Alendronate Alleviated Femoral Head Necrosis and Upregulated BMP2/EIF2AK3/EIF2A/ATF4 Pathway in Liquid Nitrogen Treated Rats.

BACKGROUND: Osteonecrosis of the femoral head (ONFH) seriously affects the quality of life and labor ability of patients. It is urgent and vital to find the methods for necrosis clinical treatment. OBJECTIVE: This study aims to study the potential protective effects of Alendronate in the early stage of femur head necrosis. METHODS: Ten clinal ONFH tissue samples were employed. H&E staining was employed for the observation of the pathological characteristics of ONFH. The rat model (n=12) was established by the treatment of liquid nitrogen and then treated with Alendronate. The protein expression of BMP2, EIF2AK3, EIF2A and ATF4 were detected via Western blotting and IHC. RESULTS: Fibrin and necrotizing granulation tissue were observed in ONFH tissues with lymphocytes and plasma cells infiltrating in the necrotic area, exhibiting the inflammatory muscle with abnormal shape and color. In the Model group, the BMP2 and ATF4 were mainly distributed in the cell boundaries. The relative protein expression of BMP2, EIF2AK3, EIF2A, ATF4 was decreased in the Model group, compared to the NC group, which was partially recovered by the Alendronate application. CONCLUSION: Alendronate application partially reversed the suppression of expression of BMP2, EIF2AK3, EIF2A, ATF4 caused by liquid nitrogen. Alendronate could be a promising strategy of curing ONFH via targeting BMP2/EIF2AK3/EIF2A/ATF4 pathway.

Digital Technology Combined with 3D Printing to Evaluate the Matching Performance of AO Clavicular Hook Plates.

BACKGROUND: AO clavicular hook plates have been widely used for treating acromioclavicular (AC) joint dislocation and distal clavicle fractures. Many complications have been reported, and many patients have complained about the discomfort of the plate. However, no study on the impact of clavicular hook plates in AC joint 3D printing models has been reported. OBJECTIVE: To evaluate the matching performance of hook plates with different hook depths when they were implanted in 3D printing models of normal Chinese AC joints and to propose a further design to achieve a better match. METHODS: AO clavicular hook plates with two different hook depths of 15 mm and 18 mm were implanted in 3D printing models of forty Chinese normal AC joints. The angle between the distal clavicle and plate (CPA) and the drop between acromion and distal clavicle (ACD) of normal AC joints with and without plates were measured. RESULTS: Mismatch was found when the hook plates were implanted, with an average CPA of 18.8 ± 5.1° with the 15-mm hook plate and 10.2 ± 4.9° with the 18-mm hook plate. To eliminate the CPA, the ACD decreased by 6.3 ± 1.1 mm with the 15-mm hook plate and 2.9 ± 0.9 mm with the 18-mm hook plate. CONCLUSIONS: The results revealed that AO clavicular hook plates with different hook depths were very likely to result in over-reduction of the AC joint. It is necessary to bend the hook angle to fit the patient's AC joint to achieve accurate reduction.

Articular Cartilage Stem Cells Influence the Postoperative Repair of Hip Replacement by Regulating Endoplasmic Reticulum Stress in Chondrocytes via PERK Pathway.

