no Hospital das Clínicas da Faculdade de Medicina da Universidade de São Paulo. O referido estudo envolveu pacientes adultos, de ambos os sexos e com diagnóstico de NASH tratados diariamente, durante 6 meses, com 3 cápsulas contendo mistura de óleo de linhaça e óleo de peixe [0,315 g AGPIs: sendo 0,065 g de ácido eicosapentaenoico (EPA), 0,050 g de docosahexaenoico (DHA) e 0,2 g alfa linolênico (ALA) por cápsula]. Pacientes, após o tratamento com AGPIs n-3, que apresentaram altas concentrações plasmáticas de ALA e/ou DHA e/ou baixas de ácido araquidônico (AA) mostraram melhora parcial das alterações de histologia hepática. No presente estudo, avaliamos as vias proteômicas e marcadores lipidômicos resultantes do tratamento com AGPIs n-3. Isto foi feito por meio da comparação, antes (grupo AT) e depois do tratamento (grupo DT), de pools de tecido hepático (análise por interactoma) e amostras de plasma (OPLS-DA). RESULTADOS: Foram identificadas proteínas hepáticas, exclusivamente e/ou alteradamente expressas, no grupo DT, relacionadas com vias de matriz celular, metabolismo lipídico, de estresse oxidativo, e de retículo endoplasmático e respiração celular. Com excessão da via de matriz celular, a análise do interactoma revelou alteração funcional significativa das vias moduladas por essas proteínas. Em conjunto, essas alterações foram sugestivas de diminuição de lipotoxicidade, estresse oxidativo e respiração anaeróbia, e aumento de respiração aeróbia após tratamento com AGPIs n-3. Estas modificações são marcadores potenciais de melhora de função de retículo endoplasmático e mitocondrial. Em adição, após o tratamento com AGPIs n-3, o perfil lipidômico plasmático mostrou-se alterado com significativo aumento de glicerofosfolípides, ALA e EPA, e diminuição de ácido araquidônico (n-6) e da razão AGPIs n-6/n-3. Estes dados são concordantes com potencial melhora das funções de retículo endoplasmático e mitocondriais. CONCLUSÃO: O tratamento com AGPIs n-3 em pacientes com NASH influenciou favoravelmente o perfil proteômico hepático e lipidômico sistêmico. Em conjunto, essas alterações sugerem melhora da função de retículo endoplasmático e mitocondrial, com potencial impacto na homeostase celular, por meio da modulação de diferentes vias biológicas / INTRODUCTION: Non-alcoholic steatohepatitis (NASH) is considered a public health problem, given its increasing incidence and its possible role in hepatocellular carcinogenesis. Current therapies involve diet and lifestyle changes, but its applicability suffers from low patients adherence. Pharmacological approaches are still missing. A main difficulty in the NASH management lies in the limited understanding of its pathophysiology, which seems to involve complex metabolic and inflammatory disturbances. Omega-3 polyunsaturated fatty acids (n-3 PUFAs) are recognized for its modulatory properties on lipid metabolism and inflammation and are decreased in patients with NASH. The clinical use of these PUFAs has shown benefit in controlling steatosis and the production of metabolic and inflammatory response markers in NASH, despite some conflicting reports. Understanding mechanisms modulated by n-3 PUFAs in NASH may be useful for identifying molecular targets that could assist in the design of effective pharmacologic interventions. In this sense, omics sciences are particularly useful for understanding molecular mechanisms with high translational value to clinical practice and may contribute to the identification of these targets. AIM: This study evaluated the liver proteomic and plasma lipidomics responses of patients with NASH towards treatment with n-3 PUFAs. METHODS: The proteomic and lipidomic evaluations were studied by mass spectrometry and / or gas chromatography in samples from liver biopsies and plasma collected from patients enrolled in a preliminary clinical trial of the Hospital das Clínicas da Faculdade de Medicina da Universidade de São Paulo. This study involved adult patients of both sexes diagnosed with NASH treated daily for 6 months, with 3 capsules containing a mixture of linseed and fish oils [0.315 g PUFAs: 0.065 g eicosapentaenoic acid (EPA) , 0.050 g docosahexaenoic (DHA) and 0.2 g alpha linolenic acid (ALA) per capsule]. Patients, after treatment with n-3 PUFAs, with higher concentrations of ALA and DHA and lower arachidonic acid (AA) showed improvement of liver histology alterations. In the present study we evaluated the proteomics pathways and lipidomics markers resulted from treatment with PUFAs n-3. This was performed by comparing, before (BT group) and after (AT group) treatment, liver tissue pools (analysis interactome) and plasma samples (OPLS-DA). RESULTS: It was identified, in a way exclusive and altered, the expressed liver proteins in AT group, related to pathways of cellular matrix, lipid metabolism, oxidative and endoplasmic reticulum stress and cellular respiration. With the exception of cell matrix, the analysis of the interactome revealed substantial functional alterations of the pathways modulated by these proteins. Together, these changes were suggestive of decreased lipotoxicity, oxidative stress and anaerobic respiration and increased aerobic respiration