Geniposide Ameliorated Dexamethasone-Induced Cholesterol Accumulation in Osteoblasts by Mediating the GLP-1R/ABCA1 Axis

Background: Overexposure to glucocorticoid (GC) produces various clinical complications, including osteoporosis (OP), dyslipidemia, and hypercholesterolemia. Geniposide (GEN) is a natural iridoid compound isolated from Eucommia ulmoides. Our previous study found that GEN could alleviate dexamethasone (DEX)-induced differentiation inhibition of MC3T3-E1 cells. However, whether GEN protected against Dex-induced cholesterol accumulation in osteoblasts was still unclear. Methods: DEX was used to induce rat OP. Micro-CT data was obtained. The ALP activity and mineralization were determined by the staining assays, and the total intracellular cholesterol was determined by the ELISA kits. The protein expression was detected by western blot. Results: GEN ameliorated Dex-induced micro-structure damages and cell differentiation inhibition in the bone trabecula in rats. In MC3T3-E1 cells, Dex enhanced the total intracellular cholesterol, which reduced the activity of cell proliferation and differentiation. Effectively, GEN decreased DEX-induced cholesterol accumulation, enhanced cell differentiation, and upregulated the expression of the GLP-1R/ABCA1 axis. In addition, inhibition of ABAC1 expression reversed the actions of GEN. Treatment with Exendin9-39, a GLP-1R inhibitor, could abrogate the protective activity of GEN. Conclusions: GEN ameliorated Dex-induced accumulation of cholesterol and inhibition of cell differentiation by mediating the GLP-1R/ABCA1 axis in MC3T3-E1 cells.

Evaluasi Radiografi terhadap Kepadatan Tulang Hasil Implan Scaffold 3D Bifasik Kalsium Fosfat Alginat pada Domba Lokal (Ovis aries)

This study aims to; 1) evaluate the osteogenesis ability of Biphasic Calcium Phosphate (BCP)-alginate 70/30 based on chages in size of defect and relative bone density, 2) implant response tissue based on relative muscle density peri implant. The BCP used in this study made from hydroxyapatite (HA) 70, tricalcium phosphate (TCP) 30 and alginate as porogen. The implant material made in 3D form and have diameter porous 200-400 ìm which similar size with the bone trabecula. This study used three local sheep (Ovis aries) male 1.5 years old, weight average 20 kg. The implant material is grown on the right sheep tibia. As a control, the left tibia is made defective with no implantable material. Evaluate osteogenesis and implant tissue responses using radiography and assess the grey values of a particular area. Osteogenesis is also assessed based on the size of the defect. Based on the size of the defect, the right defect shows no significant value. The size of the left defect shows a significant value between day 0 and day 30, and 90. The relative bone density of the right defect shows a significant value between day 7 and 30. In the left defect, the relative bone density shows the significant between 0 and 90. The relative density of the muscle does not show significant values either on the right or left defects.

High NESTIN Expression Marks the Endosteal Capillary Network in Human Bone Marrow

Hematopoiesis is hosted, supported and regulated by a special bone marrow (BM) microenvironment known as “niche.” BM niches have been classified based on micro-anatomic distance from the bone surface into “endosteal” and “central” niches. Whilst different blood vessels have been found in both BM niches in mice, our knowledge of the human BM architecture is much more limited. Here, we have used a combination of markers including NESTIN, CD146, and αSMA labeling different blood vessels in benign human BM. Applying immunohistochemical/immunofluorescence techniques on BM trephines and performing image analysis on almost 300 microphotographs, we detected high NESTIN expression in BM endothelial cells (BMECs) of small arteries (A) and endosteal arterioles (EA), and also in very small vessels we named NESTIN+ capillary-like tubes (NCLTs), not surrounded by sub-endothelial perivascular cells that occasionally reported low levels of NESTIN expression. Statistically, NCLTs were detected within 40 μm from bone trabecula, frequently found in direct contact to the bone line and spatially correlated with hematopoietic stem/progenitor cells. Our results support the expression of NESTIN in human BMECs of EA and A in accordance with the updated classification of murine BM micro-vessels. NCLTs for their peculiar characteristics and micro-anatomical localization have been here proposed as transitional vessels possibly involved in regulating human hematopoiesis.

