Nucleus Pulposus Cells from Calcified Discs Promote the Degradation of the Extracellular Matrix through Upregulation of the GATA3 Expression.

OBJECTIVE: Disc calcification is strongly associated with disc degeneration; however, the underlying mechanisms driving its pathogenesis are poorly understood. This study aimed to provide a gene expression profile of nucleus pulposus cells (NPCs) from calcified discs, and clarify the potential mechanism in disc degeneration. METHODS: Primary NPCs were isolated from calcified and control discs (CAL-NPC and CON-NPC), respectively. The proliferation and extracellular matrix (ECM) metabolism capacities of the cells were evaluated using MTT and Western blotting, respectively. RNA sequencing was used to identify differentially expressed genes (DEGs) in the CAL-NPCs. The biological functions of the DEGs were analyzed using the Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) databases. The transcription factor database and Cytoscape software were used to construct the transcription factor-DEGs regulatory network. The role of the verified transcription factor in NPC proliferation and ECM metabolism was also investigated. RESULTS: The CAL-NPCs exhibited a lower proliferation rate and higher ECM degradation capacity than the CON-NPCs. In total, 375 DEGs were identified in the CAL-NPCs. The GO and KEGG analyses showed that the DEGs were primarily involved in the regulation of ribonuclease activity and NF-kappa B and p53 signaling pathways. GATA-binding protein 3 (GATA3) with the highest verified levels was selected for further studies. Overexpression of GATA3 in the CON-NPCs significantly inhibited their proliferation and promoted their ECM degradation function, while the knockdown of GATA3 in the CAL-NPCs resulted in the opposite phenotypes. CONCLUSION: This study provided a comprehensive gene expression profile of the NPCs from the calcified discs and supported that GATA3 could be a potential target for reversing calcification-associated disc degeneration.

Comparison of blood loss between tranexamic acid-soaked absorbable Gelfoam and topical retrograde injection via drainage catheter plus clamping in cervical laminoplasty surgery.

BACKGROUND: To compare the safety and efficacy of tranexamic acid (TXA)-soaked absorbable Gelfoam and the retrograde injection of TXA through a drain with drain-clamping in degenerative cervical laminoplasty patients. METHODS: Patients were assigned into either TXA retrograde injection (TXA-RI), TXA-soaked absorbable Gelfoam (TXA-Gel), or control groups. The demographics, operative measurements, volume and length of drainage, length of hospital stay, complete blood cell count, coagulopathy, postoperative complications, and blood transfusion were recorded. RESULTS: We enrolled 133 patients, with 44, 44, and 45 in the TXA-RI, TXA-Gel, and control groups, respectively. The baseline characteristics did not differ significantly among the three groups. The TXA-RI group exhibited a lower volume and length of postoperative drainage compared to the TXA-Gel and control groups (126.60 ± 31.27 vs. 156.60 ± 38.63 and 275.45 ± 75.27 mL; 49.45 ± 9.70 vs 58.70 ± 10.46 and 89.31 ± 8.50 hours, all P < 0.01). The TXA-RI group also had significantly shorter hospital stays compared to the control group (5.31 ± 1.18 vs 7.50 ± 1.25 days, P < 0.05) and higher hemoglobin and hematocrit levels (12.58 ± 1.67 vs 11.28 ± 1.76 g/dL; 36.62 ± 3.66% vs 33.82 ± 3.57%, both P < 0.05) at hospital discharge. In the TXA-RI and TXA-Gel groups, the D-dimmer (DD) and fibrinogen (FIB) were significantly lower than those in the control group after surgery (P < 0.05). None of the patients required blood transfusion. No complications, including thromboembolic events, were reported. CONCLUSION: Topical retrograde injection of TXA through a drain with drain-clamping at the conclusion of unilateral posterior cervical expansive open-door laminoplasty may effectively reduce postoperative blood loss and the length of hospital stays without increasing postoperative complications.

