Valproic acid attenuates traumatic spinal cord injury-induced inflammation via STAT1 and NF-κB pathway dependent of HDAC3.

BACKGROUND: Microglial polarization with M1/M2 phenotype shifts and the subsequent neuroinflammatory responses are vital contributing factors for spinal cord injury (SCI)-induced secondary injury. Nuclear factor-κB (NF-κB) is considered the central transcription factor of inflammatory mediators, which plays a crucial role in microglial activation. Lysine acetylation of STAT1 seems necessary for NF-kB pathway activity, as it is regulated by histone deacetylases (HDACs). There have been no studies that have explained if HDAC inhibition by valproic acid (VPA) affects the NF-κB pathway via acetylation of STAT1 dependent of HDAC activity in the microglia-mediated central inflammation following SCI. We investigated the potential molecular mechanisms that focus on the phenotypic transition of microglia and the STAT1-mediated NF-κB acetylation after a VPA treatment. METHODS: The Basso-Beattie-Bresnahan locomotion scale, the inclined plane test, the blood-spinal cord barrier, and Nissl staining were employed to determine the neuroprotective effects of VPA treatment after SCI. Assessment of microglia polarization and pro-inflammatory markers, such as tumor necrosis factor (TNF)-α, interleukin (IL)-1ß, IL-6, and interferon (INF)-γ was used to evaluate the neuroinflammatory responses and the anti-inflammatory effects of VPA treatment. Immunofluorescent staining and Western blot analysis were used to detect HDAC3 nuclear translocation, activity, and NF-κB signaling pathway activation to evaluate the effects of VPA treatment. The impact of STAT1 acetylation on NF-kB pathway and the interaction between STAT1 and NF-kB were assessed to evaluate anti-inflammation effects of VPA treatment and also whether these effects were dependent on a STAT1/NF-κB pathway to gain further insight into the mechanisms underlying the development of the neuroinflammatory response after SCI. RESULTS: The results showed that the VPA treatment promoted the phenotypic shift of microglia from M1 to M2 phenotype and inhibited microglial activation, thus reducing the SCI-induced inflammatory factors. The VPA treatment upregulation of the acetylation of STAT1/NF-κB pathway was likely caused by the HDAC3 translocation to the nucleus and activity. These results indicated that the treatment with the VPA suppressed the expression and the activity of HDAC3 and enhanced STAT1, as well as NF-κB p65 acetylation following a SCI. The acetylation status of NF-kB p65 and the complex with NF-κB p65 and STAT1 inhibited the NF-kB p65 transcriptional activity and attenuated the microglia-mediated central inflammatory response following SCI. CONCLUSIONS: These results suggested that the VPA treatment attenuated the inflammatory response by modulating microglia polarization through STAT1-mediated acetylation of the NF-κB pathway, dependent of HDAC3 activity. These effects led to neuroprotective effects following SCI.

Oral dehydroepiandrosterone supplementation enhances osteoporotic fracture healing in the OVX rats.

