Influence of obesity on microarchitecture and biomechanical properties in patients with hip fracture.

La obesidad y la osteoporosis (OP) son dos patologías muy prevalentes en nuestra sociedad actual. El efecto de la obesidad sobre la calidad ósea se encuentra en debate en la actualidad. Objetivo: Valorar el efecto del peso corporal sobre la microestructura y las propiedades biomecánicas de hueso trabecular procedente de biopsias de extremidad proximal de fémur de pacientes con fractura de cadera por fragilidad. Material y método: Estudio transversal de 16 pacientes con fractura de cadera. 2 grupos según su IMC: (A) sujetos normopeso y (B) con obesidad. Recogimos biopsias de hueso trabecular de cabeza femoral. Valoramos determinaciones bioquímicas (PTH, 25(OH) vitamina D e IGF-1), marcadores de remodelado óseo (PINP,CTX), masa ósea (DMO cuello y cadera total), microestructura ósea y estudio biomecánico (µCt). El análisis estadístico: t-Student (SPSS 22.0) significación p<0,05. Resultados: Todos los pacientes presentaron DMO de cadera en rango osteoporótico. El grupo de obesos presentó niveles superiores de PTH e inferiores de IGF-1, vitamina D y PINP. No encontramos diferencias en los parámetros relacionados con el metabolismo óseo. El grupo de obesos presentó mejores índices microestructurales alcanzando la significación: mayor volumen óseo (BV/TV: 36,6±12,7 vs. 19,4±11,4%, BS/TV: 5,5±1,1 vs. 3,9±1,3%), mayor número de trabéculas (Tb.N: 1,6±0,4 vs. 1,01±0,4), mayor anchura de trabéculas (Tb.Th: 0,22±0,003 vs. 0,17±0,05) y menor separación trabecular (Tb.Sp: 0,51±0,12 vs. 0,66±0,16). Los parámetros biomecánicos confirman una mayor resistencia del hueso trabecular en pacientes obesos. Conclusión: La obesidad puede ser un factor protector de la calidad ósea en la región femoral y tiene menos efecto sobre la densidad mineral ósea.Obesity and osteoporosis (OP) are two very prevalent diseases in our society today. The effect of obesity on bone quality is currently a subject under discussion. Objective: To assess the effect of body weight on the microstructure and biomechanical properties of trabecular bone biopsies from the proximal end of the femur in patients with hip fracture fragility. Material and methods: Cross-sectional study of 16 patients with hip fracture. The 2 groups are divided according to their BMI: (A) normal weight individuals and (B) those with obesity. We collected biopsies of cancellous bone from the femoral head and assessed biochemical determinations (PTH, 25 (OH) vitamin D and IGF-1), bone remodeling markers (PINP, CTX), bone mass (BMD neck and total hip), bone microstructure and biomechanical study (µCt). Statistical analysis: Student's t test (SPSS 22.0) significance p<0.05. Results: All patients had hip BMD in osteoporotic range. The obese group had higher levels of PTH and lower IGF-1, vitamin D and PINP. We found no differences in the parameters related to bone metabolism. The obese group showed better indices reaching microstructural significance: increased bone volume (BV/TV: 36.6±12.7 vs 19.4±11.4%, BS/TV: 5.5±1.1 vs 3.9±1.3%), higher trabecular number (Tb.N: 1.6±0.4 vs 1,01±0,4), greater trabecular width (Tb.Th: 0.22±0.003 vs 0.17±0.05) and lower trabecular separation (Tb.Sp: 0.51±0.12 vs 0.66±0.16). Biomechanical parameters confirm greater strength of trabecular bone in obese patients. Conclusion: Obesity may be a protective factor of bone quality in the femoral region and has less effect on bone mineral density

Influence of vitamin D on biomechanical microstructure and properties of patients with hip fracture.

