Zhejiang da xue xue bao. Yi xue ban = Journal of Zhejiang University. Medical sciences最新文献

Objectives: To explore the diagnostic value of ultrasonography for injuries of anterior talofibular ligament (ATFL) and anterior inferior tibiofibular ligament distal fascicle (ATiFL-DF) in patients with ankle fractures.

Methods: Clinical data of 51 patients with ankle fractures who were clinically suspected of ligament injuries and underwent ankle ultrasonography examination and arthroscopy in Sir Run Run Shaw Hospital, Zhejiang University School of Medicine from April 2019 to March 2023 were retrospectively analyzed. Using arthroscopic results as the gold standard, the sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of ultrasonography in diagnosing ATFL and ATiFL-DF injuries were evaluated, and Kappa consistency test was performed.

Results: The sensitivity and specificity of ultrasonography in diagnosis of ATFL injury were 100.0% and 92.3%, with the PPV of 92.6% and NPV of 100.0%. Ultrasonography findings exhibited excellent concordance with arthroscopic results (kappa=0.849). The sensitivity and specificity of ultrasonography in diagnosis of ATiFL-DF injury was 86.7% and 33.3%, with the PPV of 90.7% and NPV of 25.0%. However, the consistency between ultrasonography and arthroscopic results was poor (kappa=0.168).

Conclusions: Ultrasonography is reliable in assessing injuries of ATFL in patients with ankle fractures, but its specificity in diagnosing ATiFL-DF is poor. Therefore, ankle arthroscopy remains necessary for ankle fracture patients with negative findings of ATiFL-DF in ultrasonography.

Basivertebral foramen is a natural orifice in the posterior wall of the vertebral body existing in humans and mammals, through which the basal vertebral vein, branch of lumbar artery and recurrent branch of spinal nerve enter and exit the vertebral body. Basivertebral foramen changes the local microstructure of the vertebral body, resulting in cortical defect and sparse trabecular bone in the central region of the vertebral body, thus affecting its biomechanical characteristics and making its central region a "weak" area of the vertebra. Some characteristic injuries of the vertebra are related to basivertebral foramen, such as vertebral compression fracture and intervertebral cleft, vertebral burst fracture and posterior upper vertebral fracture fragment, and cement leakage during treatment. In this article, the anatomical and developmental biological characteristics of basivertebral foramen, the impact of basivertebral foramen on biomechanical characteristics, and the treatment of basivertebral foramen related vertebral diseases are reviewed, in order to provide references for the clinical diagnosis and treatment of vertebral injuries.

Objectives: To investigate the effect of sinusoidal alternating electromagnetic field (SEMF) on fracture healing and its mechanism.

Methods: Femoral fracture model was established using specific pathogen free male Wistar rats. Thirty rats were randomly divided into the control and SEMF groups with 15 rats in each group. The SEMF group was given 50 Hz 1.8 mT for 90 min every day, while the control group was not treated. X-ray examinations were performed every two weeks to determine the formation of bone scabs. Three rats from both groups were sacrificed after 2 and 4 weeks of treatment. Protein was extracted from the fractured femurs, and the expression of type Ⅰ collagen (COL-1), osterix (OSX), Runt-related transcription factor 2 (RUNX2), and vascular endothelial growth factor (VEGF) was detected by Western blotting. After 8 weeks, the femur on the operated side was taken for micro-CT scanning to observe fracture healing, angiography to observe blood vessel growth, and organs such as hearts, livers, spleens, lungs, and kidneys were taken for safety evaluation by hematoxylin-eosin staining (HE staining).

Results: The bone scab scores of the SEMF group were significantly higher than those of the control group after 2, 4, 6, and 8 weeks of treatment (all P<0.01). The fracture healing of the SEMF group was better than that of the control group after 8 weeks, and the bone volume scores of the two groups were 0.243±0.012 and 0.186±0.008, respectively (P<0.01); the number of blood vessels in the SEMF group was also more than that of the control group after 8 weeks. Western blotting results showed that the expressions of COL-1, OSX, RUNX2, and VEGF were higher in the SEMF group than those in the control group after 2 and 4 weeks of treatment (all P<0.05). HE staining showed that histopathological results of the examined organs were normal in both groups.

Conclusions: SEMF can accelerate fracture healing by promoting the expression of osteogenic factors and vascular proliferation without significant adverse effects.

Objectives: To investigate the expression of signal recognition particle 14 (SRP14) in hepatocellular carcinoma (HCC) and its clinical significance.