following treatment with PUFAs n-3. These modifications are potential markers of endoplasmic reticulum and mitochondrial functions improvement. In addition, after treatment with n-3 PUFAs, the lipidomics profile was modified, with significant increase in glycerophospholipids, ALA and EPA and decrease of arachidonic acid (AA) and n-6/n-3 AGPIs ratio. These findings are concordant with potential improvement of reticulum endoplasmic and mitochondrial functions. CONCLUSION: In patients with NASH the treatment with n-3 PUFAs favorably influenced hepatic proteomic and systemic lipidomics profiles. Together, these changes suggest improved endoplasmic reticulum and mitochondrial functions, with potential impact on cellular homeostasis through the modulation of different biological pathways

INTRODUÇÃO: A esteato-hepatite não alcóolica (NASH) é considerada problema de saúde pública, dada sua crescente incidência e seu possível papel na carcinogênese hepato-celular. Terapias atuais envolvem alterações de dieta e estilo de vida, mas têm seu resultado prejudicado pela baixa aderência dos pacientes. Abordagens farmacológicas ainda são precárias. Uma grande dificuldade no manejo de NASH reside no limitado entendimento de sua fisiopatologia, que parece envolver complexas alterações metabólicas e inflamatórias. Ácidos graxos poli-insaturados ômega-3 (AGPIs n-3) são reconhecidos por suas propriedades moduladoras do metabolismo lipídico e da inflamação, e estão diminuídos em pacientes com NASH. O uso clínico de AGPIs n-3 tem mostrado benefício no controle da esteatose e na produção de marcadores da resposta metabólica e inflamatória em NASH, embora com algumas observações contraditórias. A compreensão de mecanismos moleculares modulados por AGPIs n-3 em NASH podem ser úteis para identificar alvos moleculares que auxiliem no desenho de intervenção farmacológica efetiva. Nesse sentido, ciências ômicas são particularmente úteis para a compreensão de mecanismos moleculares com alto valor translacional para a prática clínica e podem contribuir para a identificação desses alvos. OBJETIVO: O presente estudo avaliou a resposta proteômica hepática e lipidômica plasmática de pacientes com NASH perante o tratamento com AGPIs n-3. MÉTODO: As avaliações proteômicas e lipidômicas foram desenvolvidas por espectometria de massas e/ou cromatografia gasosa em amostras de biópsias hepáticas e plasma coletadas de pacientes envolvidos em estudo preliminar, realizado no Hospital das Clínicas da Faculdade de Medicina da Universidade de São Paulo. O referido estudo envolveu pacientes adultos, de ambos os sexos e com diagnóstico de NASH tratados diariamente, durante 6 meses, com 3 cápsulas contendo mistura de óleo de linhaça e óleo de peixe [0,315 g AGPIs: sendo 0,065 g de ácido eicosapentaenoico (EPA), 0,050 g de docosahexaenoico (DHA) e 0,2 g alfa linolênico (ALA) por cápsula]. Pacientes, após o tratamento com AGPIs n-3, que apresentaram altas concentrações plasmáticas de ALA e/ou DHA e/ou baixas de ácido araquidônico (AA) mostraram melhora parcial das alterações de histologia hepática. No presente estudo, avaliamos as vias proteômicas e marcadores lipidômicos resultantes do tratamento com AGPIs n-3. Isto foi feito por meio da comparação, antes (grupo AT) e depois do tratamento (grupo DT), de pools de tecido hepático (análise por interactoma) e amostras de plasma (OPLS-DA). RESULTADOS: Foram identificadas proteínas hepáticas, exclusivamente e/ou alteradamente expressas, no grupo DT, relacionadas com vias de matriz celular, metabolismo lipídico, de estresse oxidativo, e de retículo endoplasmático e respiração celular. Com excessão da via de matriz celular, a análise do interactoma revelou alteração funcional significativa das vias moduladas por essas proteínas. Em conjunto, essas alterações foram sugestivas de diminuição de lipotoxicidade, estresse oxidativo e respiração anaeróbia, e aumento de respiração aeróbia após tratamento com AGPIs n-3. Estas modificações são marcadores potenciais de melhora de função de retículo endoplasmático e mitocondrial. Em adição, após o tratamento com AGPIs n-3, o perfil lipidômico plasmático mostrou-se alterado com significativo aumento de glicerofosfolípides, ALA e EPA, e diminuição de ácido araquidônico (n-6) e da razão AGPIs n-6/n-3. Estes dados são concordantes com potencial melhora das funções de retículo endoplasmático e mitocondriais. CONCLUSÃO: O tratamento com AGPIs n-3 em pacientes com NASH influenciou favoravelmente o perfil proteômico hepático e lipidômico sistêmico. Em conjunto, essas alterações sugerem melhora da função de retículo endoplasmático e mitocondrial, com potencial impacto na homeostase celular, por meio da modulação de diferentes vias biológicas / INTRODUCTION: Non-alcoholic steatohepatitis (NASH) is considered a public health problem, given its increasing incidence and its possible role in hepatocellular carcinogenesis. Current therapies involve diet and lifestyle changes, but its applicability suffers from low patients adherence. Pharmacological approaches are still missing. A main difficulty in the NASH management lies in the limited understanding of its pathophysiology, which seems to involve complex metabolic