Valorar niveles séricos de 25-hidroxivitamina D -25(OH)D-, hormonas con influencia sobre el metabolismo óseo (parathormona -PTH- y factor de crecimiento insulínico -IGF-I-), marcadores de remodelado óseo (MRO) (telopéptido carboxilo-terminal del colágeno tipo I -β-CTX- y propéptido aminoterminal del procolágeno tipo I -PINP-), densidad mineral ósea (DMO), microestructura y biomecánica de cuello de fémur, en pacientes con fractura de cadera osteoporótica (OP) vs. pacientes artrósicos (OA). Material y métodos: Estudio observacional transversal de 29 pacientes OP y 14 OA, edad ≥50 años. Cuantificamos niveles séricos hormonales y MRO (inmunoensayo), DMO de cadera (DXA), microestructura (micro-CT) y biomecánica (ensayos de compresión uniaxial, sistema IGFA). Análisis estadístico (SPSS 20.0.) Resultados: Los pacientes OP presentaron niveles inferiores de 25(OH)D (p=0,02) y DMO de cadera (p<0,05), y superiores de PTH (p=0,029) y de β-CTX (p=0,04). Los niveles de 25(OH)D se correlacionaron positivamente con IGF-I (p=0,04) y negativamente con β-CTX (p=0,003). Los valores de PTH se correlacionaron negativamente con DMO de cadera (p=0,0005) y positivamente con la separación trabecular (Tb.Th) (p=0,006). Los pacientes con niveles de 25(OH)D <20 ng/mL presentaron niveles mayores de β-CTX (p=0,006), menores de IGF-I (p=0,007) y Tb.Th (p=0,04). Conclusiones: Los niveles de vitamina D son bajos en población anciana, sobre todo en pacientes con fractura de cadera osteoporótica. Además, en estos pacientes existen niveles elevados de PTH y MRO y descendidos de DMO. Los pacientes cuyos niveles de 25(OH)D son inferiores a 20 ng/mL presentan un remodelado óseo más elevado, con menores niveles de IGF-I y alteraciones de la estructura ósea (Tb.Th) que puedan estar en relación con un mayor riesgo de fracturas.To assess serum levels of 25-hydroxyvitamin D-25 (OH) D-hormones with influence on bone metabolism (parathormone -PTH- and insulin-like growth factor (IGF)-I), bone remodeling markers (BRM) (carboxy-terminal telopeptide of collagen type I-β-CTX- and amino-peptide pro-peptide of procollagen type I -PINP), bone mineral density (BMD), microstructure and biomechanics of the femoral neck, in patients with osteoporotic hip fracture (OH) versus arthritic patients (OA). Material and methods: A cross-sectional observational study of 29 OH and 14 OA, age ≥50 years. We quantified hormonal serum levels and BRM (immunoassay), hip BMD (DXA), microstructure (micro-CT) and biomechanics (uniaxial compression tests, IGFA system). Analysis (SPSS 20.0.) Results: OH patients had lower levels of 25(OH)D (p=0.02) and hip BMD (p<0.05), and higher PTH (p=0.029) and β-CTX (p=0.04). Levels of 25(OH)D correlated positively with IGF-I (p=0.04) and negatively with β-CTX (p=0.003). The PTH values were correlated negatively with hip BMD (p=0.0005) and positively with trabecular thickness (TbTh) (p=0.006). Patients with 25(OH)D <20 ng/mL presented higher levels of β-CTX (p=0.006), lower IGF-I (p=0.007) and TbTh (p=0.04). Conclusions: Vitamin D levels are low in the elderly population, especially in patients with osteoporotic hip fracture. These patients also presented raised levels of PTH and BRM and descended from BMD. Patients whose 25(OH)D levels are below 20 ng/mL present higher bone remodeling, with lower levels of IGF-I and alterations of the bone structure (TbTh) that may be linked to a greater risk of fractures

Electrical Impedance of Surface Modified Porous Titanium Implants with Femtosecond Laser

The chemical composition and surface topography of titanium implants are essential to improve implant osseointegration. The present work studies a non-invasive alternative of electrical impedance spectroscopy for the characterization of the macroporosity inherent to the manufacturing process and the effect of the surface treatment with femtosecond laser of titanium discs. Osteoblasts cell culture growths on the titanium surfaces of the laser-treated discs were also studied with this method. The measurements obtained showed that the femtosecond laser treatment of the samples and cell culture produced a significant increase (around 50%) in the absolute value of the electrical impedance module, which could be characterized in a wide range of frequencies (being more relevant at 500 MHz). Results have revealed the potential of this measurement technique, in terms of advantages, in comparison to tiresome and expensive techniques, allowing semi-quantitatively relating impedance measurements to porosity content, as well as detecting the effect of surface modification, generated by laser treatment and cell culture.Ministry of Science and Innovation of Spain grant PID2019-109371GB-I00Junta de Andalucía–FEDER (Spain) Project US-1259771Junta de Andalucía-Proyecto de Excelencia (Spain) P18-FR-203

