The Protective Effect of Chronic Intermittent Hypobaric Hypoxia on Preventing the Destruction of CD34+ Haematopoietic Stem Cells in Aplastic Anaemia by Modulating the Th1/Th2 Balance.

Aplastic anaemia (AA) is a haematopoietic disorder caused by immune-mediated attack on haematopoietic stem cells (HSCs). Stem cell transplantation and immunosuppressive therapy remain the major treatment choice for AA patients but have limited benefits and undesired side effects. The aim of our study was to clarify the protective role of immunity of chronic intermittent hypobaric hypoxia (CIHH) and the underlying mechanism in AA. Our integrative analysis demonstrated that CIHH pre-treatment significantly improved haematopoiesis and survival in an AA rat model. We further confirmed that CIHH pre-treatment was closely associated with the Th1/Th2 balance and a large number of negative regulatory haematopoietic factors, such as TNF-α and IFN-γ, produced by hyperactive Th1 lymphocytes released in AA rats, which induced the death program in a large number of CD34+ HSCs by activating the Fas/FasL apoptosis pathway, while CIHH pre-treatment effectively downregulated the expression of TNF-α and IFN-γ, resulting in a reduction in Fas antigen expression in CD34+ HSCs. In summary, this study provides evidence that CIHH has good protective effect against AA by modulating immune balance in Th1/Th2 cells and may provide a new therapeutic strategy.

Anti-resorption role of low-intensity pulsed ultrasound (LIPUS) during large-scale bone reconstruction using porous titanium alloy scaffolds through inhibiting osteoclast differentiation.

BACKGROUND: Ti6Al4V biomaterials combine with low-intensity pulsed ultrasound (LIPUS) has been reported with great bone regeneration capacity. It is important to better understand how LIPUS benefits bone microenvironment to seek for target of therapeutic medicine. Osteoclast differentiation plays a crucial role in bone resorption. Recent advances in molecular biology have revealed that N6-methyladenosine (m6A) RNA modifications can modulate biological processes, but their role in bone biology, particularly in osteoclast differentiation, remains unclear. We aim to understand how LIPUS regulates bone microenvironment especially osteoclast formation during bone regeneration to provide new therapeutic options for preventing and delaying bone resorption, thus with better bone regeneration efficiency. RESULTS: 1. LIPUS promoted bone ingrowth and bone maturity while inhibiting osteoclast formation within Ti6Al4V scaffolds in large-scale bone defect model. 2. LIPUS was found to inhibit osteoclast differentiation by decreasing the overall expression of osteoclast markers in vitro. 3. LIPUS decreases RNA m6A-modification level through upregulating FTO expression during osteoclast differentiation during. 4. Inhibiting FTO expression and function leads to less inhibition during osteoclast differentiation. CONCLUSION: LIPUS suppresses osteoclast differentiation during bone regeneration through reducing m6A modification of osteoclastic RNAs by up regulating FTO expression.

Piezo1 channel activation in response to mechanobiological acoustic radiation force in osteoblastic cells.

Mechanobiological stimuli, such as low-intensity pulsed ultrasound (LIPUS), have been shown to promote bone regeneration and fresh fracture repair, but the fundamental biophysical mechanisms involved remain elusive. Here, we propose that a mechanosensitive ion channel of Piezo1 plays a pivotal role in the noninvasive ultrasound-induced mechanical transduction pathway to trigger downstream cellular signal processes. This study aims to investigate the expression and role of Piezo1 in MC3T3-E1 cells after LIPUS treatment. Immunofluorescence analysis shows that Piezo1 was present on MC3T3-E1 cells and could be ablated by shRNA transfection. MC3T3-E1 cell migration and proliferation were significantly increased by LIPUS stimulation, and knockdown of Piezo1 restricted the increase in cell migration and proliferation. After labeling with Fluo-8, MC3T3-E1 cells exhibited fluorescence intensity traces with several high peaks compared with the baseline during LIPUS stimulation. No obvious change in the fluorescence intensity tendency was observed after LIPUS stimulation in shRNA-Piezo1 cells, which was similar to the results in the GsMTx4-treated group. The phosphorylation ratio of ERK1/2 in MC3T3-E1 cells was significantly increased (P < 0.01) after LIPUS stimulation. In addition, Phalloidin-iFluor-labeled F-actin filaments immediately accumulated in the perinuclear region after LIPUS stimulation, continued for 5 min, and then returned to their initial levels at 30 min. These results suggest that Piezo1 can transduce LIPUS-induced mechanical signals into intracellular calcium. The influx of Ca2+ serves as a second messenger to activate ERK1/2 phosphorylation and perinuclear F-actin filament polymerization, which regulate the proliferation of MC3T3-E1 cells.