Osteoporosis easily causes delayed fracture union, even non-union. It has been demonstrated that dehydroepiandrosterone (DHEA) supplementation can increase estrogen levels and improve bone mineral density (BMD) in the elderly, while the role of DHEA on fracture healing remains unknown. This study aimed to elucidate the impact of DHEA supplementation on osteoporotic fracture healing. Seventy-two female Sprague-Dawley rats were used. Forty-eight rats received ovariectomy (OVX), and the remaining rats received a sham OVX operation (sham group). A right transverse femoral osteotomy was performed in all rats at 12 weeks post-OVX. OVX rats were randomly allocated into 2 groups (n = 24 in each group): (i) ovariectomized rats (control group) and (ii) ovariectomized rats treated with DHEA (DHEA group, 5 mg/kg/day). The DHEA supplementation was initiated on the first day post-fracture for 3, 6, and 12 weeks. Fracture healing was evaluated by radiography, histology, biomechanical analysis, and dual-energy X-ray absorptiometry (DEXA). Serum biomarkers were analyzed using enzyme-linked immunosorbent assay (ELISA). At 3 and 6 weeks, radiographs revealed reduced calluses formation and lower radiographic scores in the control group than in other groups. The sham and DHEA groups showed higher BMD and bone mineral content (BMC) at the fracture site than the control group after fracture. Histological analysis revealed the fracture callus was remodeled better in the sham and DHEA groups than in the control group. At the early phase of healing, DHEA supplementation increased osteoblast number, callus area, and cartilage area than the control group. An increased bone area was observed in the DHEA group than in the control group at the late phase of healing. Additionally, improved biomechanical characteristics were observed in both the sham and DHEA groups than those in the control group post-fracture. ELISA showed higher levels of insulin-like growth factor-1 (IGF-1) and 17ß-estradiol (E2) in the DHEA group than in the control group post-fracture. Furthermore, the DHEA group exhibited significantly elevated alkaline phosphatase (ALP) and osteocalcin (OC) levels compared to the control group at 6 and 12 weeks. The DHEA group and the control group did not exhibit a notable difference in TRAP-5b levels. The present study demonstrated that the DHEA treatment has a favorable impact on osteoporotic fracture healing by enhancing callus formation, consolidation, and strength in the OVX rats.

The Mechanism of Deformation Compatibility of TA2/Q345 Laminated Metal in Dynamic Testing with Split-Hopkinson Pressure Bar.

The laminated metal materials are widely used in military, automobile and aerospace industries, but their dynamic response mechanical behavior needs to be further clarified, especially for materials with joint interface paralleling to the loading direction. The mechanical properties of TA2/Q345 (Titanium/Steel) laminated metal of this structure were studied by using the split Hopkinson pressure bar (SHPB). To shed light on the stress-state of a laminated metal with parallel structure, the relative non-uniformity of internal stress R(t) was analyzed. The mechanism of deformation compatibility of welding interface was discussed in detail. The current experiments demonstrate that in the strain rate range of 931-2250 s-1, the discrepancies of the internal stress in specimens are less than 5%, so the stress-state equilibrium hypothesis is satisfied during the effective loading time. Therefore, it is reasonable to believe that all stress-strain responses of the material are valid and reliable. Furthermore, the four deformation stages, i.e., the elastic stage, the plastic modulus compatible deformation stage, uniform plastic deformation stage and non-uniform plastic deformation stage, of the laminated metal with parallel structure were firstly proposed under the modulating action of the welding interface. The deformation stages are helpful for better utilization of laminated materials.

[Mechanism of miR-26a-5p/cAMP response element binding protein 1 molecular axis regulating osteogenic differentiation of adipose-derived mesenchymal stem cells].

