João Felipe Bezerra

Protection against T1DM-Induced Bone Loss by Zinc Supplementation: Biomechanical, Histomorphometric, and Molecular Analyses in STZ-Induced Diabetic Rats

Several studies have established an association between diabetes and alterations in bone metabolism; however, the underlying mechanism is not well established. Although zinc is recognized as a potential preventive agent against diabetes-induced bone loss, there is no evidence demonstrating its effect in chronic diabetic conditions. This study evaluated the effects of zinc supplementation in a chronic (90 days) type 1 diabetes-induced bone-loss model. Male Wistar rats were distributed in three groups: control, type 1 diabetes mellitus (T1DM), and T1DM plus zinc supplementation (T1DMS). Serum biochemical analysis; tibia histomorphometric, biomechanical, and collagen-content analyses; and femur mRNA expression were evaluated. Relative to T1DM, the zinc-supplemented group showed increased histomorphometric parameters such as TbWi and BAr and decreased TbSp, increased biomechanical parameters (maximum load, stiffness, ultimate strain, and Young’s modulus), and increased type I collagen content. Interestingly, similar values for these parameters were observed between the T1DMS and control groups. These results demonstrate the protective effect of zinc on the maintenance of bone strength and flexibility. In addition, downregulation of OPG, COL1A, and MMP-9 genes was observed in T1DMS, and the anabolic effects of zinc were evidenced by increased OC expression and serum ALP activity, both related to osteoblastogenesis, demonstrating a positive effect on bone formation. In contrast, T1DM showed excessive bone loss, observed through reduced histomorphometric and biomechanical parameters, characterizing diabetes-associated bone loss. The bone loss was also observed through upregulation of OPG, COL1A, and MMP-9 genes. In conclusion, zinc showed a positive effect on the maintenance of bone architecture and biomechanical parameters. Indeed, OC upregulation and control of expression of OPG, COL1A, and MMP-9 mRNAs, even in chronic hyperglycemia, support an anabolic and protective effect of zinc under chronic diabetic conditions. Furthermore, these results indicate that zinc supplementation could act as a complementary therapy in chronic T1DM.

Association of polymorphisms in IL6 gene promoter region with type 1 diabetes and increased albumin-to-creatinine ratio.

Pro‐inflammatory cytokines, such as interleukin‐6 (IL‐6), have been considered as key factors in type 1 diabetes mellitus (T1DM) and diabetic nephropathy, thus, our aim was to investigate the association of IL6‐174G>C (rs1800795) and ‐634C>G (rs1800796) polymorphisms with T1DM susceptibility and diabetic nephropathy.

Anabolic Effect of Insulin Therapy on the Bone: Osteoprotegerin and Osteocalcin Up-Regulation in Streptozotocin-Induced Diabetic Rats.

Type 1 diabetes mellitus (T1DM) is associated with several skeletal alterations, particularly in conditions of poor glycaemic control. Insulin therapy is the major conservative treatment for T1DM; however, the effects of this hormone on bone markers of T1DM rats are limited, and the regulatory mechanisms remain elusive. Therefore, the evaluation of molecular and non‐molecular parameters in a chronic animal model of T1DM‐induced bone loss, treated with and without insulin, may help in elucidating the insulin mechanisms. Male Wistar rats were assigned into three groups: control, T1DM (T1DM rats induced with streptozotocin [STZ] at 40 mg/kg intravenously) and T1DM plus insulin therapy (T1DMI). After 8 weeks, we evaluated the serum biochemical, tibia histomorphometric and biomechanical parameters, as well as the gene expression of the receptor activator of nuclear factor kappa‐B ligand (RANKL), osteoprotegerin (OPG) and osteocalcin (OC) of femur mRNA. Compared with T1DM, the T1DMI group showed less bone loss, which was revealed by the increased trabecular width (TbWi, p < 0.001) and trabecular bone area (BAr, p < 0.01), reduced trabecular separation (TbSp, p < 0.01) and increased Youngs modulus (p < 0.05). Moreover, molecular analyses indicated that the expression of OPG and OC was up‐regulated (p < 0.001 and p < 0.05, respectively). In summary, the up‐regulation of OPG and OC in the T1DMI group supports an anabolic effect of insulin, which was demonstrated by the maintenance of bone architecture and flexibility. These results suggest that insulin therapy may prevent T1DM‐induced bone loss via the effects on the bone formation.

Zinc supplementation reduces RANKL/OPG ratio and prevents bone architecture alterations in ovariectomized and type 1 diabetic rats

Type 1 diabetes mellitus (T1DM) and estrogen deficiency are associated with several alterations in bone turnover. Zinc (Zn) is required for growth, development, and overall health. Zinc has been used in complementary therapy against bone loss in several diseases. We hypothesized that Zn supplementation represents a potential therapy against severe bone loss induced by the combined effect of estrogen deficiency and T1DM. We evaluated the protective effect of Zn against bone alterations in a chronic model of these disorders. Female Wistar rats were ramdomized into 3 groups (5 rats each): control, OVX/T1DM (ovariectomized rats with streptozotocin-induced T1DM), and OVX/T1DM+Zn (OVX/T1DM plus daily Zn supplementation). Serum biochemical, bone histomorphometric, and molecular analyses were performed. Histomorphometric parameters were similar between the control and OVX/T1DM+Zn groups, suggesting that Zn prevents bone architecture alterations. In contrast, the OVX/T1DM group showed significantly lower trabecular width and bone area as well as greater trabecular separation than the control. The OVX/T1DM and OVX/T1DM+Zn groups had significantly higher serum alkaline phosphatase activity than the control. The supplemented group had higher levels of serum-ionized calcium and phosphorus than the nonsupplemented group. The RANKL/OPG ratio was similar between the control and OVX/T1DM+Zn groups, whereas it was higher in the OVX/T1DM group. In conclusion, Zn supplementation prevents bone alteration in chronic OVX/T1DM rats, as demonstrated by the reduced RANKL/OPG ratio and preservation of bone architecture. The findings may represent a novel therapeutic approach to preventing OVX/T1DM-induced bone alterations.

