Effect of eNOS gene G894T mutation on the ability of vascular endothelial cells to produce nitric oxide and its significance / 中国医师杂志

Objective To investigate the relationship between vascular endothelial cell nitric oxide synthase (eNOS) gene polymorphism and the pathogenesis of avascular femoral head necrosis (ANFH).Methods The eNOS full-length CDS fragments,containing 894G or 894T separately,was subcloned into lentiviral expression vector,and then infect the human umbilical vein endothelial cells (HUVEC).The contents of nitric oxide NO and cyclic guanosine monophosphate (cGMP) in cell culture supernatant were detected in a time-dependent manner.The luciferase-labeled reporter plasmid pNF-κB-luc was co-transfected with the reference control plasmid pRL-TK into HUVEC cells infected with LV-eNOS-894G,LV-eNOS-894T,and control lentivirus (LV-NC) for 48 h.The luciferase activity of each group was detected.The expression of nuclear factor (NF)-κB protein and eNOS protein in HUVEC cells were detected by Western blot assay.The HUVEC cells in each group were co-cultured with hFOB1.19 cells,and the concentration of alkaline phosphatase (ALP) or osteocalcin (OCN) in the supernatant were detected at different time points.Results The contents of NO and cGMP in the cell culture supernatant of the full-length lentivirus expressing eNOS gene (containing 894G or 894T) were significantly higher than that in the empty cell group and the empty vector group,and the contents of NO and cGMP in the cell culture supernatant of the 894G group were significantly higher than that of 894T group (P ＜ 0.01).Compared with blank cells,the expression levels of NF-κB/p65 protein and eNOS protein were significantly increased in cells expressing eNOS,and the expression of NF-κB/p65 protein in 894G group was significantly increased than 894T group,but there was no difference in eNOS protein expression between the 894G and 894T groups;each group of HUVEC cells were co-cultured with hFOB1.19 osteoblasts,and at each of the same time points,the concentrations of ALP and OCN in the cell culture supernatant expressing lentivirus were significantly higher than that in the empty cell group and the empty vector group,and the ALP and OCN concentrations in the cell culture supernatant of the 894G group were significantly higher than those in the 894T group.Conclusions The eNOS gene exon G894T mutation reduces the levels of nitric oxide and cGMP produced by vascular endothelial cells through the eNOS-NO pathway,affecting the expression levels of NF-κB/p65 protein and eNOS protein,and reducing osteoblast activity,and blood supply to blood vessels.It may be one of the pathogenic mechanisms leading to femoral head necrosis.

Retraction: Differential expression of exosomal miRNAs in osteoblasts in osteoarthritis Journal of Central South University. Medical Science, 2018, 43(12):1294-1300. DOI: 10.11817/j.issn.1672-7347.2018.12.003 / 中南大学学报(医学版)

The article entitled "Differential expression of exosomal miRNAs in osteoblasts in osteoarthritis" published on Journal of Central South University (Medical Science), in Volume 43, Issue 12, 2018 (DOI: 10.11817/j.issn.1672-7347.2018.12.003) may have an unclear risk of bias due to insufficient understanding for some results. Further experimental studies are needed. We all agree to retract this article, and apologize to the Journal and readers for the possible negative impact.

Differential expression of exosomal miRNAs in osteoblasts in osteoarthritis / 中南大学学报(医学版)

To analyze the differentially expressed exosomal miRNAs in subchondral osteoblasts in patients with osteoarthritis (OA) and to investigate the key miRNAs potentially involved in the occurrence and progression of OA.
 Methods: Subchondral bones were harvested from 6 patients with OA. All subjects were divided into two groups which was based on the severity of joint wear: An OA group, severely worn side of subchondral bone, and a control group, less worn side of subchondral bone. The exosomes were extracted from osteoblast cells and their characteristics were identified. Then exosomal miRNAs were extracted and sequencing analysis was conducted to compare the expression in the two groups. The most differentially expressed ones (log2Ratio≥2) were subject to miRNA target prediction and quantitative reverse transcription PCR (RT-qPCR) to further quantify the difference.
 Results: Osteoblast extractions were confirmed to be exosomes, which were small double-membranous vesicles with 30-200 nm in diameter and 50-150 nm in peak value of particle size under the scanning microscope. High-throughput sequencing revealed 124 miRNAs whose expression significantly increased in the OA group. The most differentially expressed one with maximum fold change was hsa-miR-4717-5p and its target gene was RGS2. RT-qPCR demonstrated hsa-miR-4717-5p expression in the OA group was relatively higher than that in the control group (2.243 vs 0.480, P<0.01).
 Conclusion: There is distinct difference in expression profiles of exosomal miRNAs in subchondral osteoblasts between patients with OA and normal subjects. Up-regulated expression of miRANs might participate in OA occurrance and progression.