What are microRNAs? Potential biomarkers and therapeutic targets in osteoporosis

Los micro-ARN (miRs) son pequeñas moléculas de ARN no codificantes que regulan la expresión génica a nivel post-transcripcional. Generalmente actúan sobre la expresión genética mediante el silenciamiento o degradación de los ARNm, y están implicados en la regulación de varios procesos biológicos, como la diferenciación celular, la proliferación, la apoptosis y en el desarrollo embrionario y tisular. Actualmente son un importante foco de interés para el estudio de diversas enfermedades como el cáncer o la diabetes mellitus tipo 2. A nivel del metabolismo óseo, están surgiendo diversos miRs implicados en su regulación, abriendo un campo de investigación importante para identificar nuevos biomarcadores para el diagnóstico de la enfermedad osteoporótica, de su evolución, así como para diseñar nuevas terapias farmacológicas.Micro-RNAs (miRs) are small non-coding RNA molecules that regulate gene expression at post-transcriptional level. Generally, they act on gene expression by silencing or degrading mRNAs, and are involved in regulating various biological processes, such as cell differentiation, proliferation, apoptosis and in embryonic and tissue development. They are currently a major focus of interest in the study of various diseases such as cancer or type 2 diabetes mellitus. At level of bone metabolism, various miRs are emerging that are involved in their regulation, opening an important research field to identify new biomarkers for diagnosis of osteoporosis and its development, and to design new drug therapies

Modifying RANKL/OPG mRNA expression in differentiating and growing human primary osteoblasts

The OPG / RANKL system in primary cultures of human osteoblasts has been studied by di ff erent authors. However, very few studies have been performed on gene expression of RANKL and OPG at di ff erent stages of maturation on human osteoblast cultures.The e ff ect of 17-estradiol and 1,25dihydroxyvitamin D3 on the OPG / RANKL system is not known during the di ff erent states of cellular maturation. In this work we quantifi ed OPG and RANKL protein levels (ELISA) and the mRNA of OPG, RANKL, collagen type I, alkaline phosphatase, and osteocalcin (semi-quantita-tive RT-PCR) in human osteoblasts. We analyzed these in basal conditions and after incubation with 17- -estradiol and 1,25dihydroxyvitamin D3 in the first and second phases. We found that OPG secretion and expression levels increased throughout cellular growth.RANKL proteins were detected only in the first stage, and the expression increased throughout the first phase. Thus, the RANKL / OPG ratio was higher in immature osteoblasts than in mature osteoblasts. The evolution of RANKL gene expression was related to collagen I and alkaline phosphatase, while OPG was related to osteocalcin. We observed no modi fi cations after estradiol and 1,25dihydroxyvitamin D3 treatment. Our results suggest that the OB is a positive stimulator at precocious stages of diff erentiation on osteoclastogenic modulates

DC electrical stimulation enhances proliferation and differentiation on N2a and MC3T3 cell lines

Background: Electrical stimulation is a novel tool to promote the differentiation and proliferation of precursor cells. In this work we have studied the effects of direct current (DC) electrical stimulation on neuroblastoma (N2a) and osteoblast (MC3T3) cell lines as a model for nervous and bone tissue regeneration, respectively. We have developed the electronics and encapsulation of a proposed stimulation system and designed a setup and protocol to stimulate cell cultures. Methods: Cell cultures were subjected to several assays to assess the effects of electrical stimulation on them. N2a cells were analyzed using microscope images and an inmunofluorescence assay, differentiated cells were counted and neurites were measured. MC3T3 cells were subjected to an AlamarBlue assay for viability, ALP activity was measured, and a real time PCR was carried out. Results: Our results show that electrically stimulated cells had more tendency to differentiate in both cell lines when compared to non-stimulated cultures, paired with a promotion of neurite growth and polarization in N2a cells and an increase in proliferation in MC3T3 cell line. Conclusions: These results prove the effectiveness of electrical stimulation as a tool for tissue engineering and regenerative medicine, both for neural and bone injuries. Bone progenitor cells submitted to electrical stimulation have a higher tendency to differentiate and proliferate, filling the gaps present in injuries. On the other hand, neuronal progenitor cells differentiate, and their neurites can be polarized to follow the electric field applied.Universidad de Sevilla US-1380661Junta de Andalucía P18-FR-2038. 2020–202

