T7 RNA polymerase (T7 RNAP) is one of the simplest known RNA polymerases. Its unique structural features make it a critical model for studying the mechanisms of RNA synthesis. This review systematically examines the static crystal structure of T7 RNAP, beginning with an in-depth examination of its characteristic “thumb”, “palm”, and “finger” domains, which form the classic “right-hand-like” architecture. By detailing these structural elements, this review establishes a foundation for understanding the overall organization of T7 RNAP. This review systematically maps the functional roles of secondary structural elements and their subdomains in transcriptional catalysis, progressively elucidating the fundamental relationships between structure and function. Further, the intrinsic flexibility of T7 RNAP and its applications in research are also discussed. Additionally, the review presents the structural diagrams of the enzyme at different stages of the transcription process, and through these diagrams, it provides a detailed description of the complete transcription process of T7 RNAP. By integrating structural dynamics and kinetics analyses, the review constructs a comprehensive framework that bridges static structure to dynamic processes. Despite its advantages, T7 RNAP has a notable limitation: it generates double-stranded RNA (dsRNA) as a byproduct. The presence of dsRNA not only compromises the purity of mRNA products but also elicits nonspecific immune responses, which pose significant challenges for biotechnological and therapeutic applications. The review provides a detailed exploration of the mechanisms underlying dsRNA formation during T7 RNAP catalysis, reviews current strategies to mitigate this issue, and highlights recent progress in the field. A key focus is the semi-rational design of T7 RNAP mutants engineered to minimize dsRNA generation and enhance catalytic performance. Beyond its role in transcription, T7 RNAP exhibits rapid development and extensive application in fields, including gene editing, biosensing, and mRNA vaccines. This review systematically examines the structure-function relationships of T7 RNAP, elucidates the mechanisms of dsRNA formation, and discusses engineering strategies to optimize its performance. It further explores the engineering optimization and functional expansion of T7 RNAP. Furthermore, this review also addresses the pressing issues that currently need resolution, discusses the major challenges in the practical application of T7 RNAP, and provides an outlook on potential future research directions. In summary, this review provides a comprehensive analysis of T7 RNAP, ranging from its structural architecture to cutting-edge applications. We systematically examine: (1) the characteristic right-hand domains (thumb, palm, fingers) that define its minimalistic structure; (2) the structure-function relationships underlying transcriptional catalysis; and (3) the dynamic transitions during the complete transcription cycle. While highlighting T7 RNAP’s versatility in gene editing, biosensing, and mRNA vaccine production, we critically address its major limitation—dsRNA byproduct formation—and evaluate engineering solutions including semi-rationally designed mutants. By synthesizing current knowledge and identifying key challenges, this work aims to provide novel insights for the development and application of T7 RNAP and to foster further thought and progress in related fields.

Apical microsurgery is accurate and minimally invasive, produces few complications, and has a success rate of more than 90%. However, due to the lack of awareness and understanding of apical microsurgery by dental general practitioners and even endodontists, many clinical problems remain to be overcome. The consensus has gathered well-known domestic experts to hold a series of special discussions and reached the consensus. This document specifies the indications, contraindications, preoperative preparations, operational procedures, complication prevention measures, and efficacy evaluation of apical microsurgery and is applicable to dentists who perform apical microsurgery after systematic training.
Microsurgery/standards* ; Humans ; Apicoectomy ; Contraindications, Procedure ; Tooth Apex/diagnostic imaging* ; Postoperative Complications/prevention & control* ; Consensus ; Treatment Outcome