OBJECTIVE: Endoplasmic reticulum stress (ERS) is present in chondrocytes of osteoarthritis, and the intensity of ERS is related to the degree of cartilage degeneration. In vitro intervention strategies can change the status of ERS and induce the inhibition of ERS-related pathway. Therefore, this study is designed to explore the role and molecular mechanism of cartilage stem cells (ACSCs) of ERS in chondrocytes after hip replacement. METHODS: Human cartilage cell lines C28/I2 were cultured as the control group. The ERS inducer was added into C28/I2 as ERS group. The third ERS + stem cells group was formed by adding cartilage stem cells into ERS group, and further transfection of si-PERK was defined as si-PERK + ERS + stem cells group. Cell cycle and apoptosis in the four groups were determined by flow cytometry. The protein expression of GRP78, PERK, ATF4, TMEM119, CDK4, Cyclin D, and BMP6 in chondrocytes in the four groups were investigated by western blot, and the distribution of PERK, TMEM119, and BMP6 in chondrocytes were observed by immunofluorescence assay. In addition, the transcriptional levels of Bcl2, Bax, and Caspase 3 were also determined by RT-PCR. RESULTS: In cell cycle assay, ERS increased the accumulation of cells in G0 /G1 and G2 /M, while cartilage stem cells weakened the effects. The apoptosis rates in control group, ERS, ERS + stem cells, si-PERK + ERS + stem cells were 0%, 21.3%, 18.9%, and 15.9%, respectively, and the difference of apoptosis rate between the latter three groups and control group was statistically significant (P < 0.01). Stem cells could weaken the ERS-induced cell apoptosis, especially reducing the number of cells in the late stage of apoptosis from 5.4% to 1.1%. The protein level of GRP78, PERK, ATF4, TMEM119, and BMP6 in the group of ERS, ERS + stem cells, and si-PERK + ERS + stem cells were all significantly higher than those in control group, and the group of ERS + stem cells was the highest, all of the differences were significant (P < 0.01). However, the protein level of CDK4 and Cyclin D presented an absolutely opposite trend and the difference was still significant (P < 0.05). The group of si-PERK + ERS + stem cell was lower than those in the group of ERS + stem cell but higher than those in the group of ERS (P < 0.05). The level of Caspase 3 in the latter three groups was significantly higher than those in the control group, and the group of ERS was the highest (P < 0.01). Besides, the relative level of Bcl-2/Bax in control group was 1, but the group of ERS was about 0.5, and there was significant difference (P < 0.01). The ratio of Bcl-2/Bax in the group of ERS + stem cells was more than 2 and significantly higher than those of other groups. CONCLUSION: ACSCs could reduce ERS-induced chondrocyte apoptosis by PERK and Bax/Bcl-2 signaling pathway.

Lysosomal autophagy promotes recovery in rats with acute knee injury through TFEB mediation.

BACKGROUND: To study the role of lysosomal decomposition and elimination of old bone matrix, as well as the mechanism of promoting chondrocyte growth and bone recovery through the perspective of TFEB-mediated lysosomal autophagy. METHODS: Rat models of acute knee injury were designed, and autophagy flow was detected by injection of autophagy inhibitors 3-methyladenine. Autophagy flow was detected by RFP-GFP-LC3 double fluorescence molecule. The expression of TFEB, DRAM, MAPLC3, and MITF were analyzed by Western blot, and the expression of genes NITF, Bcl2, and TYR in rat cartilage tissues were detected by RT-PCR. RESULTS: The number of autophagosomes was increasing in the auto group compared with the inhibitor-auto group and normal group. There was a significant difference of LC3 levels in the auto group and inhibitor-auto group compared with the normal control. The expression of TFEB, DRAM, MAPLC3, and MITF proteins by Western blot analysis were significantly increased in the auto group and decreased in the inhibitor-auto group. The expression of NITF, Bcl2, and TYR by RT-PCR determination were higher in the auto group and inhibitor-auto group than the normal group. CONCLUSIONS: Autophagy can inhibit apoptosis, promote chondrocyte growth and bone regeneration, and restore knee joint injury of rats. The main mechanism is to promote the effect of TFEB-mediated lysosomal autophagy.

Gotfried positive reduction promotes the repair of femoral neck fracture potentially via enhancing osteogenesis and angiogenesis.

