Glycemic control and study of lipid and bone metabolism in type 1 diabetic children / Control glucémico y estudio del metabolismo lipídico y del óseo en niños con diabetes de tipo 1

Introduction. Type 1 diabetes mellitus is considered one of the most common chronic diseases of childhood. It is a high-risk factor for developing early cardiovascular disease and it also affects bone health. Objective. To describe demographic characteristics and biochemical parameters of a population of children with type 1 diabetes, evaluated in the pediatric diabetes unit of a tertiary Spanish hospital. Materials and methods. In this retrospective study, we determined metabolic, lipid, and bone parameters in 124 children with type 1 diabetes who were monitored in the pediatric diabetes unit of the Hospital Universitario Miguel Servet in Zaragoza (Spain) from May 2020 to July 2021. Results. Children with type 1 diabetes have worse metabolic control of the disease at puberty, but their lipid control is considered acceptable. We found an inverse correlation between bone formation markers and disease duration, as well as with metabolic control. Conclusion. Bone formation markers are inversely correlated with the percentage of glycated hemoglobin and diabetes evolution time. Patients' lipid and bone profiles are more favorable when metabolic control of the disease is achieved.

Introducción. La diabetes mellitus de tipo 1 se considera una de las enfermedades crónicas más frecuentes de la infancia. Es un factor de gran riesgo de desarrollar enfermedad cardiovascular temprana y afecta también la salud ósea. Objetivo. Describir las características demográficas y los parámetros bioquímicos de una población de niños con diabetes de tipo 1, supervisados en la unidad pediátrica de diabetes de un hospital español de tercer nivel. Materiales y métodos. En este estudio retrospectivo, se determinaron los parámetros de control metabólico, lipídico y óseo en 124 niños con diabetes de tipo 1, a los que se hizo seguimiento en la Unidad Pediátrica de Diabetes del Hospital Universitario Miguel Servet de Zaragoza, desde mayo del 2020 hasta julio del 2021. Resultados. Los niños con diabetes de tipo 1 presentan peor control metabólico de la enfermedad en la pubertad, pero su control lipídico se puede considerar aceptable. Existe una correlación inversa de los marcadores de formación ósea con el tiempo de evolución de la enfermedad, así como con el control metabólico. Conclusión. Los marcadores de formación ósea se encuentran correlacionados de forma inversa con el porcentaje de hemoglobina glicosilada y con el tiempo de evolución de la diabetes. En estos pacientes, el perfil lipídico y el óseo son más favorables cuando existe un buen control metabólico de la enfermedad.

Control glucémico y estudio del metabolismo lipídico y del óseo en niños con diabetes de tipo 1 / Glycemic control and study of lipid and bone metabolism in type 1 diabetic children

Resumen Introducción. La diabetes mellitus de tipo 1 se considera una de las enfermedades crónicas más frecuentes de la infancia. Es un factor de gran riesgo de desarrollar enfermedad cardiovascular temprana y afecta también la salud ósea. Objetivo. Describir las características demográficas y los parámetros bioquímicos de una población de niños con diabetes de tipo 1, supervisados en la unidad pediátrica de diabetes de un hospital español de tercer nivel. Materiales y métodos. En este estudio retrospectivo, se determinaron los parámetros de control metabólico, lipídico y óseo en 124 niños con diabetes de tipo 1, a los que se hizo seguimiento en la Unidad Pediátrica de Diabetes del Hospital Universitario Miguel Servet de Zaragoza, desde mayo del 2020 hasta julio del 2021. Resultados. Los niños con diabetes de tipo 1 presentan peor control metabólico de la enfermedad en la pubertad, pero su control lipídico se puede considerar aceptable. Existe una correlación inversa de los marcadores de formación ósea con el tiempo de evolución de la enfermedad, así como con el control metabólico. Conclusión. Los marcadores de formación ósea se encuentran correlacionados de forma inversa con el porcentaje de hemoglobina glicosilada y con el tiempo de evolución de la diabetes. En estos pacientes, el perfil lipídico y el óseo son más favorables cuando existe un buen control metabólico de la enfermedad.

Abstract Introduction. Type 1 diabetes mellitus is considered one of the most common chronic diseases of childhood. It is a high-risk factor for developing early cardiovascular disease and it also affects bone health. Objective. To describe demographic characteristics and biochemical parameters of a population of children with type 1 diabetes, evaluated in the pediatric diabetes unit of a tertiary Spanish hospital. Materials and methods. In this retrospective study, we determined metabolic, lipid, and bone parameters in 124 children with type 1 diabetes who were monitored in the pediatric diabetes unit of the Hospital Universitario Miguel Servet in Zaragoza (Spain) from May 2020 to July 2021. Results. Children with type 1 diabetes have worse metabolic control of the disease at puberty, but their lipid control is considered acceptable. We found an inverse correlation between bone formation markers and disease duration, as well as with metabolic control. Conclusion. Bone formation markers are inversely correlated with the percentage of glycated hemoglobin and diabetes evolution time. Patients' lipid and bone profiles are more favorable when metabolic control of the disease is achieved.

Final irrigation with bioglass solution in regenerative endodontic procedure induces tissue formation inside the root canals, collagen maturation, proliferation cell and presence of osteocalcin.