ObjectiveAlthough expression of the TEAD1 protein in preadipocytes has been established, its function remains unclear. In this study, we sought to detect transcripts of TEAD1 in chicken and to examine the effects of this protein on the proliferation, migration, apoptosis, and differentiation of immortalized chicken preadipocyte cell lines (ICP1). MethodsThe full-length sequence of the TEAD1 gene was cloned and the two transcripts were subjected to bioinformatics analysis. The subcellsular localization of TEAD1 transcripts was determined based on indirect immunofluorescence. The effects of TEAD1 transcripts overexpression on the proliferation of ICP1 cells were examined by RT-qPCR, CCK-8, and EdU assays; the effects of TEAD1 transcripts on ICP1 cells migration were examined based on the scratch test; and the effects of TEAD1 transcripts overexpression on ICP1 cells apoptosis were analyzed using apoptosis-Hoechst staining and RT-qPCR. The expression of TEAD1 transcripts in different tissues, cells lines, and ICP1 at different periods of differentiation was analyzed by RT-qPCR. The effects of TEAD1 transcripts overexpression on lipid droplet accumulation and adipogenic-related gene expression in ICP1 cells were analyzed based on Oil Red O and BODIPY staining, RT-qPCR, Western blot, and dual-luciferase reporter gene assays. Finally, the content of triglyceride (TG) was measured in TEAD1 overexpressed ICP1 cells. ResultsThe full-length TEAD1 was cloned and two TEAD1 transcripts were identified. The TEAD1-V1 protein was found to be localized primarily in the cell nucleus, whereas the TEAD1-V2 protein is localized in the cell cytoplasm and nucleus. The overexpression of both TEAD1-V1 and TEAD1-V2 significantly inhibited the proliferation of ICP1 cells. Whereas the overexpression of TEAD1-V1 promoted ICP1 cell migration, the overexpression of TEAD1-V2 had no significant effects on ICP1 migration; the overexpression of both TEAD1-V1 and TEAD1-V2 significantly promoted the apoptosis of ICP1 cells. We found that the different transcripts of TEAD1 have similar expression pattern in different tissues and cells lines. During induced preadipocyte differentiation, the expression of these genes initially declined, although subsequently increased. Overexpression of TEAD1-V1 promoted a significant reduction in lipid droplet formation and inhibited C/EBPα expression during the differentiation of ICP1 cells (P<0.05). However, the overexpression of TEAD1-V2 had no significant effect on lipid droplet accumulation or the expression of adipogenic-related proteins (P>0.05). Overexpression of TEAD1-V1 significantly decreased triglyceride content in ICP1 cells (P<0.05), while overexpression of TEAD1-V2 had no effect on triglyceride content in ICP1 cells (P>0.05). ConclusionIn this study, for the first time, identified two TEAD1 transcripts. Overexpressed transcripts TEAD1-V1 and TEAD1-V2 both inhibited the proliferation of chicken preadipocytes and promoted apoptosis of chicken preadipocytes. TEAD1-V1 inhibited the differentiation of preadipocytes and promoted the migration of preadipocytes, while TEAD1-V2 had no effect on the differentiation and migration of preadipocytes.

Background: Twenty-four-hour urinary free cortisol (UFC) measurement is the initial diagnostic test for Cushing’s syndrome (CS). We compared UFC determination by both direct and extraction immunoassays using Abbott Architect, Siemens Atellica Solution, and Beckman DxI800 with liquid chromatography-tandem mass spectrometry (LC-MS/MS). In addition, we evaluated the value of 24-hr UFC measured by six methods for diagnosing CS. Methods: Residual 24-hr urine samples of 94 CS and 246 non-CS patients were collected.A laboratory-developed LC-MS/MS method was used as reference. UFC was measured by direct assays (D) using Abbott, Siemens, and Beckman platforms and by extraction assays (E) using Siemens and Beckman platforms. Method was compared using Passing–Bablok regression and Bland–Altman plot analyses. Cut-off values for the six assays and corresponding sensitivities and specificities were calculated by ROC analysis. Results: Abbott-D, Beckman-E, Siemens-E, and Siemens-D showed strong correlations with LC-MS/MS (Spearman coefficient r = 0.965, 0.922, 0.922, and 0.897, respectively), while Beckman-D showed weaker correlation (r = 0.755). All immunoassays showed proportionally positive bias. The areas under the curve were 0.975 for Abbott-D, 0.972 for LCMS/MS, 0.966 for Siemens-E, 0.948 for Siemens-D, 0.955 for Beckman-E, and 0.877 for Beckman-D. The cut-off values varied significantly (154.8–1,321.5 nmol/24 hrs). Assay sensitivity and specificity ranged from 76.1% to 93.2% and from 93.0% to 97.1%, respectively. Conclusions Commercially available immunoassays for measuring UFC show different levels of analytical consistency compared to LC-MS/MS. Abbott-D, Siemens-E, and Beckman-E have high diagnostic accuracy for CS.

Endodontic diseases are a kind of chronic infectious oral disease.Common endodontic treatment concepts are based on the removal of inflamed or necrotic pulp tissue and the replacement by gutta-percha.However,it is very essential for endodontic treatment to debride the root canal system and prevent the root canal system from bacterial reinfection after root canal therapy(RCT).Recent research,encompassing bacterial etiology and advanced imaging techniques,contributes to our understanding of the root canal system's anatomy intricacies and the technique sensitivity of RCT.Success in RCT hinges on factors like patients,infection severity,root canal anatomy,and treatment techniques.Therefore,improving disease management is a key issue to combat endodontic diseases and cure periapical lesions.The clinical difficulty assessment system of RCT is established based on patient conditions,tooth conditions,root canal configuration,and root canal needing retreatment,and emphasizes pre-treatment risk assessment for optimal outcomes.The findings suggest that the presence of risk factors may correlate with the challenge of achieving the high standard required for RCT.These insights contribute not only to improve education but also aid practitioners in treatment planning and referral decision-making within the field of endodontics.