Phosphoglycerate kinase 1 silencing by a novel microRNA microRNA-4523 protects human osteoblasts from dexamethasone through activation of Nrf2 signaling cascade.

Nuclear-factor-E2-related factor 2 (Nrf2) cascade activation can ameliorate dexamethasone (DEX)-induced oxidative injury and death in human osteoblasts. Phosphoglycerate kinase 1 (PGK1) depletion is shown to efficiently activate Nrf2 signaling by inducing methylglyoxal modification of Kelch-like ECH-associated protein 1 (Keap1). We here identified a novel PGK1-targeting microRNA: microRNA-4523 (miR-4523). RNA fluorescent in situ hybridization, RNA pull-down, and Argonaute-2 RNA immunoprecipitation results confirmed a direct binding between miR-4523 and PGK1 mRNA in primary human osteoblasts and hFOB1.19 osteoblastic cells. Forced overexpression of miR-4523, using a lentiviral construct, robustly decreased PGK1 3'-UTR (untranslated region) luciferase activity and downregulated its expression in human osteoblasts and hFOB1.19 cells. Furthermore, miR-4523 overexpression activated the Nrf2 signaling cascade, causing Keap1-Nrf2 disassociation, Nrf2 protein stabilization, and its nuclear translocation as well as transcription activation of Nrf2-dependent genes (NQO1, GCLC, and HO1) in human osteoblasts. By expressing a UTR-null PGK1 construct, miR-4523 overexpression-induced Nrf2 cascade activation was however largely inhibited. Importantly, DEX-induced reactive oxygen species production, oxidative injury, and cell apoptosis were significantly attenuated by miR-4523 overexpression in human osteoblasts and hFOB1.19 cells. Such actions by miR-4523 were abolished by Nrf2 shRNA or knockout, but mimicked by PGK1 knockout (using CRISPR/Cas9 method). In PGK1 knockout human osteoblasts, miR-4523 overexpression failed to further increase Nrf2 cascade activation and offer osteoblast cytoprotection against DEX. Significantly, miR-4523 is downregulated in human necrotic femoral head tissues of DEX-taking patients. Together, PGK1 silencing by miR-4523 protected human osteoblasts from DEX through activation of the Nrf2 signaling cascade.

DCAF1-targeting microRNA-3175 activates Nrf2 signaling and inhibits dexamethasone-induced oxidative injury in human osteoblasts.

Activation of nuclear-factor-E2-related factor 2 (Nrf2) signaling can protect human osteoblasts from dexamethasone-induced oxidative injury. DDB1 and CUL4 associated factor 1 (DCAF1) is a novel ubiquitin E3 ligase for Nrf2 protein degradation. We identified a novel DCAF1-targeting miRNA, miR-3175. RNA pull-down, Argonaute 2 RNA-immunoprecipitation, and RNA fluorescent in situ hybridization results confirmed a direct binding between miR-3175 and DCAF1 mRNA in primary human osteoblasts. DCAF1 3'-untranslated region luciferase activity and its expression were significantly decreased after miR-3175 overexpression but were augmented with miR-3175 inhibition in human osteoblasts and hFOB1.19 osteoblastic cells. miR-3175 overexpression activated Nrf2 signaling, causing Nrf2 protein stabilization, antioxidant response (ARE) activity increase, and transcription activation of Nrf2-dependent genes in human osteoblasts and hFOB1.19 cells. Furthermore, dexamethasone-induced oxidative injury and apoptosis were largely attenuated by miR-3175 overexpression in human osteoblasts and hFOB1.19 cells. Importantly, shRNA-induced silencing or CRISPR/Cas9-mediated Nrf2 knockout abolished miR-3175 overexpression-induced osteoblast cytoprotection against dexamethasone. Conversely, DFAC1 knockout, by the CRISPR/Cas9 method, activated the Nrf2 cascade and inhibited dexamethasone-induced cytotoxicity in hFOB1.19 cells. Importantly, miR-3175 expression was decreased in necrotic femoral head tissues of dexamethasone-taking patients, where DCAF1 mRNA was upregulated. Together, silencing DCAF1 by miR-3175 activated Nrf2 signaling to inhibit dexamethasone-induced oxidative injury and apoptosis in human osteoblasts.