Efeito do tamoxifeno no perfil das proteínas plasmáticas em condição de diabetes mellitus tipo 1

PURPOSE Considering that important scientific advances have been obtained through studies based on experimental Diabetes mellitus, and that tamoxifen action in humans remains unknown, the aim of the present work is to follow the modifications promoted by diabetes and tamoxifen in the electrophoretic profile of plasmatic proteins. METHODS It was used 27 Wistar female rats (180-250 body weight), randomicaly divided into five groups: C1 (n = 3, received vehicle), C2 (n = 3, no treatment), T (n =5, treated with tamoxifen, 0.3mg/Kg/day), D (n = 8, experimental diabetes by estreptozotocin, 45mg/Kg and DT (n = 8, diabetic treated with tamoxifen). The electrophoresis was accomplished in cellulose acetate. pH 8.6-8.8, TECNOW chamber, and the strains were stained by Ponceau S. The total proteins were determined by the Biuret method (Labtest). Proteinograms were obtained in densitometer BioSystems BTS-235. RESULTS Albumin decreased progressively in the groups T, D and DT; a1 fraction increased in groups T and DT; a2 fraction increased in groups T and D, including a synergic effect in group DT; a fraction increased in groups T and D; a fraction increased in groups T, D and DT. CONCLUSIONS The results indicate an acute phase resposta, with synergic effect of tamoxifen and diabetes, suggesting a probable hepatic lesion.

Application of high-resolution array platform for genome-wide copy number variation analysis in patients with nonsyndromic cleft lip and palate

Although more than 14 loci may be involved in the development of nonsyndromic cleft lip and palate (NSCLP), the etiology has not been fully elucidated due to genetic and environmental risk factor interactions. Despite advances in identifying genes associated with the NSCLP development using traditional genetic mapping strategies of candidate genes, genome‐wide studies, and epidemiologic and linkage analysis, microarray techniques have become important complementary tools in the search for potential causative oral clefts genes in genetic studies. Microarray hybridization enables scanning of the whole genome and detecting copy number variants (CNVs). Although common benign CNVs are often smaller, with sizes smaller than 20 kb, here we reveal small exonic CNVs based on the importance of the encompassed genes in cleft lip and palate phenotype.

Cardiovascular abnormalities in patients with oral cleft: a clinical-electrocardiographic-echocardiographic study

OBJECTIVES: The present study aims to describe the clinical, electrocardiographic, and echocardiographic cardiological findings in a group of patients with oral clefts. METHODS: This is a prospective cross-sectional study on 70 children (age range from 13 days to 19 years) with oral clefts who attended the multidisciplinary program of a university hospital from March 2013 to September 2014. The patients were evaluated by a pediatric cardiologist and underwent detailed anamnesis, physical examination, electrocardiogram, and echocardiogram. RESULTS: Sixty percent of the patients were male; 55.7% presented with cleft lip and palate, and 40.0% presented with health complaints. Comorbidities were found in 44.3%. Relevant pregnancy, neonatal, family and personal antecedents were present in 55.7%, 27.1%, 67.2%, and 24.3% of the patients, respectively. Regarding the antecedents, 15.2% of the patients presented with a cardiac murmur, 49.0% with a familial risk of developing plurimetabolic syndrome, and 6% with family antecedents of rheumatic fever. Electrocardiographic evaluation showed one case of atrioventricular block. Echocardiograms were abnormal in 35.7% of the exams, including 5 cases of mitral valve prolapse — one of which was diagnosed with rheumatic heart disease. CONCLUSION: The finding of a family risk of developing plurimetabolic syndrome and a diagnosis of rheumatic heart disease indicates that patients with oral clefts may be more prone to developing acquired heart disease. Thus, our findings highlight the importance of anamnesis and methodological triangulation (clinical-electrocardiographic-echocardiographic) in the investigation of patients with oral clefts and emphasize that cardiological follow-up to evaluate acquired and/or rhythm heart diseases is necessary. This strategy permits comorbidity prevention and individualized planned treatment.

Risk factors and comorbidities in Brazilian patients with orofacial clefts

Considering that environmental risk factors substantially contribute to the etiology of orofacial clefts and that knowledge about the characteristics and comorbidities associated with oral clefts is fundamental to promoting better quality of life, this study aimed to describe the risk factors, main characteristics, and comorbidities of a group of patients with cleft lip and/or cleft palate (CL/P) from Rio Grande do Norte (RN), Brazil. Data were obtained from 173 patients with CL/P using a form from the Brazilian database on Orofacial Clefts. Most patients were male with cleft lip and palate and had a normal size and weight at birth; presented few neonatal intercurrent events; and had anemia and respiratory and cardiovascular diseases as main associated comorbidities. They also required timely surgical rehabilitation and multidisciplinary care to stimulate their neuropsychomotor development. In addition, a high frequency of familial recurrence and of parental consanguinity was evidenced in the studied population, especially for the cleft lip and cleft palate type. Other relevant findings were the considerable maternal exposure to alcohol, infections, smoking, and hypertension, as well as low supplementation with vitamins and minerals and deliberate consumption of analgesics, antibiotics, and antihypertensives during pregnancy. Characterization of the CL/P patient profile is essential for the planning of health services and integration among the health professionals involved in the diagnosis and treatment of these malformations. Our results reinforce the need for additional research to confirm the association between environmental factors and the development of orofacial clefts.