AIM: To evaluate the influence of an experimental solution of cobalt-doped F18 bioactive glass (F18Co) on tissue repair following regenerative endodontic procedure (REP) in rat molars. METHODOLOGY: The F18Co solution was prepared at a ratio of 1:5 F18Co powder to distilled water. The right or left upper first molars of 12 Wistar rats were used, where the pulps were exposed, removed, and irrigated with 2.5% sodium hypochlorite (NaOCl), followed by 17% ethylenediaminetetraacetic acid (EDTA) (5 min each). Subsequently, the molars were divided into two groups (n = 6): REP-SS and REP-F18Co, where they received a final irrigation (5 min) with saline solution (SS) or F18Co solution, respectively. Then, intracanal bleeding was induced, and the tooth was sealed. Untreated molars were used as controls (n = 3). At 21 days, the rats were euthanized, and the specimens were processed for analysis of mineralized tissue and soft tissue formation inside the root canal using haematoxylin-eosin. The presence and maturation of collagen were evaluated by Masson's trichrome and picrosirius red staining. Immunolabelling analyses of proliferating cell nuclear antigen (PCNA) and osteocalcin (OCN) were performed. The data were submitted to the Mann-Whitney U-test (p < .05). RESULTS: There was a similar formation of mineralized tissue in thickness and length in REP-SS and REP-F18Co groups (p > .05). Regarding the presence of newly formed soft tissue, most specimens of the REP-F18Co had tissue formation up to the cervical third of the canal, whilst the REP-SS specimens showed formation up to the middle third (p < .05), and there was higher maturation of collagen in REP-F18Co (p < .05). The number of PCNA-positive cells found in the apical third of the root canal was significantly higher in the F18Co group, as well as the OCN immunolabelling, which was severe in most specimens of REP-F18Co, and low in most specimens of REP-SS. CONCLUSION: The final irrigation with F18Co bioactive glass solution in REP did not influence mineralized tissue formation but induced soft tissue formation inside the root canals, with higher collagen maturation, and an increase in PCNA-positive cells and OCN immunolabelling.

Additional Insights Into the Role of Osteocalcin in Osteoblast Differentiation and in the Early Steps of Developing Alveolar Process of Rat Molars.

This study investigated whether osteocalcin (OCN) is present in osteoblast precursors and its relationship with initial phases of alveolar process formation. Samples of maxillae of 16-, 18-, and 20-day-old rat embryos (E16, E18, and E20, respectively), and 05-, 10-, and 15-day-old postnatal rats (P05, P10, and P15, respectively) were fixed and embedded in paraffin or araldite. Immunohistochemistry for osterix (Osx), alkaline phosphatase (ALP), and OCN detection was performed and the number of immunolabelled cells was computed. Non-decalcified sections were subjected to the von Kossa method combined with immunohistochemistry for Osx or OCN detection. For OCN immunolocalization, samples were fixed in 0.5% glutaraldehyde/2% formaldehyde and embedded in LR White resin. The highest number of ALP- and OCN-immunolabelled cells was observed in dental follicle of E16 specimens, mainly in basal portions of dental alveolus. In corresponding regions, osteoblasts in differentiation adjacent to von Kossa-positive bone matrix exhibited Osx and OCN immunoreactivity. Ultrastructural analysis revealed OCN immunoreactive particles inside osteoblast in differentiation, and in bone matrix associated with collagen fibrils and within matrix vesicles, at early stages of alveolar process formation. Our results indicate that OCN plays a role in osteoblast differentiation and may regulate calcium/phosphate precipitation during early mineralization of the alveolar process.

Refining Evaluation of Bone Mass and Adipose Distribution in Dunnigan Syndrome.

Familial partial lipodystrophies (FPLD) are rare diseases characterized by selective loss of subcutaneous adipose tissue at different sites. This cross-sectional observational study aimed to estimate adipose tissue in the bone marrow (BMAT), intra (IMCL) and extra-myocyte lipids (EMCL), and define the bone phenotype in the context of FPLD2/Dunnigan syndrome (DS). The subjects comprised 23 controls (C) and 18 DS patients, matched by age, weight and height. Blood samples, dual-energy X-ray absorptiometry for bone mineral density (BMD) and trabecular bone score (TBS) and 1H-spectroscopy using magnetic resonance to estimate BMAT in the lumbar spine, IMCL, EMCL and osteoclastogenesis were assessed. The prevalence of diabetes mellitus was 78% in DS patients. Glucose, HbA1c, triglycerides, insulin and HOMA-IR levels were elevated in DS, whereas HDLc, 25(OH)D, PTH and osteocalcin levels were reduced. BMD was similar between groups at all sites, except 1/3 radius, which was lower in DS group. TBS was reduced in DS. DS presented increased osteoclastogenesis and elevated BMAT, with greater saturation levels and higher IMCL than the C group. HOMA-IR and EMCL were negatively associated with TBS; osteocalcin and EMCL were correlated negatively with BMD. This study contributes to refining the estimation of adipose tissue in DS by showing increased adiposity in the lumbar spine and muscle tissue. DXA detected lower TBS and BMD in the 1/3 radius, suggesting impairment in bone quality and that bone mass is mainly affected in the cortical bone.

Determinación de rangos de BCTx y N-MID Osteocalcina y asociación con Osteoporosis en mujeres chilenas posmenopáusicas

Introducción: La densitometría ósea (DO) tiene alta especificidad para el diagnóstico de osteoporosis, pero sensibilidad bajo lo óptimo para estimar el riesgo de fracturas. Éste, se puede calcular por algoritmos de predictores, y se ha propuesto incluir los marcadores óseos (MO), pero la magnitud de su asociación es incierta. El MO recomendado para medir resorción ósea es Beta-Cross Laps (B-CTx), y uno de formación ósea es la osteocalcina. Objetivos: 1) Establecer rangos de B-CTx y N-MID osteocalcina (N-MID) en mujeres posmenopáusicas (MPM), y comparar los niveles entre grupos: 1) Control sano y 2) Con DO alterada. Material y Métodos: Se reclutaron MPM, con DO del último año. Se realizó encuesta de factores de riesgo de fracturas y medición de MO. Se excluyeron MPM con causa secundaria para compromiso del recambio óseo. Resultados: 117 MPM (57 control, 60 DO alterada), 18 % osteoporosis, comparables. Los rangos de B-CTx y N-MID fueron 0,41 ± 0,18 [IC95% 0,37- 0,45] y 22,76 ± 7,73 [IC95% 21,29-24,24] ng/mL. Los niveles promedios de B-CTx y N-MID fueron más altos en grupo con DO alterada (0,46 ± 0,19 y 24,29 ± 8,04 ng/mL). Se encontró correlación moderada entre ambos MO, pero débil con DO alterada. Conclusiones: Se determinaron por primera vez rangos de B-CTx y N-MID en MPM chilenas, corroborando la similitud a otros países. Se encontraron valores discretamente más elevados de MO en grupo DO alterada, probablemente atribuible a causas secundarias no reportadas. Estos MO podrían constituir una herramienta complementaria a la DO y FRAX en la evaluación ósea.