Methods: The data of SRP14 expression in HCC were obtained from bioinformatics study, and from investigation with quantitative reverse transcription polymerase chain reaction (qRT-PCR), immunohistochemical staining and Western blotting in clinical samples. The Kaplan-Meier analysis was used to determine the associations between SRP14 mRNA expression and the overall survival, progression-free survival, and disease-specific survival of HCC patients. The effect of SRP14 on the proliferation and migration of HCC cells were determined by EdU staining, MTS, Transwell and wound-healing assays. The potential mechanism for SRP14 regulating HCC was explored through Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) enrichment analysis as well as qRT-PCR.

Results: According to the data from GSE14520, TNMplot database and clinical samples, compared with paired tumor-adjacent tissues, non-paired tumor-adjacent tissues and normal tissues, the mRNA expression of SPR14 in HCC tissues was upregulated (all P<0.05). In clinical samples, compared with paired tumor-adjacent tissues, the protein expression of SPR14 in HCC tissues was increased (P<0.05). The increased mRNA expression of SRP14 was associated with good overall survival, progression-free survival, and disease-specific survival in HCC patients. SRP14 inhibited the proliferation and migration of HCC cells in vitro. According to the KEGG and GO enrichment analysis, in non-specific HCC, the genes co-expressed with SRP14 may predominantly regulate protein synthesis, processing, and transport, while in nonalcoholic fatty liver disease related HCC, the genes co-expressed with SRP14 could control multiple signaling pathways such as MAPK, cAMP, PI3K-Akt, and Wnt. Mechanistically, SRP14 up-regulated the mRNA expression of tumor suppressor gene GPRC5A inHCC cells (P<0.05).

Conclusions: SRP14 may regulate HCC progression and influence patient prognosis.

Objectives: To compare the clinical outcomes of anterolateral femoral interregional flap with turbocharge technique and traditional anterolateral femoral flap in repair of large limb wounds.

Methods: Clinical data of 38 patients with large limb surface wound (11 cm×39 cm-16 cm×65 cm) admitted to the Sir Run Run Shaw Hospital, Zhejiang University School of Medicine from May 2018 to May 2022 were retrospectively analyzed. Eighteen patients were treated by anterolateral thigh perforator flap combined with superficial circumflex iliac artery flap (ALTP-SCIAP) with turbocharge technique (interregional flap group); while 20 patients were treated with unilateral or bilateral anterolateral femoral flaps, combined with skin grafting if necessary (traditional anterolateral femoral flap group). The survival of skin flap, repair of donor area, complications and patient satisfaction were compared between the two groups.

Results: In interregional flap group, 18 flaps were harvested and transplanted, the flap width, length and the viable area were (9.9±2.0) cm, (44.2±3.5) cm and (343.2±79.9) cm², respectively. In traditional anterolateral femoral flap group, 29 flaps were harvested and transplanted, the flap width, length and the viable area were (11.0±2.8) cm, (21.7±3.2) cm and (186.4±49.2) cm², respectively. There were significant differences in the flap length and the viable area between the two groups (t=22.365 and 8.345, both P<0.05). In the interregional flap group, the donor site of flap was closed by direct suture in 11 flaps, by skin retractor assisted suture in 6 flaps, and by skin grafting in one flap. In traditional anterolateral femoral flap group, the donor site of flap was closed by direct suture in 12 flaps, by skin retractor assisted suture in 11 flaps, and by skin grafting in 6 flaps. The skin graft rates of the two groups were 5.6% (1/18) and 20.7% (6/29), respectively (χ²=2.007, P>0.05). The interregional flap group had lower postoperative complications rate (5.6% vs. 35.0%, χ²=4.942, P<0.05) and higher patient satisfaction rate (94.4% vs. 70.0%, χ²=4.448, P<0.05) than traditional anterolateral femoral flap group.

Conclusions: Compared with the traditional anterolateral femoral flap, the anterolateral femoral interregional flap with turbocharge technique has a larger flap area, most of the donor areas of the flap can be sutured directly without skin grafting and with less complications and a higher patient satisfaction rate.

Objectives: To investigate the association of R-loop binding proteins with prognosis and chemotherapy efficacy in lung adenocarcinoma.

Methods: The data related to R-loop regulatory genes were obtained from literature of R-loop proteomics and relevant databases. We used 403 cases of lung adenocarcinoma in the Cancer Genome Atlas as training set, and two datasets GSE14814 and GSE31210 in Gene Expression Omnibus as validation sets. The weighted gene co-expression network analysis (WGCNA) was employed to identify R-loop genes with a significant impact on the clinical phenotype of lung adenocarcinoma. Least absolute shrinkage and selection operator (LASSO) regression analysis was utilized to eliminate genes exhibiting multicollinearity. A multivariate Cox regression analysis was employed to scrutinize clinical variables and R-loop characteristic genes that exert independent prognostic effects on patient survival. Subsequently, a risk score model was constructed. The predictive capacity of this model for the prognosis of patients was analyzed and validated. Additionally, the performance of risk model on the anti-tumor drug sensitivity was assessed. The mutations of R-loop genes were analyzed by maftools. The effect of PLEC expression on anti-tumor drug sensitivity was tested on non-small cell lung adenocarcinoma H1299 and A549 cells in vitro.

