Objective: To assess the feasibility of nuclear score combined with cyclin D1 immunocytochemistry in classifying indeterminate thyroid nodules with fine-needle aspiration (FNA) cytological diagnosis of Bethesda category Ⅲ-Ⅴ. Methods: A consecutive cohort of 118 thyroid FNA specimens with indeterminate diagnosis (TBSRTC category Ⅲ-Ⅴ) and available histopathologic follow-up data were collected between December 2018 and April 2022 at the Department of Pathology, Beijing Hospital, China. These cases were subjected to cytological evaluation and cyclin D1 immunocytochemistry. The optimal cut-off points of a simplified nuclear score and the percentage of cyclin D1-positive cells for the diagnosis of malignancy or low-risk neoplasm were determined using the receiver operating characteristic (ROC) curves and area under the ROC curve (AUC). The specificity, sensitivity, positive predictive value (PPV) and negative predictive value (NPV) of nuclear score and cyclin D1 immunostaining were evaluated from the crosstabs based on cut-off points. The diagnostic accuracy of simplified nuclear score combined with cyclin D1 immunostaining was estimated using ROC curve analysis. Results: Nuclear grooves, intra-nuclear inclusions and chromatin clearing were more commonly found in malignancy/low-risk neoplasms than benign lesions (P=0.001, P=0.012 and P=0.001 respectively). A cut-off point of≥2 for the simplified nuclear score was sensitive for defining malignancy/low-risk neoplasm, and its PPV, NPV, sensitivity and specificity were 93.6%, 87.5%, 99.0% and 50.0% respectively. A positive cut-off point of 10% positive thyroid cells in cyclin D1 immunostaining demonstrated sensitivity of 88.5%, specificity of 100%, PPV of 100% and NPV of 53.8% for correctly detecting thyroid malignancy or low-risk neoplasm. The sensitivity and PPV of simplified nuclear score combined with cyclin D1 immunostaining were 93.3% and 100%, respectively. Both specificity and NPV were maintained at high levels (100% and 66.7%, respectively). The diagnostic accuracy of simplified nuclear score combined with cyclin D1 immunostaining in detecting thyroid malignancy/low-risk neoplasm was increased to 94.1% compared to using either of them alone. Conclusions: Combing simplified nuclear score and cyclin D1 immunostaining on FNA cytology specimens can increase the diagnostic accuracy in classifying thyroid nodules of indeterminate cytological categories. Thus, this supplementary approach provides a simple, accurate, and convenient diagnostic method for cytopathologists so that may reduce unnecessary thyroidectomies.
Humans ; Thyroid Nodule/pathology* ; Biopsy, Fine-Needle ; Cyclin D1 ; Immunohistochemistry ; Thyroid Neoplasms/pathology* ; Retrospective Studies

OBJECTIVE: To observe the effect of needle-knife on the chondrocyte apoptosis of knee joint in rabbits with knee osteoarthritis (KOA) based on the CircSERPINE2-miR-1271-5P-E26 specific transformation-related gene (ERG) axis, and to explore the mechanism of needle-knife for KOA. METHODS: Thirty-six New Zealand white rabbits were randomly divided into a normal group, a model group, a needle-knife group and a sham needle-knife group, 9 rabbits in each group. The rabbits in the model group, the needle-knife group and the sham needle-knife group were treated with modified Videman method to prepare KOA model. After successful modeling, the rabbits in the needle-knife group were treated with needle-knife at cord adhesion and nodules near quadriceps femoris tendon and internal and external collateral ligament on the affected knee joint; the rabbits in the sham needle-knife group were treated with sham needle-knife baside the needle insertion point of the needle-knife group (needle-knife was only inserted, without any operation). The treatment was given once a week, 3 times in total. The Lequesne MG behavioral score was used to evaluate the knee joint damage in each group before and after intervention. After intervention, HE staining and transmission electron microscopy were used to observe the cartilage tissue morphology and ultrastructure of chondrocytes in the knee joint in each group; TUNEL method was used to detect the level of chondrocyte apoptosis in the knee joint; real-time fluorescence quantitative PCR was used to detect the expression of CircSERPINE2, miR-1271-5P and ERG mRNA in knee cartilage tissue in each group. RESULTS: After intervention, compared with the normal group, the Lequesne MG behavioral score in the model group was increased (P<0.01). Compared with the model group and the sham needle-knife group, the Lequesne MG behavioral score in the needle-knife group was decreased (P<0.01). In the model group and the sham needle-knife group, the number of chondrocytes and organelles was decreased, the cell nucleus was shrunk, mitochondria was swelling or disappeared; in the needle-knife group, the number of chondrocytes and organelles was increased, the cell nucleus was not obviously shrunk and the mitochondria was not obviously swelling. Compared with the normal group, the level of chondrocyte apoptosis in the model group was increased (P<0.01); compared with the model group and the sham needle-knife group, the level of chondrocyte apoptosis in the needle-knife group was decreased (P<0.01, P<0.05). Compared with the normal group, the expression of CircSERPINE2 and ERG mRNA in the model group was decreased (P<0.01), and the expression of miR-1271-5P mRNA was increased (P<0.01); compared with the model group and the sham needle-knife group, the expression of CircSERPINE2 and ERG mRNA in the needle-knife group was increased (P<0.01), and the expression of miR-1271-5P mRNA was decreased (P<0.01). CONCLUSION Needle-knife could reduce the knee joint damage and chondrocyte apoptosis in KOA rabbits, which may be related to up-regulating the expression of CircSERPINE2 and ERG mRNA, and inhibiting the expression of miR-1271-5P mRNA.
Rabbits ; Animals ; Osteoarthritis, Knee/metabolism* ; Chondrocytes/metabolism* ; Knee Joint/surgery* ; Apoptosis ; MicroRNAs/genetics*