Introduction: Bone densitometry (BD) has high specificity in the osteoporosis diagnosis but suboptimal sensitivity to estimate fracture risk. It was proposed that bone turnover markers (BTM) could be included in the osteoporosis risk algorithm, although the extent of its association is unknown. One recommended BTM to assess bone resorption is Beta-Cross Laps (B-CTx), while a BTM to assess bone formation is osteocalcin. Aims: To establish BCTx and N-MID osteocalcin (N-MID) ranges in postmenopausal women (PM) and compare BTM levels in two groups: control and with abnormal BD. Methods: PM with BD within the last year were recruited. A questionnaire of risk factors for fractures was applied, and BTM was measured. Volunteers with diseases that would affect bone remodeling were excluded. Results: 117 PM (57 control and 60 with abnormal BD) were recruited. 18% had osteoporosis, and the groups were comparable. The ranges for B-CTx and N-MID were 0.41 ± 0.18 [IC95% 0.37-0.45] and 22.76 ± 7.73 [IC95% 21.29-24.24] ng/mL. The mean levels of B-CTx and N-MID were higher in the group with abnormal BD (0.46 ± 0.19 and 24.29 ± 8.04 ng/mL). A moderate correlation between both BTM was found, but it was weak with abnormal BD. Conclusions: B-CTx and N-MID ranges were assessed for the first time in Chilean PM, similar to values found in other countries. Slightly higher values of BTM were found in the group with abnormal BD, which the presence of omitted secondary causes could explain. These BTM could be a complementary tool to BD and FRAX in bone evaluation.

Hyperglycemic microenvironment compromises the homeostasis of communication between the bone-brain axis by the epigenetic repression of the osteocalcin receptor, Gpr158 in the hippocampus.

Diabetes mellitus (DM) is a chronic metabolic disease, mainly characterized by increased blood glucose and insulin dysfunction. In response to the persistent systemic hyperglycemic state, numerous metabolic and physiological complications have already been well characterized. However, its relationship to bone fragility, cognitive deficits and increased risk of dementia still needs to be better understood. The impact of chronic hyperglycemia on bone physiology and architecture was assessed in a model of chronic hyperglycemia induced by a single intraperitoneal administration of streptozotocin (STZ; 55 mg/kg) in Wistar rats. In addition, the bone-to-brain communication was investigated by analyzing the gene expression and methylation status of genes that encode the main osteokines released by the bone [Fgf23 (fibroblast growth factor 23), Bglap (bone gamma-carboxyglutamate protein) and Lcn2 (lipocalin 2) and their receptors in both, the bone and the brain [Fgfr1 (fibroblast growth factor receptor 1), Gpr6A (G-protein coupled receptor family C group 6 member A), Gpr158 (G protein-coupled receptor 158) and Slc22a17 (Solute carrier family 22 member 17)]. It was observed that chronic hyperglycemia negatively impacted on bone biology and compromised the balance of the bone-brain endocrine axis. Ultrastructural disorganization was accompanied by global DNA hypomethylation and changes in gene expression of DNA-modifying enzymes that were accompanied by changes in the methylation status of the osteokine promoter region Bglap and Lcn2 (lipocalin 2) in the femur. Additionally, the chronic hyperglycemic state was accompanied by modulation of gene expression of the osteokines Fgf23 (fibroblast growth factor 23), Bglap (bone gamma-carboxyglutamate protein) and Lcn2 (lipocalin 2) in the different brain regions. However, transcriptional regulation mediated by DNA methylation was observed only for the osteokine receptors, Fgfr1(fibroblast growth factor receptor 1) in the striatum and Gpr158 (G protein-coupled receptor 158) in the hippocampus. This is a pioneer study demonstrating that the chronic hyperglycemic state compromises the crosstalk between bone tissue and the brain, mainly affecting the hippocampus, through transcriptional silencing of the Bglap receptor by hypermethylation of Gpr158 gene.

Biocompatibility and bioactive potential of an experimental tricalcium silicate-based cement in comparison with Bio-C repair and MTA Repair HP materials.

AIM: To evaluate the tissue reaction of a tricalcium silicate-based repair material associated with 30% calcium tungstate (TCS + CaWO4 ) in comparison to Bio-C Repair (Bio-C; Angelus) and to MTA Repair HP (MTA HP; Angelus). METHODOLOGY: Polyethylene tubes filled with one of the materials or left empty (control group, CG) were implanted into the subcutaneous tissues of rats for 7, 15, 30 and 60 days (n = 32/group). The capsule thickness, number of inflammatory cells, collagen content, interleukin-6 (IL-6), osteocalcin (OCN), von Kossa reaction and analysis under polarized light were evaluated. The data were subjected to generalized linear models for repeated measures, except the OCN. OCN data were submitted to Kruskal-Wallis and Dunn's post hoc test and Friedman followed by Nemenyi's test at significance level of 5%. RESULTS: At all time points, significant differences in the number of inflammatory cells were not observed between TCS + CaWO4 and Bio-C, whereas, at 15, 30 and 60 days, no significant difference was detected between TCS + CaWO4 and MTA HP. At all periods, significant differences were not detected in the number of fibroblasts in TCS + CaWO4 versus MTA HP, and, at 60 days, no significant difference was demonstrated between these groups and CG. Significant differences in the immunoexpression of IL-6 were not detected amongst bioceramic materials at all periods. From 7 to 60 days, significant reduction in the number of inflammatory cells, number of IL-6-immunopositive cells and in the capsule thickness was accompanied by significant increase in the collagen in all groups. OCN-immunolabelled cells, von Kossa-positive structures and amorphous calcite deposits were observed around all materials, whereas, in the CG, these structures were not seen. CONCLUSIONS: These findings indicate that the experimental material (TCS + CaWO4 ) is biocompatible and has a bioactive potential, similar to the MTA HP and Bio-C Repair, and suggest its use as a root repair material.