Surface-treated 3D printed Ti-6Al-4V scaffolds with enhanced bone regeneration performance: an in vivo study.

BACKGROUND: Given their highly adjustable and predictable properties, three-dimensional(3D) printed geometrically ordered porous biomaterials offer unique opportunities as orthopedic implants. The performance of such biomaterials is, however, as much a result of the surface properties of the struts as it is of the 3D porous structure. In our previous study, we have investigated the in vitro performances of selective laser melted (SLM) Ti-6Al-4V scaffolds which are surface modified by the bioactive glass (BG) and mesoporous bioactive glass (MBG), respectively. The results demonstrated that such modification enhanced the attachment, proliferation, and differentiation of human bone marrow stromal cells (hBMSC). Here, we take the next step by assessing the therapeutic potential of 3D printed Ti-6Al-4V scaffolds with BG and MBG surface modifications for bone regeneration in a rabbit bone defect model. METHODS: 3D printed Ti-6Al-4V scaffolds with BG and MBG surface modifications were implanted into the femoral condyle of the rabbits, the Ti-6Al-4V scaffolds without surface modification were used as the control. At week 3, 6, and 9 after the implantation, micro-computed tomography (micro-CT) imaging, fluorescence double-labeling to determine the mineral apposition rate (MAR), and histological analysis of non-decalcified sections were performed. RESULTS: We found significantly higher volumes of regenerated bone, significantly higher values of the relevant bone morphometric parameters, clear signs of bone matrix apposition and maturation, and the evidence of progressed angiogenesis and blood vessel formation in the groups where the bioactive glass was added as a coating, particularly the MGB group. CONCLUSIONS: The MBG coating resulted in enhanced osteoconduction and vascularization in bone defect healing, which was attributed to the release of silicon and calcium ions and the presence of a nano-mesoporous structure on the surface of the MBG specimens.

The fluoride coated AZ31B magnesium alloy improves corrosion resistance and stimulates bone formation in rabbit model.

This study aimed to evaluate the effect of fluorine coated Mg alloy and clarify its mechanism in bone formation. We implanted the fluorine coated AZ31B Mg alloy screw (group F) in rabbit mandibular and femur in vivo. Untreated AZ31B Mg alloy screw (group A) and titanium screw (group T) were used as control. Then, scanning electron microscopy, the spectral energy distribution analysis, hard and decalcified bone tissues staining were performed. Immunohistochemistry was employed to examine the protein expressions of bone morphogenetic protein 2 (BMP-2) and collagen type I in the vicinity of the implant. Compared with the group A, the degradation of the alloy was reduced, the rates of Mg corrosion and Mg ion release were slowed down, and the depositions of calcium and phosphate increased in the group F in the early stage of implantation. Histological results showed that fluorine coated Mg alloy had well osteogenic activity and biocompatibility. Moreover, fluoride coating obviously up-regulated the expressions of collagen type I and BMP-2. This study confirmed that the fluorine coating might improve the corrosion resistance of AZ31B Mg alloy and promote bone formation by up-regulated the expressions of collagen type I and BMP-2.

Loss of mechanical properties in vivo and bone-implant interface strength of AZ31B magnesium alloy screws with Si-containing coating.