Objective: To investigate the regulatory effects of miR-26a-5p on the osteogenic differentiation of adipose-derived mesenchymal stem cells (ADSCs) by regulating cAMP response element binding protein 1 (CREB1). Methods: The adipose tissues of four 3-4 weeks old female C57BL/6 mice were collected and the cells were isolated and cultured by digestion separation method. After morphological observation and identification by flow cytometry, the 3rd-generation cells were subjected to osteogenic differentiation induction. At 0, 3, 7, and 14 days after osteogenic differentiation induction, the calcium deposition was observed by alizarin red staining, ALP activity was detected, miR- 26a-5p and CREB1 mRNA expressions were examined by real-time fluorescence quantitative PCR, and CREB1 protein and its phosphorylation (phospho-CREB1, p-CREB1) level were measured by Western blot. After the binding sites between miR-26a-5p and CREB1 was predicted by the starBase database, HEK-293T cells were used to conduct a dual-luciferase reporter gene experiment to verify the targeting relationship (represented as luciferase activity after 48 hours of culture). Finally, miR-26a-p inhibitor (experimental group) and the corresponding negative control (control group) were transfected into ADSCs. Alizarin red staining, ALP activity, real-time fluorescent quantitative PCR (miR-26a-5p) and Western blot [CREB1, p-CREB1, Runt-related transcription factor 2 (RUNX2), and osteocalcin (OCN)] were performed at 7 and 14 days after osteogenic induction culture. Results: The cultured cells were identified as ADSCs. With the prolongation of osteogenic induction culture, the number of calcified nodules and ALP activity significantly increased ( P<0.05). The relative expression of miR-26a-5p in the cells gradually decreased, while the relative expressions of CREB1 mRNA and protein, as well as the relative expression of p-CREB1 protein were increased. The differences were significant between 7, 14 days and 0 day ( P<0.05). There was no significant difference in p-CREB1/CREB1 between different time points ( P>0.05). The starBase database predicted that miR-26a-5p and CREB1 had targeted binding sequences, and the dual-luciferase reporter gene experiment revealed that overexpression of miR-26a-5p significantly suppressed CREB1 wild-type luciferase activity ( P<0.05). After 7 and 14 days of osteogenic induction, compared with the control group, the number of calcified nodules, ALP activity, and relative expressions of CREB1, p-CREB1, OCN, and RUNX2 proteins in the experimental group significantly increased ( P<0.05). There was no significant difference in p-CREB1/CREB1 between the two groups ( P>0.05). Conclusion: Knocking down miR-26a-5p promoted the osteogenic differentiation of ADSCs by up-regulating CREB1 and its phosphorylation.

Histone Deacetylase 3 Inhibition Ameliorates Microglia-Mediated Neuro-Inflammation Via the SIRT1/Nrf2 Pathway After Traumatic Spinal Cord Injury.

BACKGROUND: Microglial-induced inflammation plays a crucial role in the pathophysiological process of nervous system injury, however, still lacks effective therapeutic agents. Previously, we discovered that the inhibition of histone deacetylase 3 (HDAC3) exerts anti-inflammatory effects after traumatic spinal cord injury (SCI), whereas little is known about its underlying mechanism. Therefore, the present study aimed to explore the effects and potential mechanisms of HDAC3 on neuroinflammation and microglial function. METHODS: Rats were randomized into 4 groups: sham group, SCI group, SCI + vehicle group, and SCI + RGF966 group. To examine the effect of HDAC3 on neurological deficit after SCI, we gathered data using the Basso Beattie Bresnahan locomotion scale, the inclined plane test, the blood-spinal cord barrier, junction protein expression, and Nissl staining. We also evaluated microglial activation and inflammatory factor levels. Immunofluorescence analysis, immunohistochemical analysis, western blotting, and quantitative real-time polymerase chain reaction were performed to examine the regulation of the Sirtuin 1 (SIRT1)/nuclear factor-erythroid 2-related factor 2 (Nrf2) pathway. RESULTS: The results showed that HDAC3 inhibition significantly ameliorated Basso-Beattie-Bresnahan (BBB) permeability, brain edema, and improved neurological functions and junction protein levels. Additionally, HDAC3 inhibition significantly inhibited microglial activation, thereby reducing the levels of SCI-induced pro-inflammatory factors. Moreover, HDAC3 inhibition dramatically enhanced the expression of SIRT1 and increased both Nrf2 nuclear accumulation and transcriptional activity, thereby increasing downstream heme oxygenase-1 and NAD(P)H quinone oxidoreductase 1 expression. CONCLUSIONS: The results of this study suggest that HDAC3 inhibition protects the spinal cord from injury following SCI by inhibiting SCI-induced microglial activation and the subsequent inflammatory response via SIRT1/Nrf2 signaling pathway, highlighting HDAC3 as a potential therapeutic target for the treatment of SCI.

[Effectiveness of a modified posterior approach for arthroscopic resection on painful talocalcaneal coalition in adults].