Topical Injection of Tranexamic Acid via a Drain Plus Drain-Clamping to Reduce Blood Loss in Degenerative Lumbar Scoliosis Surgery.

OBJECTIVE: The aim of the present study was to investigate whether an innovative way of administering tranexamic acid (TXA), that is, injecting it retrogradely through the drain and clamping it for 1 h, can reduce postoperative bleeding after degenerative lumbar scoliosis surgery. METHODS: Sixty degenerative lumbar scoliosis patients who underwent posterior lumbar decompression with fusion of three or more levels were retrospectively enrolled and categorized into three groups (TXA, Gelfoam, and control groups). The demographic distribution, operative parameters, length and amount of Hemovac drainage, blood transfusion rate, length of stay, laboratory results (complete blood count and coagulogram), and the postoperative complications were collected and analyzed. RESULTS: The age of patients in the Gelfoam group was significantly younger than in the TXA and control groups (59.75 ± 6.95 vs 66.10 ± 8.80, P = 0.016 and 59.75 ± 6.95 vs 67.90 ± 5.33, P = 0.000, respectively). There were no significant differences in sex, body mass index, comorbid medical status, and operation level between each of the two groups. The three groups did not differ significantly in estimated blood loss during surgery, the mean red blood cell transfusion requirement during hospitalization, and the entire perioperative allogenic blood transfusion rate. The postoperative total blood loss and total drainage were lower in the TXA group than in the control group (1027.14 ± 466.56 vs 1390.07 ± 314.85 mL, P = 0.006; 322.20 ± 187.32 vs 605.50 ± 184.70 mL, P = 0.000, respectively). The length of drainage retention in the TXA group was significantly shorter than in the Gelfoam and control groups (46.10 ± 9.00 vs 68.00 ± 12.31 h, P = 0.000 and 46.10 ± 9.00 vs 76.40 ± 10.97 h, P = 0.000, respectively). The TXA and Gelfoam groups also had significantly shorter hospital stays than the control group (7.50 ± 0.95 vs 9.80 ± 2.44 days, P = 0.000, and 7.90 ± 1.16 vs 9.80 ± 2.44 days, P = 0.003, respectively). At discharge, the mean hemoglobin and hematocrit level were significantly higher in the TXA group compared with the control group (11.77 ± 1.78 g/dL vs 10.67 ± 0.94 g/dL, P = 0.002; 34.82 ± 3.57% vs 31.79 ± 3.85%, P = 0.014). No significant difference was identified with respect to prothrombin time, activated partial thromboplastin time, and D-dimmer among groups (P > 0.05). The three groups were comparable in wound problem incidences. Symptomatic deep vein thrombosis and pulmonary embolism were not observed in this study. CONCLUSION: Topical injection of TXA retrogradely via a drain at the end of a degenerative lumbar scoliosis operation and clamping the drain for an hour can effectively decrease the postoperative blood loss and the length of hospitalization without increasing the complication rate.

Revision Surgery after Percutaneous Endoscopic Transforaminal Discectomy Compared with Primary Open Surgery for Symptomatic Lumbar Degenerative Disease.