Biocompatibility and osseointegration study of new prosthetic materials

Objetivos Los implantes óseos son utilizados cada vez con mayor frecuencia en la práctica clínica y, entre los materiales, el Ti o sus aleaciones son los de mejor rendimiento por sus propiedades fisicoquímicas. Aleaciones como TiNbTa han demostrado mejorar las características biomecánicas del Ti puro comercial (c.p.), sin embargo, su capacidad osteointegradora necesita ser evaluada. El objetivo del presente estudio fue valorar la citotoxicidad y la capacidad de adhesión, proliferación y diferenciación de células osteoblásticas en cultivo, influida por discos de material TiNbTa frente a Ti c.p. Material y métodos Analizamos a los 4 y 7 días del cultivo la línea celular MC3T3, la viabilidad celular (AlamarBlue Cell Viability Reagent. Invitrogen, España), así como la proliferación y diferenciación celular (actividad de fosfatasa alcalina (ALP) y microscopía electrónica de barrido (Fijación para SEM). Se realizó la prueba t de Student para determinar diferencias estadísticamente significativas entre los dos grupos de discos de estudio. Resultados Los resultados obtenidos demuestran muy buena viabilidad celular durante el periodo de estudio, sin diferencias significativas para ambos materiales. Así mismo, detectamos una caída en los niveles de ALP que fue significativa para ambos componentes entre los días 4 y 7 del estudio (p<0,05). Las imágenes de microscopía electrónica revelaron buena capacidad de adhesión al material, así como diferenciación celular frente a ambos tipos de discos. Conclusiones La aleación de TiNbTa como material para implantes óseos cuenta con una buena capacidad osteointegradora, además de resolver problemas de biomecánica que presenta el titanio puro como componente.Objetive Bone implants are increasingly used in clinical practice and, among the materials, Ti or its alloys are offer the best performance given their physicochemical properties. Alloys such as TiNbTa have been shown to improve the biomechanical characteristics of commercial pure Ti (c.p.), however, its osseointegration capacity needs to be evaluated. The objective of the present study was to assess the cytotoxicity and the adhesion, proliferation and differentiation capacity of osteoblastic cells in culture, influenced by discs of TiNbTa material versus Ti c.p. Material and methods At 4 and 7 days after culture, we analyzed the MC3T3 cell line, cell viability (AlamarBlue Cell Viability Reagent. Invitrogen, Spain), as well as cell proliferation and differentiation (alkaline phosphatase activity (ALP) and scanning electron microscopy (Fixation for SEM) Student's t test was performed to determine statistically significant differences between the two groups of study discs. Results The results obtained show very good cell viability during the study period, with no significant differences for both materials. Likewise, we detected a drop in ALP levels that was significant for both components between days 4 and 7 of the study (p <0.05). Electron microscopy images revealed good adhesion capacity to the material, as well as cell differentiation against both types of discs. Conclusions The TiNbTa alloy as a material for bone implants offers good osseointegrative capacity, in addition to solving biomechanical problems that pure titanium presents as a component

Surface Modification of Porous Titanium Discs Using Femtosecond Laser Structuring

The failure of titanium implants is associated with two main problems that include the bone resorption and fracture of the surrounding bone tissue (stiffness incompatibility) and implant loosening (poor osseointegration). The development of porous titanium implants with low Young modulus solve the stress shielding phenomenon, while the modification of the implant surface must be implemented to promote a fast bond between the implant and bone. In this work, femtosecond laser micromachining was applied to modify the topography of the surface of Ti porous samples obtained by a space-holder technique to obtain hierarchical structures (micro and nano roughness patterns) to enhance osseointegration. Scanning electron microscopy, confocal laser microscopy, andimageanalysiswereusedforcharacterizationofthesurfacemorphology,roughness,andporosity before and after performing the laser treatment. Based on these results, the effect of the treatment on the mechanical behavior of the samples was estimated. In addition, a preliminary in-vitro test was performed to verify the adhesion of osteoblasts (filopodia presence) on modified titanium surface. Resultsrevealedthatlasertexturinggeneratedclustersofmicro-holesandmicro-columnsbothonthe flat surface of the samples and inside the macro-pores, and periodic nanometric structures across the entire surface. The porous substrate offers suitable biomechanics (stiffness and yield strength) and bio-functional behavior (bone ingrowth and osseointegration), which improves the clinic success of titanium implants.FEDER-Junta de Andalucía Research Project US-125977