The aim of this study was to investigate the effect of Gotfried positive reduction (GPR) on repair of femoral neck fracture in rabbits and its underlying mechanisms. Male New Zealand white rabbits were employed to establish the model of femoral neck fracture. All the rabbits were randomly divided into four groups: control, open accurate reduction (OR), closed Gotfried negative support reduction (CR-N) and closed Gotfried positive support reduction (CR-P). At the 8th and 12th week after surgery, the anteroposterior and lateral radiographs of their hip joints were taken by X-ray, and local hemodynamics of their hip joints was detected by ultrasound. Histological examination was evaluated by HE staining and bone biological strength test was measured by testing machine, which was performed to study the repair of femoral neck fracture. Osteogenesis and angiogenesis-related proteins were measured by western blot in bone tissues and synovial tissues. The results revealed that the fracture healing intensity and blood supply in CR-P were better than those in CR-N and much more excellent than those in OR. In addition, the content of bone morphogenetic protein2 (BMP2), platelet derived growth factor (PDGF) and ocsteocalcin was higher in CR-P group than in CR-N, while lower in CR-P than OR. Furthermore, the expression of BMP2, COL-2 and angiopoietin (ANGPT) was upregulated in CR-P compared to CR-N and OR. Taken together, our results indicated that GPR was able to promote the repair of femoral neck fracture via enhancing osteogenesis and angiogenesis, which is valuable to us and shows good application prospect in bone tissue repair.

3D Printed Model Used as Preoperative Tool for Treating Acromioclavicular Joint Dislocation with Pre-Contoured Clavicle Hook Plate: Technical Note.

OBJECTIVE: To introduce a new technique using real-size 3D-printed acromioclavicular joint models as preoperative tools for treating acromioclavicular (AC) joint dislocation. METHODS: Both the injured AC joint and the mirrored normal side AC joint are 3D printed into real-size models. A proper hook plate is selected and pre-bended to fit the normal side AC joint. The bended plate is then used to perform a reduction operation on a real-size injured AC joint model and the result is compared with the mirrored normal AC joint model to see if the reduction is accurate. RESULTS: It is easy to select and bend a clavicle hook plate to achieve an accurate reduction with the help of a 3D printing technique. CONCLUSION: With this technique, surgeons can achieve an accurate reduction of the injured AC joint with a clavicle hook plate which might be helpful to reduce the risk of complications.

Preliminary exploration of a quantitative assessment index for the matching performance of anatomical bone plates using computer.

BACKGROUND: To explore a new quantitative index to assess the matching performance of anatomical bone plates using digital technology. METHODS: CT data of normal tibias of 40 adults were collected. Two brands of medial distal tibia plates were digitized. Two trained orthopedists simulated the surgical operation in Rhino 5.1 software by placing the plate curve on the medial distal tibia surface. The volume of the interstice between the plate curve and the bone surface was measured. The inverse value of this average interstice distance was used as the matching performance index (MPI). A wall thickness analysis tool was used to mark various interstice distances with varied colors. RESULTS: The Kangli medial distal tibia plate had a MPI of 0.55 ± 0.08 by operator A and 0.55 ± 0.06 by operator B. The general care medial distal tibia plate had a MPI of 0.32 ± 0.06 by operator A and 0.31 ± 0.05 by operator B. There were significant variations in the MPI between the two types of plates by both operators (p < 0.001). And significant variations were observed in the MPI of general care medial distal tibia plates among various operator groups (p = 0.028). CONCLUSION: This quantitative index of matching performance is straightforward and intuitive. However, we still need a method to improve the experimental repeatability, especially when it comes to a plate with poor matching performance.

Increasing Level of Leisure Physical Activity Could Reduce the Risk of Hip Fracture in Older Women: A Dose-Response Meta-analysis of Prospective Cohort Studies.

We carried out the study to investigate and quantitatively assess the potential association between current level of physical activity and the risk of osteoporosis hip fracture in older women. Relevant publications before October 2015 were identified using the PubMed and Ovid searching tools. A dose-response meta-analysis was carried out to combine and analysis results. Fourteen prospective studies were included in the meta-analysis. A general analysis of 9 studies showed a significant inverse relationship between increasing level of physical activity and risk of hip fracture in older women [relative risk (RR) = 0.93, 95% confidence interval (95% CI): 0.91-0.96]. The result of a sensitivity analysis was consistent with the general analysis (RR = 0.94, 95% CI: 0.93-0.96). The association between increasing level of physical activity and risk of wrist fracture was not statistically significant in a general analysis of three studies (RR = 1.004, 95% CI: 0.98-1.03). A potential direct association between increasing level of physical activity and risk of wrist fracture was observed after removing 1 study with the greatest weight (RR = 1.01, 95% CI: 1.00-1.03). No significant publication bias was observed in our analysis. Our results show that increasing level of physical activity within an appropriate range may reduce the risk of hip fracture but not the risk of wrist fracture in older women.