OBJECTIVE: To explore the potential mechanism of resistance to axitinib in clear cell renal cell carcinoma (ccRCC), with a view to expanding the understanding of axitinib resistance, facilitating the design of more specific treatment options, and improving the treatment effectiveness and survival prognosis of patients. METHODS: By exploring the half maximum inhibitory concentration (IC₅₀) of axitinib on ccRCC cell lines 786-O and Caki-1, cell lines resistant to axitinib were constructed by repeatedly stimulated with axitinib at this concentration for 30 cycles in vitro. Cell lines that were not treated by axitinib were sensitive cell lines. The phenotypic differences of cell proliferation and apoptosis levels between drug resistant and sensitive lines were tested. Genes that might be involved in the drug resistance process were screened from the differentially expressed genes that were co-upregulated in the two drug resistant lines by transcriptome sequencing. The expression level of the target gene in the drug resistant lines was verified by real-time quantitative polymerase chain reaction (RT-qPCR) and Western blot (WB). The expression differences of the target gene in ccRCC tumor tissues and adjacent tissues were analyzed in the Gene Expression Profiling Interactive Analysis (GEPIA) public database, and the impact of the target gene on the prognosis of ccRCC patients was analyzed in the Kaplan-Meier Plotter (K-M Plotter) database. After knocking down the target gene in the drug resistant lines using RNA interference by lentivirus vector, the phenotypic differences of the cell lines were tested again. WB was used to detect the levels of apoptosis-related proteins in the different treated cell lines to find molecular pathways that might lead to drug resistance. RESULTS: Cell lines 786-O-R and Caki-1-R resistant to axitinib were successfully constructed in vitro, and their IC₅₀ were significantly higher than those of the sensitive cell lines (10.99 μmol/L, P < 0.01; 11.96 μmol/L, P < 0.01, respectively). Cell counting kit-8 (CCK-8) assay, colony formation, and 5-ethynyl-2 '-deoxyuridine (EdU) assay showed that compared with the sensitive lines, the proliferative ability of the resistant lines decreased, but apoptosis staining showed a significant decrease in the level of cell apoptosis of the resistant lines (P < 0.01). Although resistant to axitinib, the resistant lines had no obvious new replicated cells in the environment of 20 μmol/L axitinib. Nuclear protein 1 (NUPR1) gene was screened by transcriptome sequencing, and its RNA (P < 0.0001) and protein expression levels significantly increased in the resistant lines. Database analysis showed that NUPR1 was significantly overexpressed in ccRCC tumor tissue (P < 0.05); the ccRCC patients with higher expression ofNUPR1had a worse survival prognosis (P < 0.001). Apoptosis staining results showed that knockdown ofNUPR1inhibited the anti-apoptotic ability of the resistant lines to axitinib (786-O, P < 0.01; Caki-1, P < 0.05). WB results showed that knocking downNUPR1decreased the protein level of B-cell lymphoma-2 (BCL2), increased the protein level of BCL2-associated X protein (BAX), decreased the protein level of pro-caspase3, and increased the level of cleaved-caspase3 in the resistant lines after being treated with axitinib. CONCLUSION ccRCC cell lines reduce apoptosis through theNUPR1 -BAX/ BCL2 -caspase3 pathway, which is involved in the process of resistance to axitinib.
Humans ; Carcinoma, Renal Cell/metabolism* ; Axitinib/pharmacology* ; Kidney Neoplasms/metabolism* ; bcl-2-Associated X Protein ; Nuclear Proteins ; Cell Line, Tumor ; Apoptosis ; Cell Proliferation