and inflammatory disturbances. Omega-3 polyunsaturated fatty acids (n-3 PUFAs) are recognized for its modulatory properties on lipid metabolism and inflammation and are decreased in patients with NASH. The clinical use of these PUFAs has shown benefit in controlling steatosis and the production of metabolic and inflammatory response markers in NASH, despite some conflicting reports. Understanding mechanisms modulated by n-3 PUFAs in NASH may be useful for identifying molecular targets that could assist in the design of effective pharmacologic interventions. In this sense, omics sciences are particularly useful for understanding molecular mechanisms with high translational value to clinical practice and may contribute to the identification of these targets. AIM: This study evaluated the liver proteomic and plasma lipidomics responses of patients with NASH towards treatment with n-3 PUFAs. METHODS: The proteomic and lipidomic evaluations were studied by mass spectrometry and / or gas chromatography in samples from liver biopsies and plasma collected from patients enrolled in a preliminary clinical trial of the Hospital das Clínicas da Faculdade de Medicina da Universidade de São Paulo. This study involved adult patients of both sexes diagnosed with NASH treated daily for 6 months, with 3 capsules containing a mixture of linseed and fish oils [0.315 g PUFAs: 0.065 g eicosapentaenoic acid (EPA) , 0.050 g docosahexaenoic (DHA) and 0.2 g alpha linolenic acid (ALA) per capsule]. Patients, after treatment with n-3 PUFAs, with higher concentrations of ALA and DHA and lower arachidonic acid (AA) showed improvement of liver histology alterations. In the present study we evaluated the proteomics pathways and lipidomics markers resulted from treatment with PUFAs n-3. This was performed by comparing, before (BT group) and after (AT group) treatment, liver tissue pools (analysis interactome) and plasma samples (OPLS-DA). RESULTS: It was identified, in a way exclusive and altered, the expressed liver proteins in AT group, related to pathways of cellular matrix, lipid metabolism, oxidative and endoplasmic reticulum stress and cellular respiration. With the exception of cell matrix, the analysis of the interactome revealed substantial functional alterations of the pathways modulated by these proteins. Together, these changes were suggestive of decreased lipotoxicity, oxidative stress and anaerobic respiration and increased aerobic respiration following treatment with PUFAs n-3. These modifications are potential markers of endoplasmic reticulum and mitochondrial functions improvement. In addition, after treatment with n-3 PUFAs, the lipidomics profile was modified, with significant increase in glycerophospholipids, ALA and EPA and decrease of arachidonic acid (AA) and n-6/n-3 AGPIs ratio. These findings are concordant with potential improvement of reticulum endoplasmic and mitochondrial functions. CONCLUSION: In patients with NASH the treatment with n-3 PUFAs favorably influenced hepatic proteomic and systemic lipidomics profiles. Together, these changes suggest improved endoplasmic reticulum and mitochondrial functions, with potential impact on cellular homeostasis through the modulation of different biological pathways

Indiana University-Purdue University Indianapolis (IUPUI) / Obesity increases cardiovascular disease risk and is associated with factors of the “metabolic syndrome” (MetS), a disorder including hypertension, hypercholesterolemia and/or impaired glucose tolerance. Expanding adipose and subsequent inflammation is implicated in vascular dysfunction in MetS. Perivascular adipose tissue (PVAT) surrounds virtually every artery and is capable of releasing factors that influence vascular reactivity, but the effects of PVAT in the coronary circulation are unknown. Accordingly, the goal of this investigation was to delineate mechanisms by which lean vs. MetS coronary PVAT influences vasomotor tone and the coronary PVAT proteome. We tested the hypothesis that MetS alters the functional expression and vascular contractile effects of coronary PVAT in an Ossabaw swine model of the MetS. Utilizing isometric tension measurements of coronary arteries in the absence and presence of PVAT, we revealed the vascular effects of PVAT vary according to anatomical location as coronary and mesenteric, but not subcutaneous adipose tissue augmented coronary artery contractions to KCl. Factors released from coronary PVAT increase baseline tension and potentiate constriction of isolated coronary arteries relative to the amount of adipose tissue present. The effects of coronary PVAT are elevated in the setting of MetS and occur independent of endothelial function. MetS is also associated with substantial alterations in the coronary PVAT proteome and underlying increases in vascular smooth muscle Ca2+ handling via CaV1.2 channels, H2O2-sensitive K+ channels and/or upstream mediators of these ion channels. Rho-kinase signaling participates in the increase in coronary artery contractions to PVAT in lean, but not MetS swine. These data provide novel evidence that the vascular effects of PVAT vary according to anatomic location and are influenced by the MetS phenotype.