Objective:To analyze the effectiveness and interobserver agreement of the Parer five-tier, the National Institute of Child Health and Human Development (NICHD) three-tier, and the International Federation of Gynecology and Obstetrics (FIGO) three-tier electronic fetal monitoring (EFM) systems in identification of neonatal acidosis during labor.Methods:This retrospective study was conducted on full-term singleton cephalic deliveries with neonatal acidosis (umbilical artery blood gas pH≤7.1) and normal newborns (umbilical artery blood gas pH≥7.2) in the Nanjing Drum Tower Hospital, Nanjing University Medical School from January to December 2020. EFM tracings during the last 30-60 min before delivery were collected. Four obstetricians independently described the features of randomly sorted and coded EFM tracings. Another obstetrician categorized these tracings using the NICHD three-tier, FIGO three-tier, and Parer five-tier evaluation systems based on the features. All researchers were masked to the clinical characteristics and maternal and neonatal outcomes. The sensitivity and specificity for identifying neonatal acidosis, as well as the interobserver agreement, were analyzed for all three systems. Independent sample t-test, Chi-square (or Fisher's exact test) and Mann-Whitney U tests were used for statistical analysis. Inter-group comparisons of sensitivity and specificity between the three evaluation systems were assessed using McNemar's test. The Kappa statistic was used to analyze interobserver agreement. Results:This study included a total of 3 558 cases. After propensity score matching, there were 44 cases of neonatal acidosis and 78 control cases. There were no significant differences in parity, gestational weeks, modes of delivery, placental abruption, or analgesia rates between the two groups. The rates of instrumental vaginal delivery and neonatal intensive care unit (NICU) admission in the acidosis group were significantly higher than those in the control group [15.8% (7/44) vs. 2.6% (2/78), χ2=8.45, P=0.003; 31.8% (14/44) vs. 12.8% (10/78), χ2=8.45, P=0.004], while the umbilical artery blood pH and mean base excess were lower in the acidosis group than in the control group [7.04±0.07 vs. 7.30±0.05, t=4.98; (－12.40±3.32) vs. (－5.64±1.95) mmol/L, t=13.61; both P<0.001]. (2) Using the NICHD three-tier system, 95.5% (42/44) of the acidosis cases and 89.7% (70/78) of the control cases were classified as having category Ⅱ EFM tracings, indicating potential fetal acid-base imbalance; category Ⅲ EFM tracings were only observed in 4.5% (2/44) of the cases in the acidosis group. With the FIGO three-tier system, 81.8% (36/44) of the acidosis cases were categorized as having "pathological" tracings, and with the Parer five-tier system, 86.4% (38/44) of the acidosis cases were correctly classified into the "orange or red" risk zones that indicated acid-base imbalance. Among the control cases, there were 28.2% (22/78) with EFM tracings of "normal patterns" categorized by the FIGO three-tier system, and 41.0% (32/78) classified into the "green or blue" risk zones by the Parer five-tier system, which indicated good fetal conditions. None of the acidosis cases were misdiagnosed as being normal by the Parer five-tier system. (3) Compared with the NICHD three-tier system, both the FIGO three-tier and the Parer five-tier systems showed increased diagnostic sensitivity [4.5% (1.2%- 14.5%) vs. 81.8% (66.8%-89.4%) and 86.4% (71.8%-92.4%)], but decreased specificity [100.0% (95.3%- 100.0%) vs. 87.2% (78.0%-92.9%) and 84.6% (75.0%-91.0%)]. There was no statistically significant difference in the sensitivity or specificity between the FIGO three-tier and Parer five-tier systems for identifying neonatal acidosis ( P=0.727 and 0.791). (4) When reading the tracings of control cases, the total agreement rate for the NICHD three-tier system by different observers was as high as 94.2%, while the total agreement rates for the FIGO three-tier and Parer five-tier systems were 69.7% and 67.7%, respectively. In the interpretation of EFHR tracings for acidosis cases, the interobserver agreement for the Parer five-tier system was excellent [Kappa (95% CI): 0.87 (0.79-0.95)], while both the NICHD three-tier and FIGO three-tier systems showed good agreement [Kappa (95% CI): 0.77 (0.66-0.88) and 0.72 (0.60-0.84)]. Conclusions:The Parer five-tier and the FIGO three-tier systems have higher sensitivity in identifying neonatal acidosis than the NICHD three-tier system, and the Parer five-tier system achieves a higher negative predictive value and a greater agreement in the interpretation of pathological EFM patterns.