OBJECTIVE: To investigate the effectiveness of arthroscopic talocalcaneal coalition resection in painful adults via a modified posterior approach. METHODS: Between January 2015 and December 2017, 9 patients with painful talocalcaneal coalition accepted arthroscopic resection via the posterior malleolus high lateral observation approach combined with the lower medial operation approach. Of them, 6 were male and 3 were female, aged from 19 to 30 years (mean, 24 years). Among them, 2 cases had no definite local trauma and 7 cases had a history of sprain of foot and ankle. The disease duration ranged from 6 to 30 months, with a median of 12 months. Rozansky classification of talocalcaneal coalition for the 9 patients: 5 cases (5 feet) were type Ⅰ, 2 cases (2 feet) type Ⅱ, and 2 cases (2 feet) type Ⅲ. The patients had no sequelae of limb dysfunction and no limb joint surgery in the past. All the patients received anteroposterior and lateral X-ray films and CT scans of the ankle joint during follow-up. The visual analogue scale (VAS) score and American Orthopaedic Foot and Ankle Society (AOFAS) ankle-hind foot score were used to evaluate the effectiveness. RESULTS: The operation time was 60-90 minutes (mean, 76 minutes). All patients were followed up 12-24 months (mean, 18 months). All the incisions healed by first intention, without infection, skin necrosis, lower extremity deep vein thrombosis, vascular nerve and tendon injury, bone bridge recurrence, and other complications. The ankle function recovered well and the pain was relieved obviously after operation, and the patients returned to work at 3-5 months after operation, with an average of 3.9 months. At last follow-up, the VAS score was 0.7±0.5, which was significantly improved ( t=20.239, P=0.000) when compared with preoperative score (4.2±0.5); the AOFAS ankle-hind foot score was 94±4, which was significantly improved ( t=-27.424, P=0.000) when compared with preoperative score (62±2). According to AOFAS ankle-hindfoot scoring system, the results were excellent in 7 cases and good in 2 cases at last follow-up. CONCLUSION: It is more intuitive, more space, and more flexibility for operation via the modified posterior malleolus high lateral observation approach combined with the lower medial operation approach in talocalcaneal coalition. It is feasible to remove talocalcaneal coalition programmatically according to the specific anatomic signs during the operation.

[Treatment of traumatic anterior dislocation of sternoclavicular joint with allogeneic tendon of "W" type knit].

OBJECTIVE: To investigate effectiveness of allogeneic tendon of "W" type knit in repair of traumatic anterior dislocation of sternoclavicular joint. METHODS: Between June 2013 and June 2017, 12 patients with traumatic anterior dislocation of sternoclavicular joint after poor conservative treatment were treated with allogeneic tendon of "W" type knit. Of them, 10 were males and 2 were females, aged from 25 to 58 years (mean, 42 years). All injuries were caused by traffic accidents. The time from injury to operation was 4-12 weeks (median, 6 weeks). All of them were closed injuries. The patients had no fracture around the shoulder, or blood vessels, nerves, and other adjacent limb joint injuries. The operation time, intraoperative blood loss, incision healing, and complications were recorded. The sternoclavicular joint was observed by X-ray film and CT at 1 year after operation. Visual analogue scale (VAS) score, University of California Los Angeles (UCLA) score, Rockwood score, modified Hospital for Special Surgery (HSS) score, and Constant-Murley score were used to evaluate the function of shoulder joint after operation. RESULTS: The operation time was 60-80 minutes (mean, 70 minutes). The intraoperative blood loss was 50-100 mL (mean, 60 mL). Primary healing of incision was obtained in all patients without complications. All the patients were followed up 12-24 months (mean, 18 months). At 1 year after operation, X-ray film and CT examination showed that the position of sternoclavicular joint was satisfactory. At 1 year after operation, the Rockwood score was 12-14 (mean, 13). The UCLA score was 28-34 (mean, 31). The VAS score was significant lower than that before operation ( P<0.05), and the Constant-Murley score and modified HSS score were significantly higher than those before operation ( P<0.05). CONCLUSION: The repair of traumatic anterior dislocationr of sternoclavicula joint with allogeneic tendon of "W" type knit can effectively reconstruct the stability of the joint, retain the physiological fretting, and obtain satisfactory results.