Indiana University-Purdue University Indianapolis (IUPUI) / Signal Transducer and Activator of Transcription 3 (Stat3) is a transcription factor expressed in bone and joint cells that include osteoblasts, osteocytes, osteoclasts, and chondrocytes. Stat3 is activated by a variety of cytokines and growth factors, including IL-6/gp130 family cytokines. These cytokines not only regulate the differentiation of osteoblasts and osteoclasts, but also regulate proliferation of chondrocytes through Stat3 activation. In 2007, mutations of Stat3 have been confirmed to cause a rare human immunodeficiency disease – Job syndrome which presents skeletal abnormalities like: reduced bone density (osteopenia), scoliosis, hyperextensibility of joints, and recurrent pathological bone fractures. Changes in the Stat3 gene alter the structure and function of the Stat3 proteins, impairing its ability to control the activity of other genes. However, little is known about the effects of Stat3 mutations on bone cells and tissues. To investigate the in vivo physiological role of Stat3 in bone homeostasis, osteoblast/osteocyte-specific Stat3 knockout (KO) mice were generated via the Cre-LoxP recombination system. The osteoblast/osteocyte-specific Stat3 KO mice showed bone abnormalities and an osteoporotic phenotype because of a reduced bone formation rate. Furthermore, inactivation of Stat3 decreased load-driven bone formation, and the disruption of Stat3 in osteoblasts suppressed load-driven mitochondrial activity, which led to an elevated level of reactive oxygen species (ROS) in cultured primary osteoblasts. Stat3 has been found to be responsive to mechanical stimulation, and might play an important role in mechanical signal transduction in osteocytes. To investigate the role Stat3 plays in mechanical signaling transduction, osteocyte-specific Stat3 knockout (KO) mice were created. Inactivation of Stat3 in osteocytes presented a significantly reduced load-driven bone formation. Decreased osteoblast activity indicated by reduced osteoid surface was also found in osteocyte-specific Stat3 KO mice. Moreover, sclerostin (SOST) protein which is a critical osteocyte-specific inhibitor of bone formation, its encoded gene SOST expression has been found to be enhanced in osteocyte-specific Stat3 KO mice. Thus, these results clearly demonstrated that Stat3 plays an important role in bone homeostasis and mechanotransduction, and Stat3 is not only involved in bone-formation-important genes regulation in the nucleus but also in mediation of ROS and oxidative stress in mitochondria.

Indiana University-Purdue University Indianapolis (IUPUI) / Multiple sclerosis (MS) is a disabling disease that afflicts an estimated two million people worldwide. The disease is characterized by degradation of the myelin sheath that insulates neurons of the central nervous system manifesting as a heterogeneous collection of symptoms. Two enzymes, protein arginine deaminases type 2 and 4 (PAD2 and PAD4) have been implicated to play an etiologic role in demyelination and neurodegeneration by catalyzing a post-translational modification of arginine peptide residues to citrulline. The pathogenesis of MS is poorly understood, though vitamin D deficiency is a well-associated risk factor for developing the disorder. Using the experimental autoimmune encephalomyelitis (EAE) model of MS we demonstrate vitamin D treatment to attenuate over-expression of PAD 2 and 4 in the brain and spine during EAE. In addition, we identify two molecules produced by peripheral immune cells, IFNɣ and IL-6, as candidate signaling molecules that induce PAD expression in the brain. We demonstrate vitamin D treatment to inhibit IFNɣ mediated up regulation of PAD2 and PAD4 both directly within the brain and by modulating PAD-inducing cytokine production by infiltrating immune cells. These results provide neuroprotective rational for the supplementation of vitamin D in MS patients. More importantly, these results imply an epigenetic link between vitamin D deficiency and the pathogenesis of MS that merits further investigation.