Influência da fibrina rica em plaquetas em procedimento endodôntico regenerativo em molares de ratos / Influence of injectable platelet rich fibrin on regenerative endodontic procedure in rat's immature molars

Este estudo avaliou a influência da fibrina rica em plaquetas injetável (i-PRF) no reparo após procedimento endodôntico regenerativo (REP) em molares imaturos de ratos. Molares superiores direito ou esquerdo de 18 ratos foram divididos em grupos (n = 6): SI ­ foi removido o tecido pulpar do canal mesial, feita irrigação com hipoclorito de sódio e ácido etilenodiaminotetracético, e induzido o sangramento intracanal (SI); iPRF ­ após protocolo de irrigação, foi inserido no interior do canal radicular a i-PRF, sem indução do SI; i-PRF-SI ­ foi realizado o tratamento como no grupo i-PRF, e então, induzido o SI. Ao término, os dentes foram selados. Para produzir a i-PRF, outros 3 animais foram utilizados para a coleta sanguínea intracardíaca, e o sangue foi centrifugado (1200 rpm, 8 min). Aos 21 dias, os animais foram eutanasiados e as peças preparadas para análise histológica e imunohistoquímica para antígeno nuclear de proliferação celular (PCNA) e osteocalcina (OCN). Teste estatístico foi aplicado (p < 0,05). Houve formação de tecido mineralizado em comprimento ou espessura da raiz em todos os grupos (p > 0,05). A formação de tecido conjuntivo nos canais ocorreu até terço médio na maior parte dos espécimes de SI, e até terço cervical em i-PRF e i-PRF-SI (p < 0,05). Houve células semelhantes a odontoblastos no terço apical de metade dos espécimes de i-PRF, e terços apical e médio na maior parte dos espécimes de i-PRF-SI; estas células não foram encontradas no grupo SI (p < 0,05). Os espécimes dos grupos i-PRF e i-PRF-SI tiveram significante número de células positivas para PCNA (p < 0,05) e maior imunomarcação de OCN, principalmente o grupo i-PRF-SI quando comparado ao grupo SI (p < 0,05). Conclui-se que i-PRF auxiliou o processo de reparo após REP em ratos, induzindo formação de tecido conjuntivo nos canais radiculares, presença de células semelhantes a odontoblastos e células positivas para PCNA, e imunomarcação de OCN, principalmente quando associada ao SI.

This study evaluated the influence of injectable platelet-rich fibrin (i-PRF) on repair after regenerative endodontic procedure (REP) in immature molars of rats. Right or left upper molars of 18 rats were divided into groups (n = 6): IB ­ the pulp tissue of the mesial canal was removed, irrigation with sodium hypochlorite and ethylenediaminetetraacetic acid was performed, and intracanal bleeding (IB) was induced; i-PRF ­ after the irrigation protocol, i-PRF was inserted inside the root canal, without inducing of IB; i-PRF-IB ­ the treatment was carried out as in the i-PRF group and then the IB was induced. At the end, the teeth were sealed. To produce the i-PRF, another 3 animals were used for intracardiac blood collection, and the blood was centrifuged (1200 rpm, 8 min). After 21 days, the animals were euthanized and the samples were prepared for histological and immunohistochemical analysis for proliferating cell nuclear antigen (PCNA) and osteocalcin (OCN). Statistical test was applied (P < 0.05). There was formation of mineralized tissue in root length or thickness in all groups (P > 0.05). The formation of connective tissue in the canals occurred up to the medium third in most IB specimens, and up to the cervical third in i-PRF and iPRF-IB (P < 0.05). There were odontoblast-like cells in the apical third of half of the PRF specimens, and apical and medium thirds in most of the i-PRF specimens; these cells were not found in the IB group (P < 0.05). The i-PRF and i-PRF-IB group had significant higher number of PCNA-positive cells (P < 0.05), and higher OCN immunolabeling, mainly i-PRF-IB compared to IB group (P < 0.05). It is concluded that i-PRF helped the repair process after REP in rats, inducing connective tissue formation in root canals, presence of odontoblast-like and PCNA-positive cells, and OCN immunolabeling, mainly when associated with IB.

Análise da influência de solução de biovidro no procedimento endodôntico regenerativo / Analysis of the influence of bioglass solution on the regenerative endodontic procedure