Objective: To study the therapeutic effect of Huoxue Tongluo Decoction (HXTLD) on erectile dysfunction caused by ischemic stroke and identify the mechanisms involved. Methods: Network pharmacology was used to predict the key active ingredients and targets of HXTLD. Surgical methods were used to create a rat model of ischemic stroke. The rats were then given a suspension of HXTLD by ig administration. Erectile function was evaluated by Apomorphine (APO) induction. Real-time fluorescence quantitative reverse transcription-polymerase chain reaction (Real-time PCR) and Western blotting were used to detect the expression of related mRNAs and proteins in rat penile corpus cavernous tissue and brain tissue. Hematoxylin & Eosin (HE) staining was used to investigate structural changes in the penile cavernous tissue. Results: Network pharmacology showed that tumor necrosis factor (TNF), nitric oxide synthase 3 (eNOS), and vascular endothelial growth factor (VEGF) were the key targets of HXTLD in the treatment of erectile dysfunction caused by ischemic stroke. Experimental studies showed that HXTLD improved erectile dysfunction caused by ischemic stroke. HE results showed that HXTLD improved the structure of the corpus cavernosa. HXTLD also inhibited the expression of TNF and VEGF proteins in penile tissue (P < 0.05) and enhanced the expression of eNOS protein in penile tissue (P < 0.05). Conclusion: HXTLD improved the erectile function of rats with erectile dysfunction caused by ischemic stroke by regulating the mRNA and protein levels of TNF, eNOS and VEGF.

OBJECTIVE: To observe the effect of moxibustion on proteins related with apoptosis of hippocampal neurons in rats with vascular dementia (VD), and to explore the possible mechanism of moxibustion on improving VD. METHODS: Thirty SD rats were selected from 100 rats (3 rats were excluded) and randomly divided into a normal group and a sham operation group, 15 rats in each group. The remaining 67 rats were treated with ischemia-reperfusion method at bilateral common carotid artery to establish VD model. The 45 rats with successful VD model were randomly divided into a model group, a moxibustion group and a medication group, 15 rats in each group. On the 7th day after successful modeling, the rats in the moxibustion group were treated with suspended moxibustion at "Guanyuan" (CV 4), "Mingmen" (GV 4) and "Dazhui" (GV 14), 15 min per acupoint, once a day; there was 1 d of rest after 6 d of moxibustion, and the treatment was given for 4 weeks. The rats in the medication group was treated with nimodipine tablets by gavage, 2 mg/kg per day, 3 times a day for 4 weeks. Before and after intervention, the Morris water maze test was used to detect the escape latency of rats in each group; after the intervention, the TUNEL method was used to detect the apoptosis rate of neurons in hippocampal CA1 area; the immunofluorescence double labeling method was used to detect the number of co-expression positive cells of B-cell lymphoma-2 (Bcl-2)/neuronal nuclear antigen (NeuN) and Bcl-2-associated X protein (Bax)/NeuN in hippocampal CA1 area; the immunofluorescence single labeling method was used to detect cytochrome C (cytC) and outer mitochondrial membrane receptor Tom20 (Tom20) in hippocampal CA1 area; the Western blot method was used to detect the p53 upregulated modulator of apoptosis (PUMA) in hippocampus. RESULTS: Before intervention, compared with the normal group and the sham operation group, the escape latency in the model group, the moxibustion group and the medication group was prolonged ( CONCLUSION Moxibustion could improve the cognitive function of VD rats, which may be related to reducing the expression of Bax, cytC, Tom20 and PUMA protein in hippocampal CA1 area, promoting the release of Bcl-2 and inhibiting the apoptosis of hippocampal neurons.
Animals ; Apoptosis ; Cognition ; Dementia, Vascular/therapy* ; Hippocampus ; Moxibustion ; Neurons ; Rats ; Rats, Sprague-Dawley