In this study the loss of mechanical properties and the interface strength of coated AZ31B magnesium alloy (a magnesium-aluminum alloy) screws with surrounding host tissues were investigated and compared with non-coated AZ31B, degradable polymer and biostable titanium alloy screws in a rabbit animal model after 1, 4, 12 and 21weeks of implantation. The interface strength was evaluated in terms of the extraction torque required to back out the screws. The loss of mechanical properties over time was indicated by one-point bending load loss of the screws after these were extracted at different times. AZ31B samples with a silicon-containing coating had a decreased degradation rate and improved biological properties. The extraction torque of Ti6Al4V, poly-l-lactide (PLLA) and coated AZ31B increased significantly from 1week to 4weeks post-implantation, indicating a rapid osteosynthesis process over 3weeks. The extraction torque of coated AZ31B increased with implantation time, and was higher than that of PLLA after 4weeks of implantation, equalling that of Ti6Al4V at 12weeks and was higher at 21weeks. The bending loads of non-coated AZ31B and PLLA screws degraded sharply after implantation, and that of coated AZ31B degraded more slowly. The biodegradation mechanism, the coating to control the degradation rate and the bioactivity of magnesium alloys influencing the mechanical properties loss over time and bone-implant interface strength are discussed in this study and it is concluded that a suitable degradation rate will result in an improvement in the mechanical performance of magnesium alloys, making them more suitable for clinical application.

In vivo degradation and tissue compatibility of ZK60 magnesium alloy with micro-arc oxidation coating in a transcortical model.

Magnesium alloys were studied extensively as a class of biodegradable metallic materials for medical applications. In the present study, ZK60 magnesium alloy was considered as a candidate and the micro-arc oxidation (MAO) treatment was adopted in order to reduce the degradation rate of the alloy. The in vivo degradation behaviors and biological compatibilities of ZK60 alloys with and without MAO treatment were studied with a transcortical model in rabbits. The implant and the surrounding bone tissues were characterized by CT, SEM and histological methods at 2, 4 and 12 weeks after the implantation. The results demonstrated that both the bare and MAO-coated ZK60 alloys completely degraded within 12 weeks in this animal model. The MAO coating decreased the degradation rate of ZK60 alloy and enhanced the response of the surrounding tissues within the first 2 weeks. After then, an acceleration of the degradation of the MAO-coated ZK60 alloy was observed. It was found that the alloy could be degraded before the complete degradation of the MAO coating, leading to the local peeling off of the coating. An in vivo degradation mechanism of the MAO-coated ZK60 alloy was proposed based on the experimental results. The severe localized degradation caused by the peeling off of the MAO coating was the main reason for the acceleration of the degradation of the MAO-coated ZK60 alloy.

[A study on the influence of fluid shear stress on the expression of COX-2 in HPDLF].

PURPOSE: To utilize the fluid shear stress to simulate occlusion trauma in vitro. Different density of FN was given to HPDLF and the amount of COX-2 mRNA of HPDLF was measured at different time. METHODS: Young healthy permanent teeth which were extracted for orthodontic treatment were collected. The HPDLF was primarily cultured via the method for tissue block. The third generation cells were taken as the study objects. There were two groups in this study. Group A were cultured with the fluid shear stress given by a special swing bed(35r/min). Group B were cultured quietly. Different density of FN (0,20,40,60microg/ml)was added into Group A and B. The cells were cultured for 6 and 12 hours without serum. RESULTS: After 24 hours, 80% tissue block anchoraged. After 7-10 days,there were cells moving from the tissue block,After about 14 days, the cell overgrew the culture flask.Immuocytochemistry showed that anti- filamin was positive while anti- ceratin was negative. The expression of COX-2 mRNA was positive after 6 hours in group A,and the amount of the expression decreased with the increase of the amount of FN . The expression of COX-2 mRNA after 12 hours in group A was higher than that after 6 hours. The result of electrophoresis for group B was negative. The electrophoresis optical density of beta-actin amplification fragment was uniform. CONCLUSIONS: The fluid shear stress can revoke the inflammatory reaction of HPDLF. Human plasma FN can decreased the expression of COX-2 mRNA of HPDLF,which is dose and time dependent.