Decreased Zn(2+) Influx Underlies the Protective Role of Hypoxia in Rat Nucleus Pulposus Cells.

Zn(2+) is an essential component of metalloproteinases, and is required for their activity in cartilage; however, the effect of Zn(2+) on nucleus pulposus (NP) cells has not been widely investigated. The aim of this paper was to investigate the effect of intracellular Zn(2+) concentration ([Zn(2+)]i) in hypoxia-induced regulation of metalloproteinases (MMPs) and extracellular matrix (ECM) production in NP cells. NP cells from Sprague-Dawley (SD) rats were cultured as monolayers or in alginate beads. [Zn(2+)]i was assayed by FluoZin-3 AM staining. Alcian Blue staining, immunochemistry, 1,9-dimethylmethylene blue (DMMB) assay, and real-time PCR were used to assay collagen II, proteoglycan, and COL2A1, MMP-13, and ADAMTS-5 mRNA expression. ZIP8, a main Zn(2+) transporter in chondrocytes, was assayed by immunochemistry and in Western blotting. Interleukin (IL)-1ß- and ZnCl2-induced increases of [Zn(2+)]i were significantly inhibited by hypoxia. Hypoxia did not reverse a decline of ECM expression caused by IL-1ß and ZnCl2 in monolayer cultures, but did significantly attenuate the decreases of proteoglycan, glycosaminoglycan (GAG), and COL2A1 mRNA expression following IL-1ß and ZnCl2 treatment in alginate bead cultures. However, ZnCl2 inhibited the protective effect of hypoxia. Both an intracellular Zn(2+) chelator and hypoxia prevented the increase in MMP-13 mRNA expression. IL-1ß and ZnCl2 treatment increased ZIP8 expression in NP cells, and hypoxia inhibited ZIP8 expression. In conclusion, decrease of Zn(2+) influx mediates the protective role of hypoxia on ECM and MMP-13 expression. Consequently, changes in intracellular Zn(2+) concentration maybe involved in intervertebral disc degeneration.

[Study on animal model of perforator sural neurocutaneous flap].

OBJECTIVE: To establish the experimental animal model of perforator sural neurocutaneous flap for laying a foundation of further study on its physiology and haemodynamics. METHODS: Thirty-five New Zealand rabbits were divided into four groups, weighing 2.5-3.0 kg and being male or female. In group A (n=5), vivisection was performed to observe the starting point and arrangement of sural nerve, its concomitant vessels, posterior tibial artery and perforating vessel. In groups B and C (n=5), red latex and gelatin-lead oxide were injected into the concomitant arteries of sural nerve and the posterior tibial arteries respectively to observe their arrangement, the diameter and anastomosis. In group D, forty neurocutaneous flaps based on single perforator were elevated in the twenty rabbits with a size of 7 cm x 1 cm and a pedicle of 0.5 cm. The colour and condition of flaps were observed. RESULTS: The sural nerve originated from posterior tibial nerve, passed through the lateral head of the gastrocnemius at site of the popliteal fossa, descended obliquely to exterior, entered in the deep fascia at about (5.42 +/- 0.15) cm above lateral malleolus, and descended vertically to lateral malleolus. Its concomitant artery originated from deep femoral artery with an initial diameter of (0.73 +/- 0.11) mm and extended to the lateral malleolus along the sural nerve. A perforating branch of posterior tibial artery at the position of the calcaneus originated from the midpoint of the line connecting between the medial malleolus and the calcaneus with an initial diameter of (0.45 +/- 0.01) mm. The perforating branch traversed the calcaneus to the region of the lateral malleolus, and anastomosed to the concomitant artery of the sural nerve, forming a vascular plexus around the sural nerve. In group D, two cases were excluded due to infection. The survival rate was 78.0% +/- 1.5% in other 38 flaps 10 days after operation. CONCLUSION: The perforator based sural neurocutaneous flap in rabbit is a good experimental model, which has stable anamatic features and reliable blood distribution.