Results: A collection of 1551 R-loop genes were obtained, and 78 genes exhibited significant effects on the clinical phenotype shown on WGCNA. The LASSO regression analysis retained fourteen R-loop genes. A multivariate Cox regression analysis further identified three R-loop genes (HEXIM1, GLI2, PLEC) and a clinical variable (tumor grading) that were associated with patient prognosis. Risk prediction model was established according to the regression coefficients of each parameter. Kaplan-Meier survival analysis showed that the prognosis of high-risk group was significantly worse than that of low-risk group (P<0.01). The time-dependent ROC curve showed that the risk model had good predictive ability in both training and validation sets. Predictive analyses of anti-neoplastic drug sensitivity indicated a diminished responsiveness to both chemotherapy and targeted treatment drugs among high-risk patients. The expression of PLEC was strongly correlated with sensitivity to gefitinib, a classical EGFR inhibitor.

Conclusions: R-loop binding proteins have been identified as significant determinants in the prognosis and therapeutic strategies for lung adenocarcinoma, which indicates that therapeutic interventions targeting these specific R-loop binding proteins might contribute to a better survival of the patients.

Objectives: To investigate the effect of decarbromodiphenyl ether (BDE-209) exposure on the migration ability of triple-negative breast cancer (TNBC) cells and to explore the underlying mechanism.

Methods: Human TNBC MDA-MB-231 cells were divided into blank control group and BDE-209 exposure groups (treated with 0.02, 0.20, 2.00, 20.00 and 200.00 ng/mL BDE-209 in high glucose DMEM). Extracellular vehicles (EVs) secreted by MDA-MB-231 cells were isolated by differential ultracentrifugation. Transmission electron microscopy (SEM), nanoparticle tracking analysis (NTA) and Western blotting were performed to characterize the EVs. The effect of the EVs induced by BDE-209 exposure (EVs-BDE-209) on the migration and invasion of MDA-MB-231 cells was detected by wound-healing assay and Transwell test. qRT-PCR was used to measure the miR-221 level in EVs-BDE-209. The expression of MMP9 in MDA-MB-231 cells was determined by Western blotting.

Results: Compared with the blank control, BDE-209 exposure increased the tumor cell-derived EVs in dose-dependent manner. The MDA-MB-231 cells co-cultured with EVs released by 200.00 ng/mL BDE-209 exposure showed an 86% increase in cell migration rate, a 1.32-fold higher number of membrane-penetrating cells, a 2.71-fold higher expression level of miR-221, and a 1.62-fold higher expression level of MMP9 compared with the blank control group (all P<0.05). While transfection with anti-miR-221 antibody to decrease miR-221 level in EVs significantly reversed the increased invasion ability of the MDA-MB-231 cells treated with EVs-BDE-209.

Conclusions: BDE-209 exposure may promote metastasis potential of MDA-MB-231 cells via EVs-BDE-209 transmitted miR-221.

Objectives: To establish a methodological system for reprogramming rat embryonic fibroblasts (REF) into chemically induced neurons (ciNCs) via small molecule compounds to provide safe and effective donor cells for treatment of neurodegenerative diseases.

Methods: Based on the method established by PEI Gang's research group to directly reprogram human fibroblasts into neurons, the induction medium and maturation medium was optimized by replacing the coating solution, mitigating oxidative stress injury, adding neurogenic protective factors, adjusting the concentration of trichothecenes, performing small-molecule removal experiments, and carrying out immunofluorescence and Western blotting on cells at different stages of induction to validate the effect of induction.

Results: When the original protocol was used for induction, the cell survival rate was (34.24±2.77)%. After replacing the coating solution gelatin with matrigel, the cell survival rate increased to (45.41±4.27)%; after adding melatonin, the cell survival rate increased to (67.95±5.61)% and (23.43±1.42)% were transformed into neural-like cells; after adding the small molecule P7C3-A20, the cell survival rate was further increased to (76.27±1.41)%, and (39.72±4.75)% of the cells were transformed into neural-like cells. When the concentration of trichothecene was increased to 30 μmol/L, the proportion of neural-like cells reached (55.79±1.90)%; after the removal of SP600125, (86.96±2.15)% of the cells survived, and the rate of neural-like cell production increased to (63.43±1.60)%. With the optimized protocol, REF could be successfully induced into ciNC through the neural precursor cell stage, in which the neural precursor cells were able to highly express the neural precursor cell markers SRY-related HMG-box gene 2 (Sox2) and paired box 6 (Pax6) as well as neuron-specific marker tubulin 1 (Tuj1), while the expression of fiber-associated protein vimentin was reduced. After two weeks of induction of neural precursor cells in a maturation medium, most cells displayed neuronal-like cell morphology. The induced ciNCs were able to highly express the mature neuronal surface markers Tuj1 and microtubule-associated protein 2 (MAP2), while the expression of vimentin was reduced.