Objective:Investigate the relationship between dietary patterns and sarcopenia in a Chinese elderly population.Methods:Participants in this cross-sectional study were recruited from the Tianjin Chronic Low-grade Systemic Inflammation and Health Cohort Study. The study population comprised 2 423 participants, with mean age of (67.6±5.2) years. Sarcopenia was defined based on the guidelines of the Asian Working Group for Sarcopenia. Three dietary patterns were derived by factor analysis: fruit and sweet pattern, traditional oriental pattern, and animal food pattern. The association between quartile categories of dietary pattern scores and the presence of sarcopenia was analyzed using multiple logistic regression models. Odds ratios ( OR) and 95% confidence interval ( CI) were calculated. Results:The prevalence of sarcopenia was 16.1%. After adjusting for confounding factors, compared with the lowest quartile, the adjusted OR ( 95%CI) of sarcopenia for the highest quintile of Fruit and sweet pattern score, Traditional oriental pattern score and Animal food pattern score were 1.06 (0.74, 1.50), 0.54 (0.34, 0.86), and 0.50 (0.33, 0.74), ( P for trend were 0.87,<0.01, and<0.001), respectively. Conclusions:The current study found that the traditional oriental pattern and animal food pattern has a protective relation for sarcopenia in elderly adults, which suggests its potential to attenuate or prevent the progression of sarcopenia.

Autophagy-related gene 5 (Atg5) plays an essential role in autophagy, the loss of its function impairs neurogenesis and axon regeneration. However, the biological function of Atg5 has not been characterized in planarian. Planarian is an ideal model for the study of brain regeneration. It can regenerate a new brain de novo in 1 week following amputation. To explore the role of Atg5 in planarian brain regeneration, we dissected the molecular characteristics of Atg5 in planarian Dugesia japonica (DjAtg5) and examined its function by RNAi. The full-length cDNA of DjAtg5 is 1 014 bp encoding 284 amino acids. The deduced amino sequence of DjAtg5 contains the functional Pfam domain of ATG5 and highly conserved residues for ATG5-ATG12 interaction. After amputation, the transcrips of DjAtg5 are increased and mainly distributed in the newly regenerated brain on day 3-5 of regeneration. However, knockdown of DjAtg5 by RNAi does not impair the regeneration ability and brain structure reformation, nor affects the neoblasts proliferation. Our results suggest that DjAtg5 participates in re-formation of planarian brain structure following amputation, but it is not an important regulator for planarian regeneration. However, autophagy inhibitor 3-MA can block planarian regeneration, which suggests that autophagy is necessary for planarian regeneration.

PURPOSE: Low-velocity penetrating brain injury (LVPBI) caused by foreign bodies can pose life-threatening emergencies. Their complexity and lack of validated classification data have prevented standardization of clinical management. We aimed to compare the trans-base and trans-vault phenotypes of LVPBI to help provide guidance for clinical decision-making of such injury type. METHODS: A retrospective study on LVPBI patients managed at our institution from November 2013 to March 2020 was conducted. We included LVPBI patients admitted for the first time for surgery, and excluded those with multiple injuries, gunshot wounds, pregnancy, severe blunt head trauma, etc. Patients were categorized into trans-base and trans-vault LVPBI groups based on the penetration pathway. Discharged patients were followed up by outpatient visit or telephone. The data were entered into the Electronic Medical Record system by clinicians, and subsequently derived by researchers. The demography and injury characteristics, treatment protocols, complications, and outcomes were analyzed and compared between the two groups. A t-test was used for analysis of normally distributed data, and a Mann-Whitney U test for non-parametric data. A generalized linear model was further established to determine whether the factors length of stay and performance scale score were influenced by each factor. RESULTS: A total of 27 LVPBI patients were included in this analysis, comprised of 13 (48.1%) trans-base cases and 14 (51.9%) trans-vault cases. Statistical analyses suggested that trans-base LVPBI was correlated with deeper wounds; while the trans-vault phenotype was correlated with injury by metal foreign bodies. There was no difference in Glasgow Coma Scale score and the risk of intracranial hemorrhage between the two groups. Surgical approaches in the trans-base LVPBI group included subfrontal (n = 5, 38.5%), subtemporal (n = 5, 38.5%), lateral fissure (n = 2, 15.4%), and distal lateral (n = 1, 7.7%). All patients in the trans-vault group underwent a brain convex approach using the foreign body as reference (n = 14, 100%). Moreover, the two groups differed in application prerequisites for intracranial pressure monitoring and vessel-related treatment. Trans-base LVPBI was associated with higher rates of cranial nerve and major vessel injuries; in contrast, trans-vault LVPBI was associated with lower functional outcome scores. CONCLUSION Our findings suggest that trans-base and trans-vault LVPBIs differ in terms of characteristics, treatment, and outcomes. Further understanding of these differences may help guide clinical decisions and contribute to a better management of LVPBIs.