[Early effectiveness of arthroscopic three-point suture technique in treatment of anterior cruciate ligament tibial eminence avulsion fracture].

OBJECTIVE: To investigate the early effectiveness of three-point suture technique in treatment of anterior cruciate ligament (ACL) tibial eminence avulsion fracture by arthroscopy. METHODS: Between January 2016 and December 2017, 12 patients with ACL tibial eminence avulsion fractures underwent arthroscopic fixation of avulsion fractures with Ethibon suture using three-point suture technique. There were 9 males and 3 females, with an average of 36.4 years (range, 18-50 years). The fracture caused by traffic accident in 10 cases and sports in 2 cases. Among them, 1 patient was old fracture and 11 was fresh fracture. According to the modified Meyers-McKeever classification criteria, the fractures were rated as type Ⅲ in 7 cases and type Ⅳ in 5 cases. There were 2 cases of medial collateral ligament injury and medial meniscus injury. The preoperative International Knee Documentation Committee (IKDC) score was 37.9±4.7 and Lysholm score was 46.0±3.7. RESULTS: All operations completed smoothly. The operation time was 45-70 minutes (mean, 61.3 minutes). The incisions healed by first intention in all patients. The hospitalization stays ranged from 4 to 9 days (mean, 5 days). All patients were followed up 3-20 months (mean, 9.3 months). The anterior drawer test, Lachman test, and axis shift test in all patients were negative after operation. At last follow-up, the IKDC score was 89.7±2.5 and Lysholm score was 90.2±1.9, which were significantly higher than those before operation ( t=-30.94, P=0.00; t=-33.03, P=0.00). At last follow-up, the X-ray films showed 9 cases of fracture anatomical reduction and 3 cases of almost anatomical reduction, and 12 cases of fracture healing. CONCLUSION: For ACL tibial eminence avulsion fracture, arthroscopic three-point suture technique can effectively restore the stability of knee joint and obtain satisfactory early effectiveness.

[Clinical comparative study of thoracoscopic assisted reduction and traditional manual reduction with percutaneous intramedullary nail internal fixation for mid-clavicular fractures].

Objective: To compare the effectiveness of thoracoscopic assisted reduction and traditional manual reduction with percutaneous intramedullary nail internal fixation in the treatment of mid-clavicular fractures. Methods: A prospective randomized controlled trial was conducted. Twenty-two patients with mid-clavicular fractures who met the selection criteria between March 2012 and March 2017 were recruited and randomly divided into trial group (7 cases, thoracoscopic assisted reduction and percutaneous intramedullary nail fixation) and control group (15 cases, traditional manual reduction and percutaneous intramedullary nail fixation). There was no significant difference in gender, age, side, cause of injury, fracture classification, interval between injury and operation between the two groups ( P>0.05). The operation time and fracture healing time were recorded and compared between the two groups. The effectiveness was evaluated by Constant-Murley scale at 6 months after operation, which included subjective evaluation indexes (functional activity and pain) and objective evaluation indexes (range of motion of shoulder joint and muscle strength). Results: The operation time of the trial group was significantly longer than that of the control group ( t=5.881, P=0.000). Patients in both groups were followed up 7-20 months, with an average of 11 months. Satisfactory anatomical reduction achieved in all patients, and all incisions healed by first intension. In the control group, 1 patient had difficulty in removing the intramedullary nail, and 1 patient had fracture nonunion. No fracture nonunion or intramedullary nail rupture in the other patients of two groups. There was no significant difference in fracture healing time between the two groups ( t=0.764, P=0.453). At 6 months after operation, there was no significant difference in Constant-Murley scale between the two groups ( P>0.05). Conclusion: The treatment of the mid-clavicular fracture by using thoracoscopic assisted reduction with intramedullary nail internal fixation requires longer operation time, but does not require fluoroscopy. The effectiveness is comparable to that of traditional surgery.

