BackgroundChemoresistance is one of the leading causes that severely limits the success of osteosarcoma treatment. Evaluating chemoresistance before chemotherapy poses a new challenge for researchers. We established an effective chemoresistance risk scoring model for prechemotherapy osteosarcoma using single-cell sequencing.MethodsWe comprehensively analyzed osteosarcoma data from the bulk mRNA sequencing dataset TARGET-OS and the single-cell RNA sequencing (scRNA-seq) dataset GSE162454. Chemoresistant tumor clusters were identified using enrichment analysis and AUCell scoring. Its differentiated trajectory was achieved with inferCNV and pseudotime analysis. Ligand–receptor interactions were annotated with iTALK. Furthermore, we established a chemoresistance risk scoring model using LASSO regression based on scRNA-seq-based markers of chemoresistant tumor clusters. The TARGET-OS dataset was used as the training group, and the bulk mRNA array dataset GSE33382 was used as the validation group. Finally, the performance was verified for its discriminatory ability and calibration.ResultsUsing bulk RNA data, we found that osteogenic expression was upregulated in chemoresistant osteosarcoma as compared to chemosensitive osteosarcoma. Then, we transferred the bulk RNA findings to scRNA-seq and noticed osteosarcoma tumor clusters C14 and C25 showing osteogenic cancer stem cell expression patterns, which fit chemoresistant characteristics. C14 and C25 possessed bridge roles in interactions with other clusters. On the one hand, they received various growth factor stimulators and could potentially transform into a proliferative state. On the other hand, they promote local tumor angiogenesis, bone remodeling and immunosuppression. Next, we identified a ten-gene signature from the C14 and C25 markers and constructed a chemoresistant risk scoring model using LASSO regression model. Finally, we found that chemoresistant osteosarcoma had higher chemoresistance risk score and that the model showed good discriminatory ability and calibration in both the training and validation groups (AUCtrain = 0.82; AUCvalid = 0.84). Compared with that of the classic bulk RNA-based model, it showed more robust performance in validation environment (AUCvalid-scRNA = 0.84; AUCvalid-bulk DEGs = 0.54).ConclusionsOur work provides insights into understanding chemoresistant osteosarcoma tumor cells and using single-cell sequencing to establish a chemoresistance risk scoring model. The model showed good discriminatory ability and calibration and provided us with a feasible way to evaluate chemoresistance in prechemotherapy osteosarcoma.

BackgroundOsteoporosis often occurs with carotid atherosclerosis and causes contradictory calcification across tissue in the same patient, which is called the “calcification paradox”. Circulating monocytes may be responsible for this unbalanced ectopic calcification. Here, we aimed to show how CD14+ monocytes contribute to the pathophysiology of coexisting postmenopausal osteoporosis and carotid atherosclerosis.MethodsWe comprehensively analyzed osteoporosis data from the mRNA array dataset GSE56814 and the scRNA-seq dataset GSM4423510. Carotid atherosclerosis data were obtained from the GSE23746 mRNA dataset and GSM4705591 scRNA-seq dataset. First, osteoblast and vascular SMC lineages were annotated based on their functional expression using gene set enrichment analysis and AUCell scoring. Next, pseudotime analysis was applied to draw their differentiated trajectory and identify the key gene expression changes in crossroads. Then, ligand–receptor interactions between CD14+ monocytes and osteoblast and vascular smooth muscle cell (SMC) lineages were annotated with iTALK. Finally, we selected calcification paradox-related expression in circulating monocytes with LASSO analysis.ResultsFirst, we found a large proportion of delayed premature osteoblasts in osteoporosis and osteogenic SMCs in atherosclerosis. Second, CD14+ monocytes interacted with the intermediate cells of the premature osteoblast and osteogenic SMC lineage by delivering TGFB1 and TNFSF10. This interaction served as a trigger activating the transcription factors (TF) SP1 and NFKB1 to upregulate the inflammatory response and cell senescence and led to a retarded premature state in the osteoblast lineage and osteogenic transition in the SMC lineage. Then, 76.49% of common monocyte markers were upregulated in the circulating monocytes between the two diseases, which were related to chemotaxis and inflammatory responses. Finally, we identified 7 calcification paradox-related genes on circulating monocytes, which were upregulated in aging cells and downregulated in DNA repair cells, indicating that the aging monocytes contributed to the development of the two diseases.ConclusionsOur work provides a perspective for understanding the triggering roles of CD14+ monocytes in the development of the calcification paradox in osteoporosis- and atherosclerosis-related cells based on combined scRNA and mRNA data. This study provided us with an elucidation of the mechanisms underlying the calcification paradox and could help in developing preventive and therapeutic strategies.