Objective:To observe the outcome of posterior staphyloma (PS) marginal retinal photocoagulation in pars plana vitrectomy (PPV) for high myopia macular hole retinal detachment eyes accompanied with PS.Methods:From January 2017 to June 2019, 49 patients (49 eyes) with high myopia macular hole retinal detachment accompanied with PS who were undergone PPV operation from Tianjin Eye Hospital were included in this study. There were 13 males (13 eyes) and 36 females (36 eyes). All patients underwent best corrected visual acuities (BCVA) and optical coherence tomography examinations. The standard logarithmic visual acuity chart was used for BCVA examination, and the visual acuity was converted to minimum resolution angle in logarithmic (logMAR) when recorded. The patients were randomly divided into two groups according to surgical options: conventional PPV with internal limiting membrane (ILM) peeling (group A, 24 eyes), PS marginal retinal photocoagulation in PPV with ILM peeling (group A, 25 eyes). The mean preoperative logMAR BCVA of group A and B were 1.87±0.28 and 1.80±0.37, the difference was not statistically significant ( t=0.604, P=0.551). The patients in the group A received 23G PPV, triamcinolone acetonide staining during the operation, the epiretinal membrane was peeled off, indocyanine green assisted staining, the posterior macular ILM was peeled off, and the peripheral retina was examined in detail during the operation. Areas with retinal degeneration were reinforced by laser photocoagulation, and the subretinal fluid was drained through the macular hole and filled with silicone oil. The eyes of the group B were subjected to retinal photocoagulation for 2 to 3 rows at the edge of the PS in addition to the usual surgical procedures. The average follow-up time was 8.34±3.21 months. Surgical outcome were estimated by the average number of operation, retinal reattachment rate, macular hole closure rate and BCVA. The χ2 test or Fisher exact probability was used to compare the count data. Independent sample t test was used to compare the measurement data. Results:Retinal reattachment was obtained in 17 eyes (70.8%, 17/24) and 24 eyes (96.0%, 24/25) in group A and B after first surgery respectively, the difference was statistically significant ( χ2=3.984, P=0.046). Final retinal reattachment was obtained in all 49 eyes. Final macular hole closure was in 15 eyes (62.5%, 15/24) and 19 eyes (76.0%, 19/25) in group A and B, respectively, the difference was not statistically significant ( χ2=1.051, P=0.305). The mean postoperative logMAR BCVA of group A (1.20±0.47) and B (1.08±0.39) were all improved than preoperative BCVA, the differences were all statistically significant ( t=2.899, 5.327; P=0.001, 0.000), the differences of mean postoperative logMAR BCVA between two groups was not statistically significant ( t=0.675, P=0.506). The mean number of operation of group A (2.63±0.88) was more than group B (2.08±0.28), the difference was statistically significant ( t=3.003, P=0.006). Conclusion:In comparison with conventional PPV, combined PS marginal retinal photocoagulation can improve retinal reattachment rate after first surgery, and reduce the number of reoperations.

Objective:To access the independent associations between serum ferritin quintile levels and metabolic syndrome (MS) in adults of different genders.Methods:19 563 participants over the age of 18 years were recruited from "TCLSIH Cohort Study" from 2007 to 2015. Serum ferritin concentration was measured by Enzyme-linked immunoassay, while metabolic syndrome was diagnosed according to MS diagnostic criteria formulated by Chinese Diabetes Society in 2013. Multiple logistic regression analysis was used to assess the association between serum ferritin quintile levels and the prevalence of MS in males and females. Results:After adjusting the confounding factors, the overall prevalence of MS gradually increased with the increasing of serum ferritin levels, similar results were observed in males and females. Subjects were divided into 5 subgroups according to serum ferritin levels. Compared with level 1 group, logistic regression showed that the serum ferritin quintiles of males and females ranged from low to high, the OR (95% confidence interval) for metabolic syndrome were 1.142 (0.998, 1.307), 1.382 (1.210, 1.579), 1.680 (1.472, 1.917), 2.085 (1.827, 2.380), respectively ( Ptrend<0.01), and 1.147 (0.911, 1.444), 1.346 (1.075, 1.687), 1.567 (1.268, 1.941), 2.444 (1.981, 3.023), respectively ( Ptrend<0.01). Conclusion:The elevated ferritin levels were positively related to the prevalence of metabolic syndrome in adults of different genders.