[Effectiveness of anterior talofibular ligament repair in treatment of lateral ankle stability].

Objective: To evaluate the effectiveness of anterior talofibular ligament repair in the treatment of lateral ankle stability and the effect of combined tarsal sinus syndrome on results. Methods: Between December 2013 and October 2014, 47 cases of lateral ankle instability underwent anatomical repair of anterior talofibular ligament, and the clinical data were retrospectively analyzed. Of 47 cases, 32 had no tarsal sinus syndrome (group A); 15 had tarsal sinus syndrome (group B), arthroscopic debridement of tarsal sinus was performed at the same time. There was no significant difference in gender, age, disease duration, side, American Orthopaedic Foot and Ankle Society (AOFAS), Karlsson score, and Tegner movement function score between 2 groups ( P>0.05). Results: No early surgical complication of infection occurred, and primary healing of incision was obtained in 2 groups. The patients were followed up 20-31 months (mean, 26.0 months) in group A, and 20-31 months (mean, 24.7 months) in group B. Disappearance of ankle swelling, good joints movement, and recovery of normal walking were observed in all patients. At last follow-up, AOFAS score, Karlsson score, and Tegner movement function score were significantly improved when compared with preoperative ones in 2 groups ( P<0.05), but no significant difference was found between 2 groups ( P>0.05). No ankle instability recurrence was found during follow-up period. Conclusion: The effectiveness of anatomical repair of anterior talofibular ligament in lateral ankle instability is satisfactory for patients with or without tarsal sinus syndrome.

[Case-control study on suture-assisted locking plate for the treatment of proximal humeral fractures in elderly].

OBJECTIVE: To explore clinical effects of suturing-assisted locking plate in treating elderly proximal humeral fractures. METHODS: From January 2005 to January 2013, 55 elderly patients with three- and four-part fractures of proximal humeral fractures were divided into treatment group and control group. In treatment group, there were 31 patients including 12 males, and 19 females aged from 65 to 85 with an average of (74.00±5.42) years old, and treated with suturing-assisted locking plates; 19 patients were Neer 3-part fractures, and 12 patients were Neer 4-part fractures of proximal humerus; 23 patients were suffered from low-energy injuries and 8 patients were caused by high-energy injuries. In control group, there were 24 patients including 7 males, and 17 females aged from 65 to 85 with an average of (72.79±5.34) years old, and treated with locking plates; 16 patients were Neer 3-part fractures, and 8 patients were Neer 4-part fractures of proximal humerus; 17 patients were suffered from low-energy injuries and 7 patients were caused by high-energy injuries. Operative time, blood loss during operation, and bone healing time between two groups were observed and compared. Postoperative Neer scoring were used to evaluate recovery of shoulder joint function. RESULTS: All patients were followed up from 6 to 24 months with an average of 16.1 months. In treatment group, blood loss was (495.806±143.150) ml, function of Neer scoring was 22.645±2.443, range of action was 18.194±2.613, anatomy was 7.935±1.504 and total score of Neer scoring was 77.161±8.335; while in control group, blood loss was (641.667±169.851) ml, function of Neer scoring was 13.958±1.989, range of action was 13.083±2.165, anatomy was 5.500±1.978 and total score of Neer scoring was 58.792±7.313. There were sigificant difference between two groups in these indexes. CONCLUSION: Suturing-assisted locking plate for the treatment of proximal humerus fractures in elderly, has advantages of less blood loss, simple fracture reduction and rapid recovery of shoulder joint, and is a effective method.