Objective:To retrospectively analyze the treatment status of tyrosine kinase inhibitors (TKI) in newly diagnosed patients with chronic myeloid leukemia (CML) in China.Methods:Data of chronic phase (CP) and accelerated phase (AP) CML patients diagnosed from January 2006 to December 2022 from 77 centers, ≥18 years old, and receiving initial imatinib, nilotinib, dasatinib or flumatinib-therapy within 6 months after diagnosis in China with complete data were retrospectively interrogated. The choice of initial TKI, current TKI medications, treatment switch and reasons, treatment responses and outcomes as well as the variables associated with them were analyzed.Results:6 893 patients in CP ( n=6 453, 93.6%) or AP ( n=440, 6.4%) receiving initial imatinib ( n=4 906, 71.2%), nilotinib ( n=1 157, 16.8%), dasatinib ( n=298, 4.3%) or flumatinib ( n=532, 7.2%) -therapy. With the median follow-up of 43 ( IQR 22-75) months, 1 581 (22.9%) patients switched TKI due to resistance ( n=1 055, 15.3%), intolerance ( n=248, 3.6%), pursuit of better efficacy ( n=168, 2.4%), economic or other reasons ( n=110, 1.6%). The frequency of switching TKI in AP patients was significantly-higher than that in CP patients (44.1% vs 21.5%, P<0.001), and more AP patients switched TKI due to resistance than CP patients (75.3% vs 66.1%, P=0.011). Multi-variable analyses showed that male, lower HGB concentration and ELTS intermediate/high-risk cohort were associated with lower cytogenetic and molecular responses rate and poor outcomes in CP patients; higher WBC count and initial the second-generation TKI treatment, the higher response rates; Ph + ACA at diagnosis, poor PFS. However, Sokal intermediate/high-risk cohort was only significantly-associated with lower CCyR and MMR rates and the poor PFS. Lower HGB concentration and larger spleen size were significantly-associated with the lower cytogenetic and molecular response rates in AP patients; initial the second-generation TKI treatment, the higher treatment response rates; lower PLT count, higher blasts and Ph + ACA, poorer TFS; Ph + ACA, poorer OS. Conclusion:At present, the vast majority of newly-diagnosed CML-CP or AP patients could benefit from TKI treatment in the long term with the good treatment responses and survival outcomes.

Objective:To investigate the efficacy and safety of protein A immunoadsorption (PAIA) combined with rituximab (RTX) in highly sensitized patients who underwent haplo-hematopoietic stem cell transplantation (haplo-HSCT) .Methods:The clinical data of 56 highly sensitized patients treated with PAIA and RTX before haplo-HSCT at the First Affiliated Hospital of Soochow University and Soochow Hopes Hematonosis Hospital between March 2021 and June 2023 were retrospectively analyzed. The number of human leukocyte antigen (HLA) antibody types and the mean fluorescence intensity (MFI), humoral immunity, adverse reactions during adsorption, and survival within 100 days before and after adsorption were measured.Results:After receiving the PAIA treatment, the median MFI of patients containing only HLA Ⅰ antibodies decreased from 7 859 (3 209-12 444) to 3 719 (0-8 275) ( P<0.001), and the median MFI of HLA Ⅰ+Ⅱ antibodies decreased from 5 476 (1 977-12 382) to 3 714 (0-11 074) ( P=0.035). The median MFI of patients with positive anti-donor-specific antibodies decreased from 8 779 (2 697-18 659) to 4 524 (0–15 989) ( P<0.001). The number of HLA-A, B, C, DR, and DQ antibodies in all patients decreased after the PAIA treatment, and the differences were statistically significant (A, B, C, DR: P<0.001, DQ: P<0.01). The humoral immune monitoring before and after the PAIA treatment showed a significant decrease in the number of IgG and complement C3 ( P<0.001 and P=0.002, respectively). Forty-four patients underwent HLA antibody monitoring after transplantation, and the overall MFI and number of antibody types decreased. However, five patients developed new antibodies with low MFI, and nine patients continued to have high MFI. The overall survival, disease-free survival, non-recurrent mortality, and cumulative recurrence rates at 100 days post-transplantation were 83.8%, 80.2%, 16.1%, and 4.5%, respectively. Conclusions:The combination of PAIA and RTX has a certain therapeutic effect and good safety in the desensitization treatment of highly sensitive patients before haplo-HSCT.