Conclusions: The small molecule combinations optimized in this study can reprogram REF to ciNCs under normoxic conditions.

Objectives: To investigate the effect of osteoblast-derived extracellular vesicles (OB-EVs) on the proliferation and differentiation of osteoclasts, and to explore the possible molecular mechanism of extracellular vesicles involved in the communication between osteoblasts and osteoclasts.

Methods: Primary osteoblasts were isolated from newborn mouse calvarial bone and induced by β-glycero phosphate, ascorbic acid and dexamethasone. Osteogenic feature was tested by alkaline phosphatase (ALP) and alizarin red S staining. Extracellular vesicles were isolated by ultracentrifugation from the cell culture supernatant. Vesicle morphology was observed by transmission electron microscopy, and the characteristic markers of tumor susceptibility gene 101 (TSG101), ALG-2 interacting protein X (Alix) and cluster of differentiation 9 (CD9) on the surface of extracellular vesicles were identified by Western blotting. Cell counting kit 8 (CCK-8) assay was used to determine the proliferation effect of OB-EVs on mouse mononuclear macrophage RAW264.7 cells. Furthermore, the expression level of specific markers of osteoclast differentiation in RAW264.7 cells was detected by Western blotting after the combined effect of OB-EVs and receptor activator for nuclear factor κB ligand (RANKL). The number of osteoclasts was observed and compared with OB-EVs-treated mouse bone marrow-derived macrophages (BMMs) by tartrate-resistant acid phosphatase (TRAP) staining, and the effect of OB-EVs on osteoclast differentiation was determined.

Results: The extracted OB-EVs showed a double-layer cup-like structure with a diameter of 30-150 nm, and TSG101, Alix and CD9 were expressed. RAW264.7 cells were stimulated with OB-EVs, and the results of CCK-8 assay showed that high concentration of OB-EVs (more than 20 μg/mL) inhibited cell proliferation (P<0.05). Western blotting analysis showed that the expression of osteoclast differentiation marker proteins such as c-Fos, activated T cell nuclear factor (NFATc1) and c-Jun N-terminal kinase (JNK) in RAW264.7 cells were significantly increased, and the promoting effect was enhanced with increasing of OB-EVs concentration (P<0.05). In addition, the combination of OB-EVs and RANKL on BMMs showed that the number of TRAP-positive cells was significantly higher than that of the RANKL induction group alone (P<0.05).

Conclusions: OB-EVs can promote the differentiation of osteoclast precursor cells into osteoclasts, but high concentration of OB-EVs can inhibit proliferation of RAW264.7 cells.

Segmental bone defects and nonunion of fractures caused by trauma, infection, tumor or systemic diseases with limited osteogenesis and prolonged bone healing cycles are challenging issues in orthopedic clinical practice. Therefore, identifying regulatory factors for bone tissue regeneration and metabolism is crucial for accelerating bone repair and reconstructing defective areas. Silence information regulator 6 (SIRT6), functioning as a deacetylase and nucleotide transferase, is extensively involved in the regulation of differentiation, apoptosis, metabolism, and inflammation in bone cells including osteoblasts and osteoclasts, and is considered to be an important factor in regulating bone metabolism. SIRT6 forms a complex with B lymphocyte-induced maturation protein 1 (Blimp1), down-regulates the expression of the nuclear factor κB (NF-κB) pathway, and promotes the expression of the ERα-FasL axis signal to inhibit osteoclast formation and maturation differentiation, thereby hindering bone resorption and increasing bone mass. In addition, SIRT6 activates the Akt-mTOR pathway to regulate the autophagy level and osteogenesis of bone marrow mesenchymal stem cells, inhibits glycolysis and reactive oxygen production in osteoblasts, promotes osteoblast differentiation through the CREB/CCN1/COX2 pathway and the bone morphogenetic protein (BMP) signaling pathway, enhances bone formation, and accelerates bone regeneration and repair of skeletal tissue. This article provides an overview of the research progress on SIRT6 in the pathophysiology of bone regeneration, revealing its potential as a novel therapeutic target for bone tissue repair to alleviate the progression of skeletal pathological diseases.