Este estudo avaliou os efeitos de uma solução experimental de vidro bioativo F18 dopado com cobalto (F18Co) no reparo tecidual após procedimento endodôntico regenerativo (REP, do inglês regenerative endodontic procedure) em molares de ratos. A solução de F18Co foi preparada misturando o pó de F18Co com água destilada na proporção de 1:5. Os primeiros molares superiores direito ou esquerdo de 12 ratos Wistar foram utilizados, nos quais as polpas foram expostas, removidas e os canais irrigados com hipoclorito de sódio (NaOCl) 2,5%, seguido de ácido etilenodiaminotetraacético (EDTA) 17% (5 min cada). Em seguida, os molares foram divididos em dois grupos (n = 6): REP-SS e REP-F18Co, nos quais receberam irrigação final (5 min) com solução salina (SS) ou solução de F18Co, respectivamente. Em seguida, foi induzido sangramento intracanal e o dente foi selado. Molares não tratados foram usados como controles (n = 3). Após 21 dias, os ratos foram eutanasiados e as peças processadas para análise da formação de tecido mineralizado e tecido mole dentro do canal radicular, pela técnica de hematoxilinaeosina. A presença e maturação do colágeno foram avaliadas por coloração de tricrômio de Masson e picrosírius red. Análises da imunomarcação do antígeno nuclear de proliferação celular (PCNA) e osteocalcina (OCN) foram realizadas. Os dados foram submetidos ao teste de Mann-Whitney U (p < 0,05). Houve formação semelhante de tecido mineralizado em espessura e comprimento nos grupos REP-SS e REP-F18Co (p > 0,05). Em relação à presença de tecido neoformado no interior dos canais radiculares, a maioria dos espécimes de REP-F18Co apresentou formação de tecido até o terço cervical do canal radicular, enquanto do grupo REP-SS, até o terço médio (p < 0,05), e houve maior maturação colágena em REP-F18Co (p < 0,05). O número de células positivas para PCNA encontradas no terço apical do canal radicular foi significativamente maior em REP-F18Co, assim como a imunomarcação de OCN, que foi severa na maior parte dos espécimes do grupo REP-F18Co, e leve em REPSS. Conclui-se que a irrigação final com solução de biovidro F18Co em REP não influenciou a formação de tecido mineralizado, mas induziu a formação de tecido conjuntivo no interior dos canais radiculares, com maior maturação colágena, aumento no número de células positivas para PCNA e na imunomarcação de OCN.

This study evaluated the influence of an experimental solution of cobalt-doped F18 bioactive glass (F18Co) on tissue repair following regenerative endodontic procedure (REP) in rat molars. The F18Co solution was prepared at a ratio of 1:5 F18Co powder to distilled water. The right or left upper first molars of 12 Wistar rats were used, where the pulps were exposed, removed, and irrigated with 2.5% sodium hypochlorite (NaOCl), followed by 17% ethylenediaminetetraacetic acid (EDTA) (5 min each). Subsequently, the molars were divided into two groups (n = 6): REP-SS and REP F18Co, where they received a final irrigation (5 min) with saline solution (SS) or F18Co solution, respectively. Then, intracanal bleeding was induced, and the tooth was sealed. Untreated molars were used as controls (n = 3). At 21 days, the rats were euthanized, and the specimens were processed for analysis of mineralized tissue and soft tissue formation inside the root canal using haematoxylin-eosin. The presence and maturation of collagen was evaluated by Masson's trichrome and picrosirius red staining. Immunolabeling analyses of proliferating cell nuclear antigen (PCNA) and osteocalcin (OCN) were performed. The data were submitted to the Mann-Whitney U test (P < 0.05). There was similar formation of mineralized tissue in thickness and length in REP-SS and REP-F18Co groups (P > 0.05). Regarding the presence of newly formed soft tissue, most specimens of the REP-F18Co had tissue formation up to the cervical third of the canal, while the REP-SS specimens showed formation up to the middle third (P < 0.05), and there was higher maturation collagen in REP-F18Co (P < 0.05). The number of PCNA-positive cells found in the apical third of the root canal was significantly higher in the F18Co group, as well as the OCN immunolabeling, which was severe in most specimens of REP-F18Co, and low in most specimens of REP SS. In conclusion, the final irrigation with F18Co bioactive glass solution in REP did not influence mineralized tissue formation but induced soft tissue formation inside the root canals, with higher collagen maturation, and an increase in PCNA-positive cells and OCN immunolabeling

Effect of walking with blood flow restriction in elderly women with osteoporosis/osteopenia / Efeito da caminhada com restrição do fluxo sanguíneo em idosas com osteoporose/osteopenia

Abstract Introduction The preservation of bone mass in elderly women is associated with better levels of practice of systematic physical exercises. Aerobic training combined with blood flow restriction seems to be a new alternative that determines this process, but knowledge gaps are still observed when referring to exercise associated with blood flow restriction (BFR) and adaptations on bone variables. Objective To analyze the chronic effects of aerobic training with and without BFR on bone mineral density and bone biomarker osteocalcin concentrations in older women. Methods Thirty women were randomized into the following groups: walking on a treadmill at low intensity with BFR; moderate treadmill walking with no BFR; only BFR (no exercise) for 20 minutes, twice a week, for 24 weeks. Bone mineral density was measured before and 24 weeks after intervention. Blood serum osteocalcin concentrations were measured before, 12 and 24 weeks after intervention. Results There were no differences between groups in bone mineral density (femoral neck, p = 0.31; total femur, p = 0.17; lumbar spin, p = 0.06) and osteocalcine (W(2) = 0.27; p = 0.87) ouctomes after 24 weeks of intervention. Conclusion There was no difference between walking training, blood flow restriction only, or walking+blood flow restriction on bone mineral density and osteocalcin concentrations after 24-weeks of intervention in older women with osteopenia/osteoporosis.

Resumo Introdução A preservação da massa óssea em mulheres idosas está associada a melhores níveis de prática de exercícios físicos sistemáticos. O treinamento aeróbico combinado com restrição de fluxo sanguíneo (RFS) parece ser uma nova alternativa que determina esse processo, mas ainda são observadas lacunas de conhecimento quando se refere ao exercício associado à RFS e adaptações nas variáveis ósseas. Objetivo Analisar os efeitos crônicos do treinamento aeróbico com e sem RFS na densidade mineral óssea e nas concentrações do biomarcador ósseo osteocalcina em mulheres idosas. Métodos Trinta mulheres foram randomizadas nos seguintes grupos: caminhada em esteira de baixa intensidade com RFS; caminhada moderada em esteira sem RFS; apenas RFS (sem exercícios) por 20 minutos, duas vezes por semana, durante 24 semanas. A densidade mineral óssea foi medida antes e 24 semanas após a intervenção. As concentrações séricas de osteocalcina no sangue foram medidas antes, 12 e 24 semanas após a intervenção. Resultados Não houve diferenças entre os grupos na densidade mineral óssea (colo do fêmur, p = 0,31; fêmur total, p = 0,17; giro lombar, p = 0,06) e osteocalcina (W(2) = 0,27; p = 0,87) após 24 semanas de intervenção. Conclusão Não houve diferença entre treinamento de caminhada, apenas restrição de fluxo sanguíneo ou caminhada + restrição de fluxo sanguíneo na densidade mineral óssea e nas concentrações de osteocalcina após 24 semanas de intervenção em mulheres idosas com osteopenia/osteoporose.