Objective: To analyze and compare therapy responses, outcomes, and incidence of severe hematologic adverse events of flumatinib and imatinib in patients newly diagnosed with chronic phase chronic myeloid leukemia (CML) . Methods: Data of patients with chronic phase CML diagnosed between January 2006 and November 2022 from 76 centers, aged ≥18 years, and received initial flumatinib or imatinib therapy within 6 months after diagnosis in China were retrospectively interrogated. Propensity score matching (PSM) analysis was performed to reduce the bias of the initial TKI selection, and the therapy responses and outcomes of patients receiving initial flumatinib or imatinib therapy were compared. Results: A total of 4 833 adult patients with CML receiving initial imatinib (n=4 380) or flumatinib (n=453) therapy were included in the study. In the imatinib cohort, the median follow-up time was 54 [interquartile range (IQR), 31-85] months, and the 7-year cumulative incidences of CCyR, MMR, MR(4), and MR(4.5) were 95.2%, 88.4%, 78.3%, and 63.0%, respectively. The 7-year FFS, PFS, and OS rates were 71.8%, 93.0%, and 96.9%, respectively. With the median follow-up of 18 (IQR, 13-25) months in the flumatinib cohort, the 2-year cumulative incidences of CCyR, MMR, MR(4), and MR(4.5) were 95.4%, 86.5%, 58.4%, and 46.6%, respectively. The 2-year FFS, PFS, and OS rates were 80.1%, 95.0%, and 99.5%, respectively. The PSM analysis indicated that patients receiving initial flumatinib therapy had significantly higher cumulative incidences of CCyR, MMR, MR(4), and MR(4.5) and higher probabilities of FFS than those receiving the initial imatinib therapy (all P<0.001), whereas the PFS (P=0.230) and OS (P=0.268) were comparable between the two cohorts. The incidence of severe hematologic adverse events (grade≥Ⅲ) was comparable in the two cohorts. Conclusion: Patients receiving initial flumatinib therapy had higher cumulative incidences of therapy responses and higher probability of FFS than those receiving initial imatinib therapy, whereas the incidence of severe hematologic adverse events was comparable between the two cohorts.
Adult ; Humans ; Adolescent ; Imatinib Mesylate/adverse effects* ; Incidence ; Antineoplastic Agents/adverse effects* ; Retrospective Studies ; Pyrimidines/adverse effects* ; Leukemia, Myelogenous, Chronic, BCR-ABL Positive/drug therapy* ; Treatment Outcome ; Benzamides/adverse effects* ; Leukemia, Myeloid, Chronic-Phase/drug therapy* ; Aminopyridines/therapeutic use* ; Protein Kinase Inhibitors/therapeutic use*

Objective:To evaluate the prevalence and distribution of ossification of ligamentum flavum (OLF) at the segments adjacent to the apex in patients with degenerative kyphosis.Methods:All of 74 patients with degenerative kyphosis from January 2018 to December 2021 were retrospective reviewed. All patients were taken anteroposterior and lateral radiographs, CT scan and magnetic resonance imaging (MRI) of the entire spine. Global kyphosis, the morphology of kyphosis and the occurrence of OLF at three segments adjacent to the kyphosis apex were recorded.Results:Of the 74 patients, 54 patients (73%) developed OLF in three segments adjacent to the kyphotic apex. The mean age of the 54 patients was 61.4±6.8 years, and the mean global kyphosis was 49.5°±21.2°. Among other 20 patients without OLF, the mean age was 56.1±7.5 years, and the mean kyphosis angle was 52.1°±19.1°. There was a statistically significant difference in ages ( t=2.92, P=0.005), but no statistically significant difference was observed regarding global kyphosis ( t=0.48, P=0.634). In these 74 patients, 9 patients had angular kyphosis, of which 8 (89%) developed OLF; of the 65 patients without angular kyphosis, 46 patients (71%) developed OLF. There was no significant difference between them (χ 2=1.32, P=0.251). Among the 54 patients diagnosed with OLF, 5 patients (9%) suffered ossification of the posterior longitudinal ligament (OPLL) and 20 patients (37%) suffered dural ossification; 43 patients (80%) developed OLF at proximal segments of apex, 6 patient (11%) developed OLF at distal segments of apex, and 5 patients (9%) developed OLF both at proximal and distal segments of apex. Thirty-two patients (59%) developed OLF at the first segment adjacent to the kyphotic apex, 27 patients (50%) developed OLF at the second segment, and 15 patients (28%) developed OLF at the third segment. Conclusion:Among patients with degenerative kyphosis, about 73% may development OLF within three segments adjacent to the kyphotic apex, and it mostly occurred within two segments adjacent to the apex proximally.