OBJECTIVE: To evaluate the clinical outcome of reoperation after percutaneous endoscopic lumbar discectomy (PELD) as compared with primary spinal decompression and fusion. METHODS: A retrospective study from December 2014 to December 2017 was conducted at Peking Union Medical College Hospital and comprised 39 patients with symptomatic lumbar degenerative disease (LDD): 13 post-PELD who underwent reoperation (revision surgery group) and 26 who received primary spinal decompression and fusion (primary open surgery group). The two groups were compared regarding: operative time, blood loss, transfusion, hospitalization, postoperative visual analog scale (VAS) scores, Oswestry Disability Index (ODI) scores, Japanese Orthopedic Association (JOA) improvement rate, and postoperative complications. The Mann-Whitney U-test was applied to analyze continuous parameters, and the χ2 -test for categorical parameters. Fisher's exact test was used for small data subsets. RESULTS: There was no statistically significant difference between the two groups in mean age (52.7 years vs 52.9 years), gender ratio (6 men-to-7 women vs 12 men-to-14 women), body mass index, medical history, preoperative diagnosis, or surgical spine level (P > 0.05). The mean operative time of the revision surgery group was significantly longer than that of the primary open surgery group (160.0 min vs 130.2 min, P < 0.05). The revision surgery group also had a significantly higher mean estimated blood loss, postoperative drainage, and length of hospital stay (P < 0.05). However, no significant differences were found between the two groups in terms of hemoglobin and hematocrit values, preoperatively and postoperatively. The rate of transitional neurological irritation was higher in the revision surgery group (61.5% vs 3.8%; P < 0.05), as was intraoperative durotomy and cerebrospinal fluid leakage (30.8% vs 3.8%, P < 0.05). At 1 month, the VAS and ODI scores of the primary open surgery group were significantly better than those of the revision surgery group, while the improvement in JOA scores was similar. After 6 and 12 months' follow-up, the VAS and ODI scores and the rates of JOA improvement were comparable. CONCLUSION: Patients with LDD who received primary spinal decompression and fusion experienced lower rates of perioperative complications and shorter hospitalization compared with patients who underwent revision surgery after PELD, but the clinical outcomes at the last follow-up of both groups were satisfactory.

Correlation between severity of adolescent idiopathic scoliosis and pulmonary artery systolic pressure: a study of 338 patients.

PURPOSE: Study of patients with adolescent idiopathic scoliosis. OBJECTIVE: To examine the correlation between pulmonary arterial pressure and coronal Cobb angle of idiopathic scoliosis. METHODS: A total of 338 patients (82.8 % female) with idiopathic scoliosis (average age 15.6 years; range 14-20 years) were included. Preoperatively, the coronal Cobb angle of curvature and the apex location and direction were determined from radiographic records. Tricuspid regurgitation velocity (TRV) and inferior vena cava diameter were also measured using Doppler echocardiography. Pulmonary arterial systolic pressure (sPAP) was calculated from the TRV according to the modified Bernoulli equation and correlations between sPAP and the features of scoliosis were identified by statistical analysis. RESULTS: Among the 338 patients, there were 305 thoracic curves, 276 (90.5 %) of which were right curves, and 265 thoracolumbar/lumbar curves. sPAP varied from 5.0 to 37.6 mmHg. Pulmonary hypertension could not be excluded in the case of one patient. A mild correlation (Spearman test, correlation coefficient = 0.187, P = 0.001) between sPAP and coronal Cobb angle of the main thoracic (MT) curves was identified. Correlations between sPAP and the degree of other curves were not significant. Patients with sPAP >20 mmHg also had larger thoracic curve angles (mean MT 42.16° vs. 52.45°; U test, P = 0.002). There were no differences in sPAP levels between patients with right and left thoracic curves. CONCLUSIONS: A mild positive correlation was identified between sPAP and the coronal Cobb angle of the MT curves. There was no relationship between sPAP and the direction of the curvature.

Identification of long noncoding RNA associated with osteoarthritis in humans.