Background: This study aimed to identify the key genes related to male ankylosing spondylitis (AS) and to analyze the role of immune cell infiltration in the pathological process of this disease. Methods:The AS dataset was downloaded from the Gene Expression Omnibus (GEO) public database, and the data of male healthy controls (M_HC) and male AS patients (M_AS) were extracted. R software was used to identify differentially expressed genes (DEGs). Functional and pathway enrichment analysis of the DEGs was performed. A protein-protein interaction (PPI) network was constructed, and the hub genes were screened out. All expression profile data were analyzed by weighted correlation network analysis (WGCNA) to screen out the hub genes, which were then intersected with the hub genes from the PPI network to obtain the key genes. Finally, the difference in immune cell infiltration in the two sets of samples was evaluated with CIBERSORT, and the correlation between the key genes and infiltrating immune cells was analyzed.Results: A total of 689 DEGs were obtained, of which 395 genes were up-regulated and 294 genes were down-regulated. Functional and pathway enrichment analysis showed that DEGs were mainly enriched in pathways related to immune response. Based on the PPI analysis, five clusters with high scores were selected.Through WGCNA, 14 gene modules were obtained. The green module with the highest correlation was selected and intersected with the cluster previously obtained to obtain three key genes, RAB5C, SYNJ1, and RNF19B. Immune infiltration analysis found that monocytes and gamma delta T cells may be involved in the process of AS. Also, RAB5C, SYNJ1, and RNF19B are all related to increased levels of monocytes and macrophages.Conclusions: RAB5C, SYNJ1, and RNF19B are key DEGs expressed in M_AS and may play a role in the disease's occurrence and development through regulating immune cell functions.

Objective At present, the true sagittal alignment of the cervical spine is uncertain, resulting in no standard reference for subaxial cervical surgery. So, we aimed to explore the age difference of normal cervical sagittal alignment and to further investigate the mid‐and long‐term changes of sagittal alignment after subaxial cervical spine surgery. Materials and Methods This was a retrospective study and 1223 asymptomatic volunteers and 79 patients undergoing subaxial cervical spine surgery were retrospectively reviewed in total. Asymptomatic volunteers and patients were divided into six subgroups: 20–29, 30–39, 40–49, 50–59, 60–69 and ≥70 groups. The age difference and trend with age of cervical sagittal parameters of asymptomatic volunteers were assessed by cervical lateral radiography and analyzed by ANOVA test, and the regression equation of C2‐7 Cobb was established via multiple linear regression. Based on the C2‐7 Cobb regression equations of different ages, the theoretical value, deviation value, loss value of the C2‐7 Cobb, and JOA recovery rate of patients were calculated, and the correlation among the loss value, deviation value of the C2‐7 Cobb, and JOA recovery rate of the 79 patients was evaluated by Pearson correlation analysis. Results For the asymptomatic volunteers, the C0‐2 Cobb decreased gradually with increasing age. The C2‐7 Cobb, C2‐7 SVA, T1S, NT, and TIA increased gradually with increasing age. The CBVA fluctuated with increasing age. T1S demonstrated a moderate correlation with C2‐7 Cobb ( r = 0.60, p < 0.01); C0‐2 Cobb, C2‐7 SVA, CBVA, and TIA demonstrated a fair correlation with C2‐7 Cobb ( r = −0.30, −0.33, 0.41, 0.40, p < 0.01); age demonstrated a poor correlation with C2‐7 Cobb ( r = 0.19, p < 0.01). The regression equations of C2‐7 Cobb were established using C0‐2 Cobb, C2‐7 SVA, CBVA, and T1S. For the patients with subaxial cervical spine surgery, the loss of C2‐7 Cobb was moderately correlated with the deviation of C2‐7 Cobb ( r = 0.33, p < 0.01). Conclusion The age difference of cervical sagittal alignment was obvious, and the C2‐7 Cobb increased with age especially. The closer the postoperative C2‐7 Cobb was to the theoretical value of corresponding age, the smaller the loss of correction angle was, and the better the mid‐ and long‐term outcomes. The personalized sagittal reconstruction should be performed according to age difference for subaxial cervical spine surgery.

Background: To explore the feasibility and efficiency of posterior wedge osteotomy assisted by O-arm navigation treatment of ankylosing spondylitis (AS) patients with thoracolumbar fracture. Methods: This is a case series study. A total of 16 cases of AS accompanied by thoracolumbar fractures from January 2012 to July 2015 were retrospectively analyzed. All patients underwent "posterior wedge osteotomy assisted by O-arm navigation". The operation time, blood loss volume, preoperative and postoperative visual analogue scale (VAS), American Spinal Injury Association (ASIA) classification, and spinal imaging parameters [Cobb angle, C7 plumb line (C7PL), and jaw-brow angle] were collected and analyzed. Results: The operative time consumption was 120-350 mins and the intra-operative blood loss volume was 200-800 mL. No obvious postoperative complications occurred in any participants. The back pain of all participants was relieved, and the neurological functions of eight participants with spinal cord injury (SCI)were recovered in varying degrees except for one patient with severe SCI. The spinal deformities of the participants were corrected to varying degrees. The fracture sites of 16 participants were all healed, and there was no loosening or detachment of internal fixation during the 6-month follow-up period.Conclusions: Posterior wedge osteotomy assisted by O-arm navigation was shown to be a safe and effective method to treat AS accompanied by thoracolumbar fractures. This treatment strategy can accurately decompress and reduce the fracture and significantly correct the kyphosis, with good surgical effect.