Osteocalcin and Runx2 Expression in Anterior Maxillary Reconstructions Using Bone Xenografts Associated to Bone Marrow Aspirate Concentrate.

Background: It is known that a large number of mediators involved in osteogenesis can influence bone development and repair; however, whether these mediators could be used as markers of bone maturity has yet to be determined. Aim: To evaluate the expression of osteocalcin (OC) and Runt-related transcription factor 2 (Runx2) in bone biopsies obtained during the reconstruction of atrophic anterior maxillae using particulate bone xenografts with or without association of autogenous bone marrow aspirate concentrate (BMAC). Materials and Methods: Ten patients were distributed into two groups (n = 5), according to the type of grafting material used: Control group (CG), particulate bone xenograft alone, and test group (TG), particulate bone xenograft combined with BMAC. A bone specimen was removed from the graft area 4 months after grafting, before implant placement. The specimens were processed and submitted to immunohistochemical analysis for detection of OC and Runx2. Histomorphometry was used to ascertain the percentage of stained areas in both groups. The Wilcoxon Mann-Whitney U-Test was used in the statistical analysis (P < 0.05). Results: The immunohistochemical analysis revealed a significantly higher OC expression in the TG than in the CG, namely 27.40 ± 1.34% and 11.40 ± 2.70%, respectively (P < 0.05), and a significantly higher Runx2 expression in the TG than in the CG, namely 2.80 ± 0.84% and 0.40 ± 0.55%, respectively (P < 0.05). Conclusion: The OC and Runx2 expression levels were higher when BMAC was associated with the bone xenograft than when it was not.

Lithium chloride stimulates bone formation in extraction socket repair in rats.

PURPOSE: Previous evidence shows that lithium chloride (LiCl), a suppressor of glycogen synthase kinase-3ß (GSK-3ß), may enhance bone formation in several medical and dental conditions. Thus, the purpose of the current study was to assess the effects of LiCl on extraction socket repair in rats. METHODS: Thirty rats were randomly assigned into a control group (administration of water; n = 15) or a LiCl group (administration of 150 mg/kg of LiCl; n = 15). LiCl and water were given every other day, starting at 7 days before the extraction of upper first molars until the end of each experiment period. Histological sections from five rats per group were obtained at 10, 20, and 30 days post-extractions. Histometrical analysis of newly formed bone (NB) and the levels of tartrate-resistant acid phosphatase (TRAP)-stained cells were evaluated at 10, 20, and 30 days post-extractions. Immunohistochemical staining for receptor activator of nuclear factor kappa-Β ligand (RANKL), osteoprotegerin (OPG), bone sialoprotein (BSP), osteocalcin (OCN), and osteopontin (OPN) was assessed at 10 days post-extractions. RESULTS: The LiCl group had a greater proportion of NB than the control group at 20 days (P < 0.05). At 30 days, the rate of TRAP-stained cells was lower in the LiCl group than in the control group (P < 0.05). At 10 days, the LiCl group presented stronger staining for OPG, BSP, OPN, and OCN, when compared to the control group (P < 0.05). CONCLUSION: Systemic LiCl enhanced extraction socket repair, stimulated an overall increase in bone formation markers, and restricted the levels of TRAP in rats.

Bioactive potential of Bio-C Pulpo is evidenced by presence of birefringent calcite and osteocalcin immunoexpression in the rat subcutaneous tissue.

As the biocompatibility and bioactive potential of repair materials are desired characteristics in dentistry, the tissue response of Bio-C Pulpo, a bioceramic material launched on the marked by Angelus (Brazil), was compared with Biodentine (Septodont, France) and White MTA (WMTA; Angelus, Brazil). In 32 rats, 148 polyethylene tubes filled with Bio-C Pulpo, Biodentine or WMTA, and empty (CG, control group) were implanted into subcutaneous tissues for 7, 15, 30, and 60 days. The capsule thickness, numerical density of inflammatory cells (IC) and fibroblasts (Fb), amount of collagen, immunohistochemistry detection of interleukin-6 (IL-6) and osteocalcin (OCN), von Kossa and analysis under polarized light were performed. Data were subjected to two-way ANOVA followed by Tukey's test (p ≤ 0.05). At 7 and 15 days, the capsules around Bio-C Pulpo were thicker than in WMTA while, at 30 and 60 days, significant differences were not observed among the groups. Although at 7, 15, and 30 days, a greater number of IL-6-immunostained cells was found in Bio-C Pulpo and Biodentine than in WMTA, no significant difference was detected among the groups at 60 days. In all groups, the number of Fb and collagen content increased significantly over time. The capsules around materials exhibited von Kossa-positive and birefringent structures, and OCN-immunostained cells whereas, in the CG, these structures were not observed. Bio-C Pulpo, similarly to Biodentine and WMTA, is biocompatible, allows the connective tissue repair and presents bioactive potential in connective tissue of rats.

Osteocalcin serum concentrations and markers of energetic metabolism in pediatric patients. Systematic review and metanalysis.