OBJECTIVE: Long noncoding RNAs (lncRNAs) are an important class of genes involved in various biological functions; however, knowledge about lncRNAs in osteoarthritis (OA) is limited. Therefore, the present study aimed to identify which lncRNAs are expressed in OA versus normal cartilage. METHODS: To identify lncRNAs specifically expressed in OA cartilage, expression of lncRNAs in OA cartilage was compared with that in normal cartilage using microarray analysis. The identified differences in expression of lncRNAs were validated by real time polymerase chain reaction (RT-PCR). Furthermore, expression of several key mRNAs associated with OA, including those for matrix metalloproteinase (MMP)-9, MMP-13, bone morphogenetic protein (BMP)-2, COL2A1 and ADAMTS5, was investigated by RT-PCR in OA and normal cartilage. RESULTS: Microarray analysis identified 121 lncRNAs that were up- or down-regulated in OA compared with normal tissue, 73 being upregulated and 48 downregulated compared with normal cartilage. Twenty-one of the above differently expressed lncRNAs were up-regulated twofold. Expression of six lncRNAs, including HOTAIR, GAS5, PMS2L2, RP11-445H22.4, H19 and CTD-2574D22.4, was up-regulated in OA compared with normal tissue as validated by RT-PCR after microarray analysis. Expression of mRNA for MMP-9, MMP-13, BMP-2, and ADAMTS5 in OA was significantly greater than in normal cartilage. However, expression of mRNA for COL2A1 was lower in OA than in normal cartilage. CONCLUSION: The differently expressed lncRNAs may be associated with the pathogenesis of OA. Further functional studies are critical to confirming the function of lncRNAs in OA and to exploring new potential targets for therapy.

[Experimental study of three dimensional navigation assisted spinal surgery by multi-segment registration technology based on structured light scanning].

OBJECTIVE: To study the registration method based on structured light scanning for navigation assisted spinal surgery and assess its accuracy so as to construct a registration system for the navigation assisted spinal surgery using structured light scanning. METHODS: Both the computed tomographic (CT) dataset and the structured light scanning images of thoracic vertebra were obtained. The pre-registration and multi-segment iterative closest point (ICP) algorithm were used for the registration of CT images and structured light images. Four segmentations were selected from the surface of thoracic vertebra and placed into different combinations. The accuracy for each combination was studied. Noise and perturbation were exerted to structured light and registration accuracy was studied. And calf vertebra was used for further verification. RESULTS: A combination of pre-registration and multi-segment iterative closest point (ICP) algorithm was competent for the registration of CT scanning data and the structured light scanning data. The registration error was less than 1 mm when two and more segments were selected for registration combination. The registration error was less than 1 mm when noise was exerted. CONCLUSION: With a high accuracy and a perturbation resistance, a combination of pre-registration and multi-segment registration algorithm based on structured light scanning is competent for the registration of CT scanning data and structured light scanning data.

[Selection of distal fusion levels in the surgical management of thoracolumbar/lumbar adolescent idiopathic scoliosis].

OBJECTIVE: To investigate the efficacy and safety of a new method which determines the exact distal fusion level in the treatment of adolescent idiopathic scoliosis (AIS) with posterior pedicle screw fixation and to assess its clinical outcome. METHODS: This prospective clinical study analyzed 31 AIS patients who met the inclusion criteria enrolled from July 2005 to September 2008. Based on the principle of our new criteria for selection of distal fusion level of AIS, all patients had posterior spinal fusion and instrumentation with pedicle screws. Cobb angle of the curve, tilt angle of the LIV, intervertebral angle and trunk shift were measured and analyzed. RESULTS: Preoperative (42 + or - 17) degrees of thoracic curve was corrected to (12 + or - 7) degrees , with a curve correction of 70.6%. Preoperative (44 + or - 7) degrees of lumbar curve was corrected to (9 + or - 4) degrees , with a curve correction of 80.2%. The trunk shift were significantly improved from (13 + or - 8) mm to (9 + or - 7) mm before and after surgery respectively (P < 0.05). The tilt angles of the LIV before and after surgery were (20.8 + or - 5.7) degrees and (1.5 + or - 3.1) degrees respectively. The thoracic Cobb angle was (14 + or - 8) degrees and the lumbar Cobb angle was (9 + or - 5) degrees at latest follow up. The changes were of significance in the tilt angle of the LIV after surgery compared with that before surgery (P = 0.000). This angle averaged (0.8 + or - 3.7) degrees at final follow up, but the change was not significant compared with that after surgery (P > 0.05). CONCLUSION: This is an effective method with the advantage of shortening the fusion level, reserving the distal motion segments and easing segmental degeneration adjacent to the fusion area.