Gene expression profiling of the bone trabecula in patients with osteonecrosis of the femoral head by RNA sequencing

Early diagnosis and treatment of osteonecrosis of the femoral head (ONFH) is challenging. Bone trabecula play a vital role in the severity and progression of ONFH. In the present study, the investigators used gene expression profiling of bone trabecula to investigate gene alterations in ONFH patients. Osteonecrotic bone trabecula (ONBT) such as necrosis, fibrosis, and lacuna were confirmed by histological examination in the patients. The adjacent ‘normal’ bone trabecula (ANBT) did not show any pathological changes. Gene sequencing data revealed that although ANBT showed no significant histological changes, alteration of mRNA profiling in ANBT was observed, similar to that in ONBT. Our results indicated that the alteration of mRNA profiling in ANBT may cause normal bone tissue to develop into necrotic bone. RNA-seq data indicated that 2,297 differentially abundant mRNAs were found in the ONBT group (1,032 upregulated and 1,265 downregulated) and 1,523 differentially abundant mRNAs in the ANBT group (744 upregulated and 799 downregulated) compared with the healthy control group. Gene ontology (GO) enrichment analysis suggested that fatty acid metabolism and degradation were the main zones enriched with differentially expressed genes (DEG). Kyoto encyclopedia of genes and genomes (KEGG) pathway enrichment analysis indicated that peroxisome proliferator-activated receptor γ (PPAR-γ) pathway was the most significantly regulated pathway. Lipocalin-2 (LCN2), an osteoblast-enriched secreted protein, was significantly decreased in ONBT suggesting that downregulation of LCN2 might affect lipid metabolism and lead to hyperlipidemia, and thus promote pathogenesis of ONFH.

The effect of local changes in ionic and enzymatic homeostasis induced by polycaprolactone scaffolds mineralized with vaterite on bone tissue regeneration

Цель исследования - оценка влияния на регенерацию костной ткани локальных изменений ферментативного и ионного гомеостаза минерализованными ватеритом скаффолдами из поликапролакона. Методика. Эксперимент выполнен на 47 нелинейных крысах-самцах. Животные были разделены на 4 экспериментальные группы: контроль (интактные животные), группа сравнения (ложнооперированные крысы), группа отрицательного контроля (крысы с имплантацией матрицы, на которой сорбирован нативный овальбумин) и экспериментальная группа - крысы, которым имплантировали поликапролактоновый скаффолд, минерализованный ватеритом (ПКЛ/СаСО3-скаффолд) с адсорбированной щелочной фосфатазой в дефект бедренной кости. На 7-е и 28-е сут в крови определяли содержание щелочной фосфатазы, а также концентрацию остеокальцина. Количественное определение остеокальцина производили методом иммуноферментного анализа с использованием набора реактивов фирмы «Immunodiagnostic systems» (Франция) и полуавтоматического ИФА-анализатора «Anthos Reader 2020» («Biochrom», Великобритания). Для выявления изменений ферментативной активности в сыворотке крови определяли кинетическим методом активность щелочной фосфатазы на биохимическом анализаторе «Сапфир-400» («Hirose Electronic System», Япония) с использованием набора реактивов фирмы «ДиаС» (Россия). Морфологическую верификацию процессов остеогенеза осуществляли при изучении гистологических препаратов на 28-е сут эксперимента. Поперечные срезы диафиза бедренной кости изучали при помощи микроскопа AxioImager Z2 («Carl Zeiss», Германия) и микровизора проходящего света серии μVizo-103 (ООО «ЛОМО ФОТОНИКА», Россия). Оценивали интеграцию матриц с краями пропила, изменения кости в области имплантации матриц, преобладающий тип клеточных элементов матриц, наличие в них костных балок и сосудов. Результаты. Установлено, что изменение локального ионного гомеостаза (СаСО3) и ферментативного гомеостаза щелочной фосфатазы, вызывает в матрице имплантированной в область дефекта формирование костных балок и биохимические изменения в крови, характеризующиеся повышением активности щелочной фосфатазы и увеличением концентрации остеоакальцина. Заключение. При имплантации ПКЛ/СаСО3-матриц с щелочной фосфатазой локальные изменения ферментативного и ионного гомеостаза, вызванные введением в состав кальцийсодержащих скаффолдов щелочной фосфатазы, в значительной степени стимулируют остеогенез. Биосовместимость и остеоиндуктивные свойства ПКЛ/СаСО3-матриц с щелочной фосфатазой позволяют говорить о перспективности их клинической апробации для стимуляции процессов регенерации костной ткани. Aim. To study the effect of local changes in enzymatic and ionic homeostasis induced by vaterite-mineralized polycaprolactone scaffolds with adsorbed alkaline phosphatase on bone tissue regeneration. The experiment was conducted on 47 mongrel male rats divided into four experimental groups: control (intact animals), comparison group (sham-operated rats), negative control (rats implanted with a matrix with adsorbed native ovabumin), and experimental group (rats implanted with PLC-CaCO3-scaffold with adsorbed alkaline phosphatase into the femoral bone defect). Alkaline phosphatase and osteocalcin contents were measured in blood of all animals on days 7 and 28. Osteocalcin was measured using the immune enzyme assay with an Immunodiagnostic Systems (France) kit and semi-automatic immune Anthos Reader 2020 (Biochrom, Great Britain) enzyme analyzer. Changes in serum alkaline phosphatase activity were detected using the kinetic method with a Sapphire-400 biochemical analyzer (Hirose Electronic Systems, Japan) and a DiaC (Russia) kit. Osteogenesis processes were verified morphologically on day 28 of the experiment by a histological assay. Transversal sections of the femoral diaphysis were examined with an AxioImager Z2 (Carl Zeiss, Germany) microscope and a µVizo-103 transmitted-light microvisor (OOO LOMO FOTONIKA, Russia). The evaluated indexes included matrix integration with saw-cut edges; bone changes in the area of matrix implantation; predominating type of matrix cell elements and the presence there of bone trabecula and blood vessels. Results. Changes in the local ionic homeostasis and homeostatic changes of alkaline phosphatase induced formation of bone trabecula in the matrix implanted into the defect area and biochemical changes in blood evident as upregulation of serum alkaline phosphatase associated with increased osteocalcin. Conclusion. In implantation of PLC-CaCO3 matrices with alkaline phosphatase, local changes in the enzymatic and ionic homeostasis induced by supplementation of calcium-containing scaffolds with alkaline phosphatase considerably stimulate osteogenesis. The biocompatibility and osteoinductive properties of PLC-CaCO3 matrices with alkaline phosphatase suggested that clinical testing of the use of the matrices for stimulation of bone tissue regeneration would be promising.