Background: Osteocalcin plays a role in glucose metabolism in mice, but its relevance in human energetic metabolism is controversial. Its relationship with markers of energetic metabolism in the pediatric population has not been systematically addressed in infants and adolescents. Objective: This study aims to assess the mean differences between tOC, ucOC, and cOC among healthy children and children with type 1 or type 2 diabetes (T1D or T2D) and the correlation of these bone molecules with metabolic markers. Methods: A systematic review and metanalysis were performed following PRISMA criteria to identify relevant observational studies published in English and Spanish using PubMed, Scopus, EBSCO, and Web of Science databases. The risk of bias was assessed using New Castle-Ottawa scale. Effect size measures comprised standardized mean difference (SMD) and Pearson correlations. Heterogeneity and meta-regressions were performed. Results: The 20 studies included were of high quality and comprised 3,000 pediatric patients who underwent tOC, cOC, or ucOC measurements. Among healthy subjects, there was a positive correlation of ucOC with WC and weight, a positive correlation of tOC with FPG, HDL-c, WC, height, and weight, and a negative correlation between tOC and HbA1c. Among diabetic subjects, a negative correlation of ucOC with HbA1c and glycemia in both T1D and T2D was found and a negative correlation between tOC and HbA1c in T1D but not in T2D. The ucOC concentrations were lower in T2D, T1D, and patients with abnormal glucose status than among controls. The serum concentrations of tOC concentrations were lower among T1D than in controls. The patient's age, altitude, and HbA1c influenced the levels of serum tOC. Conclusion: Osteocalcin is involved in energy metabolism in pediatric subjects because it is consistently related to metabolic and anthropometric parameters. Systematic Review Registration: https://www.crd.york.ac.uk/prospero/, identifier: CRD42019138283.

Effects of teriparatide on bone formation in rats with experimentally induced premaxillary expansion

ABSTRACT Objective: This study aimed to evaluate the effects of systemic teriparatide on sutural bone formation after premaxillary suture expansion in rats. Material and Methods: Twenty Wistar male rats (8-10 weeks old) were randomly divided into two groups, namely, control (C, n=10) and teriparatide (T, n=10). An expansion force was applied to the maxillary incisors using helical spring for a seven-day expansion period, for both groups. On the eighth day, the rats were kept for a seven-day consolidation period, and then 60 µg/kg teriparatide (once a day) was administered to group T subcutaneously for seven days. Then, all the rats were sacrificed, and histological sections were stained with hemotoxylin-eosin for examination. Anti-osteonectin, anti-osteocalcin, anti-Vascular endothelial growth factor (VEGF) and anti-transforming growth factor beta (TGF-β) were evaluated by immunohistochemical analysis in the midpalatal suture area. Results: Histologically, the newly formed bone tissue was observed to be larger in group T than in group C. The number of immunoreactive osteoblasts for osteonectin, osteocalcin and VEGF antibodies was significantly higher in group T than in group C (p = 0.0001). The TGF-β antibody showed a mild reaction in group T, but did not reach significance in comparison with group C (p ˃ 0.05). Conclusion: Systemic teriparatide application following the premaxillary expansion of the suture area may stimulate bone formation and add to the consolidation of the expansion in rats by regulating osteonectin, osteocalcin and VEGF.

RESUMO Objetivo: O presente estudo teve como objetivo avaliar os efeitos do uso sistêmico da teriparatida na formação óssea sutural após a expansão da pré-maxila em ratos. Material e Métodos: Vinte ratos machos da raça Wistar (com oito a dez semanas de vida) foram divididos aleatoriamente em dois grupos: controle (C, n=10) e teriparatida (T, n=10). Uma força de expansão foi aplicada aos incisivos superiores, usando uma mola helicoidal, por um período de expansão de sete dias em ambos os grupos. No oitavo dia, os ratos iniciaram um período de sete dias de consolidação, nos quais 60 µg/kg de teriparatida foram administrados (uma vez ao dia), por via subcutânea, para o grupo T. Posteriormente, todos os ratos foram sacrificados e cortes histológicos corados com hemotolixina-eosina foram examinados. Por meio de análise imuno-histoquímica da região da sutura palatina mediana, avaliou-se a presença de anti-ostenectina, anti-osteocalcina, anti-fator de crescimento endotelial vascular (VEGF) e anti- fator transformador de crescimento (TGF-β). Resultados: Histologicamente, observou-se que o tecido ósseo recém-formado foi maior no grupo T do que no grupo C. O número de osteoblastos imunorreativos para anticorpos de osteonectina, osteocalcina e VEGF foi significativamente maior no grupo T do que no grupo C (p = 0,0001). O anticorpo TGF-β mostrou uma pequena reação no grupo T; porém, sem diferença significativa para o grupo C (p ˃ 0,05). Conclusão: O uso sistêmico de teriparatida após a expansão da sutura na região da pré-maxila pode estimular a formação óssea e melhorar a consolidação da expansão em ratos, por meio da regulação de osteonectina, osteocalcina e VEGF.

Arhgap21 Deficiency Results in Increase of Osteoblastic Lineage Cells in the Murine Bone Marrow Microenvironment.

ARHGAP21 is a member of the RhoGAP family of proteins involved in cell growth, differentiation, and adhesion. We have previously shown that the heterozygous Arhgap21 knockout mouse model (Arhgap21+/-) presents several alterations in the hematopoietic compartment, including increased frequency of hematopoietic stem and progenitor cells (HSPC) with impaired adhesion in vitro, increased mobilization to peripheral blood, and decreased engraftment after bone marrow transplantation. Although these HSPC functions strongly depend on their interactions with the components of the bone marrow (BM) niche, the role of ARHGAP21 in the marrow microenvironment has not yet been explored. In this study, we investigated the composition and function of the BM microenvironment in Arhgap21+/- mice. The BM of Arhgap21+/- mice presented a significant increase in the frequency of phenotypic osteoblastic lineage cells, with no differences in the frequencies of multipotent stromal cells or endothelial cells when compared to the BM of wild type mice. Arhgap21+/- BM cells had increased capacity of generating osteogenic colony-forming units (CFU-OB) in vitro and higher levels of osteocalcin were detected in the Arhgap21+/- BM supernatant. Increased expression of Col1a1, Ocn and decreased expression of Trap1 were observed after osteogenic differentiation of Arhgap21+/- BM cells. In addition, Arhgap21+/- mice recipients of normal BM cells showed decreased leucocyte numbers during transplantation recovery. Our data suggest participation of ARHGAP21 in the balanced composition of the BM microenvironment through the regulation of osteogenic differentiation.