[Differential expressed proteins of knee osteoarthritic chondrocytes].

OBJECTIVE: To investigate the molecular mechanism of osteoarthritis (OA) through the differential expressed proteins of advanced osteoarthritic chondrocytes. METHODS: Normal articular cartilage (NAC, n = 22) were obtained from knee cartilage of donors without knee diseases and osteoarthritic cartilage (OAC, n = 17) from OA patients undergoing knee arthroplasty. Total protein was extracted from chondrocytes and loaded directly for two-dimensional gel electrophoresis (2-DE). Gel images were analyzed with the software Imagemaster 2-D platinum3.0 to screen the differential expression protein spots. Target spots were excised and digested and the resulting peptides submitted for mass spectrometry analysis. Identities of differential expressed proteins were recognized through matching the peptide mass finger printing of each spot with NCBI protein database by MASCOT. RESULTS: About 1000 protein spots were presented in the 2-DE gels for each group. Thirty-five NAC spots and 31 OAC spots were confirmed to be differential expression protein spots. Among these spots, 19 proteins were identified by MASCOT, including type VI collagen, enzymes involved in collagen synthesis, shock related proteins and some novel proteins with unknown functions. CONCLUSION: The differential expressed proteins isolated and identified by the present study provide valuable information on various aspects of OA pathology. It may help us to gain new insights into the molecular mechanism of OA.

A retrospective study of echocardiographic cardiac function and structure in adolescents with congenital scoliosis.

BACKGROUND: Patients with congenital scoliosis often also have intraspinal abnormalities and other organ defects, and few studies of the effects of congenital scoliosis on cardiac function and structure have been published. METHODS: A total of 215 adolescent patients with congenital scoliosis (average age, 13.58 years) underwent preoperative echocardiography and were then assigned to subgroups according to apex vertebral rotation, side of convexity, curvature severity in the coronal and sagittal planes, type of deformity, and sex. Differences between the subgroups were compared by independent-samples t test or a one-factor analysis of variance. RESULTS: We observed statistically significant differences between patients with right-sided scoliosis curvature and those with left-sided scoliosis curvature, respectively, in left ventricular inner diameter at end-diastole ((39.39 +/- 4.66) mm vs (41.74 +/- 4.90) mm), left ventricular inner diameter at end-systole ((24.80 +/- 3.45) mm vs (25.92 +/- 3.07) mm), interventricular septum thickness at end-diastole ((5.66 +/- 0.98) mm vs (5.98 +/- 1.03) mm), and posterior wall of left ventricle at end-diastole ((5.61 +/- 0.98) mm vs (6.06 +/- 1.20) mm). When the patients were evaluated by coronal plane Cobb angle, significant differences were found between those with Cobb angle of 40 degrees - 80 degrees and of > 80 degrees in left ventricular inner diameter at end-diastole ((40.97 +/- 5.06) mm vs (38.98 +/- 4.45) mm) and left ventricular inner diameter at end-systole ((25.53 +/- 3.39) mm vs (24.36 +/- 3.14) mm), respectively. When the patients were evaluated by sagittal plane Cobb angle (< 20 degrees, group 1; 20 degrees - 40 degrees, group 2; > 40 degrees, group 3), significant differences were found in right ventricular diameter between those with Cobb angle of < 20 degrees and of 20 degrees - 40 degrees ((18.27 +/- 3.66) mm vs (16.54 +/- 3.57) mm) and in diameter of aortic root between those with Cobb angle of 20 degrees - 40 degrees and of > 40 degrees ((23.83 +/- 3.39) mm vs (24.90 +/- 3.30) mm), respectively. No significant differences were found in ejection fraction and fractional shortening between patients according to apex vertebral rotation, side of convexity, coronal plane and sagittal plane Cobb angles, type of deformity, or sex. CONCLUSIONS: Congenital scoliosis influences cardiac structure, but not function.