The Experimental Study on Promoting the Ilizarov Distraction Osteogenesis by the Injection of Liquid Alg/nHAC Biocomposites

Limb lengthening is frequently utilized in treating limb length inequalities, angulation deformities, nonunions, complex fractures, and deficiencies after tumor resection in more recent year. The procedure of limb lengthening pioneered by Ilizarov is now a widely accepted method for correcting limb length inequality and short stature as well as for bridging large defects in long bones. In order to promote bone healing during distraction osteogenesis and reduce the complications caused by limb lengthening pioneered, an alginate/nanohydroxyapatite/collagen (Alg/nHAC) composite was fabricated. General observation, histologically morphological observations, X-ray examination, biomechanical test, bone density, and the percentage area of bone trabecula were used to assay the ability of Alg/nHAC composite to promote bone healing. The present study demonstrates that the injection of liquid Alg/nHAC composites can significantly promote distraction osteogenesis. Alg/nHAC composite is promising for clinical application, solving the healing problem of backbone osteotomy and the fixing problem of metaphyseal backbone.

Bone Trabecula Surface Reconstruction for Cranium Scaffold in Bone Tissue Engineering

In bone tissue engineering, the scaffold architecture is very important for cell growth, it is better to make it similar with the native bone trabecula. To imitate the nature bone morphology, this paper presents a 3D reconstruction method of bone trabecula surface for cranium scaffold. Firstly, a native human cranium specimen on forehead was scanned by micro CT equipment and a set of gray level images were obtained. Then through image denoising, image enhancement, contour extraction and triangular surface reconstruction, the 3D structure of the specimen and its internal bone trabecula were reconstructed successfully. Lastly, to evaluate the feasibility of the method, a biomaterial scaffold case was fabrication using lost-foam casting technology. Results shown that the bone trabecula architecture in the scaffold is best retained and the porous structure is highly similar with the native specimen. This reconstructions process is simple and objective, which provide a new way for clinic cranium restoration.