Pattern of Adiponectin, Osteocalcin, Irisin, FGF-21, and MCP-1 According to the Body Size Phenotype: Could They Be Markers of Metabolic Health in Mexican-Mestizo Middle-Aged Women?

Variations in levels of some adipokines, myokines, osteokines, hepatokines and inflammatory cytokines contribute to abnormal glucose and lipid metabolism. The aim of this study was to determine the pattern of adiponectin, osteocalcin (OCN), irisin, FGF-21, and MCP-1 according to the body size phenotype of middle-aged women, and their associations with BMI, visceral adipose tissue (VAT), and HOMA-IR. A cross-sectional study in 265 women aged from 40 to 65 years was performed. The biochemical characteristics were evaluated in metabolically healthy normal weight, metabolically unhealthy normal weight, metabolically healthy obese, and metabolically unhealthy obese women. There was an association of OCN with BMI (r = -0.107; p = 0.047); adiponectin with BMI (r = -0.217; p = 0.001), insulin (r = -0.415; p = 0.0001), HOMA-IR (r = -0.429; p = 0.0001), and VAT (r = -0.134; p = 0.025); irisin with BMI (r = 0.604; p = 0.001), insulin (r = 0.446; p = 0.0001), HOMA-IR (r = 0.452; p = 0.0001), and VAT (r = 0.645; p = 0.0001); FGF-21 with insulin (r = -0.337; p= 0.030) and HOMA-IR (r = -0.341; p = 0.03); and MCP-1 with BMI (r = 0.481; p = 0.0001), VAT (r = 0.497; p = 0.001), insulin (r = 0.298; p= 0.001), and HOMA-IR (r = 0.255; p = 0.004). A multivariate analysis showed that an elevation of OCN (OR 1.4 (95%CI 1.06-1.81)) and a reduction of adiponectin (OR 0.9 (0.84-0.96)) were associated factors for a metabolic unhealthy phenotype in normal weight participants. Likewise, higher irisin (OR 1.007 (1.003-1.011)) and MCP-1 (1.044 (1.008-1.083)) were risk factors for a metabolic unhealthy phenotype in woman with obesity. OCN, adiponectin, irisin, FGF-21, and MCP-1 are associated with some metabolic parameters such as BMI, HOMA-IR, and VAT, and could be possible biomarkers of an unhealthy metabolic phenotype in middle-aged women.

Bone Mineral Density in Patients with Hepatic Glycogen Storage Diseases.

The association between bone mineral density (BMD) and hepatic glycogen storage diseases (GSDs) is still unclear. To evaluate the BMD of patients with GSD I, IIIa and IXα, a cross-sectional study was performed, including 23 patients (GSD Ia = 13, Ib = 5, IIIa = 2 and IXα = 3; median age = 11.9 years; IQ = 10.9-20.1) who underwent a dual-energy X-ray absorptiometry (DXA). Osteocalcin (OC, n = 18), procollagen type 1 N-terminal propeptide (P1NP, n = 19), collagen type 1 C-terminal telopeptide (CTX, n = 18) and 25-OH Vitamin D (n = 23) were also measured. The participants completed a 3-day food diary (n = 20). Low BMD was defined as a Z-score ≤ -2.0. All participants were receiving uncooked cornstarch (median dosage = 6.3 g/kg/day) at inclusion, and 11 (47.8%) presented good metabolic control. Three (13%) patients (GSD Ia = 1, with poor metabolic control; IIIa = 2, both with high CPK levels) had a BMD ≤ -2.0. CTX, OC and P1NP correlated negatively with body weight and age. 25-OH Vitamin D concentration was decreased in seven (30.4%) patients. Our data suggest that patients with hepatic GSDs may have low BMD, especially in the presence of muscular involvement and poor metabolic control. Systematic nutritional monitoring of these patients is essential.

Maternal adaptations of pancreatic islets and glucose metabolism after lactation.

Pancreatic islets adapt to metabolic requirements and the hormonal milieu by modifying their size and hormone secretions. Maternal glucose demands and hormonal changes occur after weaning, to rapidly re-establish bone mineralization. Minimal information exists about glucose metabolism and pancreatic islets after lactation. This study investigated islet morphology and glucose homeostasis for 14 days after lactation in C57BL/6NHHsd mice. Compared to the day of weaning, rapid increases in the islets' area and number of beta cells were found from the first day post-lactation, attaining maximum values on the third day post-weaning. These changes were accompanied by modifications in glucose-induced insulin secretion, glucose tolerance and insulin sensitivity. Islet-cell proliferation was already augmented before lactation ceased. Serum undercarboxylated osteocalcin concentrations increased significantly post-lactation; however, it is unlikely that this enhancement participates in earlier cell proliferation augmentation or in decreasing insulin sensitivity. Islet serotonin content was barely expressed, and serum calcium concentrations decreased. By the 14th day post-weaning, islets' area and glucose homeostasis returned to age-matched virgin mice levels. These findings recognize for the first time that increases in islet area and insulin secretion occur during physiological post-weaning conditions. These results open up new opportunities to identify molecules and mechanisms participating in these processes, which will help in developing strategies to combat diabetes.