Effect of alendronate amount on the static mechanical properties of bone cement / 中南大学学报(医学版)

OBJECTIVE@#To determine the effect of alendronate amount on the static mechanical properties of bone cement before elution and 4 weeks after elution, and determine suitable amount of alendronate in cement from the perspective of biomechanics.@*METHODS@#Samples of compression strength and bending strength (modulus) were prepared with bone cement adding 0, 10, 50, 100, 500 and 1000 mg alendronate in 50 g Cemex®XL bone cement powders respectively (named as G0-G5 groups). The compression strength, bending strength, and bending modulus of bone cement were examinated by INSTRON 8032 tester before the elution and 4 weeks after the elution. Some broken samples of 4-point bending test were examinated by scanning electronic microscopy (SEM), and some other samples of bending strength (modulus) tests by micro-CT.@*RESULTS@#No significant difference was found in the compression strength before the elution,4 weeks after elution in the 6 groups, and before and after the elution in the respective groups (P>0.05). Compared with G0 group, G1-G4 groups had no obvious difference in the bending strength before the elution and 4 weeks after the elution (P>0.05), while G5 group had difference (P0.05). Taken G0 group as the control group, G1-G4 groups had no visible effects on the bending modulus 4 weeks after the elution (P>0.05), while G5 group decreased it significantly (P0.05), while bending modulus before and after the elution in the respective groups displayed a marked difference (P<0.05).@*CONCLUSION@#Less than 500 mg alendronate added in Cemex?XL 50 g bone cement powder does not decrease the compressive strength, flexural strength and flexural modulus before the elution and 4 weeks after the elution.

Effect of alendronate-loaded bone cement on osteoblast / 中南大学学报(医学版)

OBJECTIVE@#To determine the effect of bone cement, with different amounts of alendronate on osteoblast, and determine the cytotoxicity of alendronate-integrated bone cement from the viewpoint of cell biology.@*METHODS@#According to different additions (0, 10, 50, 100, 500, 1 000 mg) of alendronate in 50 g Cemex®XL bone cement powder, the experiments were divided into 6 groups, namely G0-G5 groups. In all groups, the adhesive capacity of osteoblast-like cells MG-63 was evaluated by electron microscope, the optical density (OD) value of cells by MTT colorimetry method, the alkaline phosphatase activity (AKP) by AKP assay kit, the apoptosis rates by Annexin-V-FITC apoptosis detection kit, and the bone mineralization potentiality by phase contrast microscope.@*RESULTS@#The adhesive capacity of MG-63 was good in all groups. Compared with the G0 group, the cell apoptosis was inhibited in G1-G4 groups while in G5 group the cell apoptosis was promoted and cell proliferation was inhibited (P0.05).@*CONCLUSION@#Less than 500 mg alendronate added in Cemex®XL 50 g bone cement powder has no cytotoxicity on osteoblasts.

Measurement of proximal femoral morphology and analysis of 500 cases in Hunan Province / 中南大学学报(医学版)

Objective:To evaluate the effect of age and gender on the femoral morphology to guide prosthesis selection in operation and design. Methods:A total of 500 females and males were collected from the departments of orthopedics and medical radiology, Second Xiangya Hospital, Central South University. Informed consent was obtained from all subjects. All patients underwent anteroposterior position scan of the left or right hip joint using Philips Digital Diagnost DR system. The shooting range included the hip joint and at least 2/3 of the proximal femur. The images were measured with Onis 2.3 software. We measured 13 parametersfromthepatients,includingtheexternalparamtersofthefemur,radius-lengthparameters of femoral medullary cavity, and morphological parameters of the femoral medullary cavity. Results:Compared with Westeners, the offset was smaller, while the neck shaft angle was signiifcantly larger in Chinese population (P0.05). The main differences between 31 and 70 years old were the diameter of femoral head, the offset of isthmus, the medullary cavity diameter and extracortical width at isthmus level and the medullary cavity diameter at the level of the lesser trochanter (P<0.05). The modullary transverse diameter at 20 mm below the lesser trochanter and isthmus and extracortical width of isthmus in the male and female group was positively correlated with age (P<0.01), while the parameters of the proximal femoral canal morphology in the female group were negatively correlated with age. The female canal parameters had a stronger correlation. Conclusion:Chinese proximal femoral parameters are signiifcantly different from Westerners. When people, especially females, get older, the medullary cavity diameter of the isthmus and proximal femur becomes wider and the morphology of the femur becomes straight. The difference in the femoral morphology between the male and female decline with the age. There is almost no difference for the over 70 years old. For the 31-70 years old, The male femoral cavity diameter is larger and the position of isthmus is lower than in the females.

Perioperative Management of Fracture with Hemophilia A / 中国医师杂志

Objective To study the perioperative treatment of the patients having fracture with hemophilia A. Methods Seven male hemophilia A patients with fractures were treated in our hospital from Jan. 1988 to Apr. 2003. All the patients received coagulation factor replacement treatment. The plasma level of factor Ⅷ was measured at preoperation, intraoperation and postoperation, fracture was treated by internal fixation, and the mean follow-up period was 16 months. Results In this series there were 1 severe, 2 moderate, 2 mild, and 2 pre-clinic hemophilia A patients, all patients had bone union during the follow-up, and no complications such as hematoma, infection and loosening or breaking of internal fixation happened. Conclusion The surgical treatment of fractures with hemophilia A can obtain excellent clinical outcome throug careful perioperative preparation.