[Analysis of the donor-site complications of the anterolateral thigh flap].

OBJECTIVE: To investigate the donor-site complications of the anterolateral thigh flap and its influencing factors. METHODS: Between July 1988 and July 2007, 427 patients were treated with anterolateral thigh flap. Among them, 33 patients had postoperative donor-site complications and their clinical data were analyzed retrospectively. There were 21 males and 12 females aged 14-47 years old (average 32.7 years old). The size of the wound defect ranged from 16 cm x 7 cm to 28 cm x 13 cm. The area of the flap harvested during operation ranged from 16 cm x 7 cm to 30 cm x 13 cm. The donor sites were treated by direct suture in 7 cases; free split-thickness skin graft in 23 cases, and reverse superficial epigastric artery flap repair in 3 cases. The size of graft in the donor site ranged from 10 cm x 5 cm to 18 cm x 8 cm. The occurrence of short-term (within 4 weeks) and long-term (over 6 months) complications were analyzed. RESULTS: All patients were followed up for 8-54 months (average 21 months). There were 26 patients (78.8%) with short-term and long-term complications and 7 patients (21.2%) with long-term complications. The short-term complications included skin graft necrosis in 23 cases (69.7%), wound infection in 17 cases (51.5%), and muscle necrosis in 2 cases (6.1%). The long-term complications included non-healing wounds in 21 cases (63.6%), serious scar in 28 cases (84.8%), discomfort in 25 cases (75.8%), and dysfunction of the quadriceps femoris in 16 cases (48.5%). CONCLUSION: The occurrence of anterolateral thigh flap donor-site complications is related to the anatomical structure of the anterolateral thigh region, the surgical procedure, and the patient's physique.

[Clinical application of the reverse neurocutaneous flap with anterior cutaneous branch of the femoral nerve supplied by the perforator of saphenous artery].

OBJECTIVE: To report the operative technique and clinical application of the neurocutaneous flap with anterior cutaneous branch of the femoral nerve supplied by the perforator of saphenous artery. METHODS: The reverse neurocutaneous flap with anterior cutaneous branch of the femoral nerve supplied by the perforator of saphenous artery was used for repairing the defect around knee or at the upper pad of leg. Since Oct. 2005, 16 cases were treated. The flap size ranged from 15 cm x 7 cm to 30 cm x 15 cm. Flap rotation angle ranged from l00 degrees to 180 degrees. RESULTS: 13 flaps survived completely. Flap necrosis happened at the 1/7 - 1/5 distal end of the 3 flaps, which healed with dressing or local flap advancement. The patients were followed up for 6 to 24 months with satisfactory functional and cosmetic results. There was no morbidity at the donor site. CONCLUSION: The flap has the advantages of both the perforator flap and the neurocutaneous flap. The size of the neurocutaneous flap with the anterior cutaneous branch of the femoral nerve can be enlarged for the large defect at lower extremity.

[Study on detection of samples of Rh-weak D and Del].

To study the detection of weak D and Del from samples initially screened RhD(-), RhD phenotype was initially screened by routine serological test, out of which weak D phenotype was detected by indirect antiglobulin test (IAT) and Del phenotype was detected by chloroform-trichloroethylene absorption-elution test. The results showed that 56 samples were RhD(-) confirmed by routine serology test, which were screened out of 26 200 donors, among them 5 samples were typed as weak D by IAT and 9 cases samples were typed as Del by absorption-elution test. In conclusion, the samples which typed as RhD(-) by routine serological test must be identified by IAT and chloroform-trchloroethylene absorption test is order to detect weak D and Del phenotype. It is important for clinical transfusion safely.