Histological staining is the basis of pathological analysis,but the traditional staining method relies on chemical reagents,which not only consumes a lot of resources,but also causes harm to the environment and human health.In recent years,with the rapid development of deep learning technology,virtual staining technology,as a new method,is expected to effectively replace and supplement the traditional histological staining methods.It uses neural networks to analyze unstained tissue images,generate digital images that are highly similar to chemical staining effects,and even realize the mutual conversion between different staining modes,reducing the laboratory's dependence on chemical reagents and providing sustainable research programs.In this paper,the basic principles of virtual staining and its potential applications in histopathology are introduced in detail,and the current challenges and future research directions are discussed.

Objective To study the influence of laser diffraction ultrasound parameters on the determination of particle size and particle size distribution of budesonide suspension for inhalation,and explore the rationality of ultrasound parameter settings of the laser diffraction method.Methods The Mastersizer 3000 was set to measure the particle size and particle size distribution of budesonide suspension for inhalation from 6 companies at different ultrasonic times,and the FlowCam was used to test the samples simultaneously.Results The ultrasound parameters have a significant impact on the particle size and particle size distribution of budesonide suspension for inhalation in some companies,and the FlowCam test results show that particles in some samples exhibit aggregation,which can be dispersed by ultrasound and affect the results.Conclusion It is necessary to optimize the ultrasound parameters of laser diffraction to determine the particle size and particle size distribution of suspension for inhalation,and verify the rationality of the ultrasound parameter settings through microscopic techniques.

Objective To explore the neuroprotective effects and mechanisms of the sesquiterpene lactone compound ACT001 on rotenone(ROT)-induced Parkinson's disease(PD)model mouse.Methods SPF C57BL/6 mice were randomly divided into 6 groups,including control group,solvent control group,ROT model group,ACT001 5 mg/kg group(ROT+ACT001-5),ACT001 20 mg/kg group(ROT+ACT001-20),and levodopa(L-dopa)positive control group(ROT+L-dopa),with 9 mice in each group.The control group received an equivalent amount of intraperitoneal injection of saline,the solvent control group received an equivalent amount of rotenone solvent without rotenone,the remaining groups of mice were used to establish a PD mouse model by intraperitoneal injection of rotenone.Mice in different ACT001 dosage groups received intraperitoneal injections of high and low doses of ACT001,while the positive control group received levodopa intraperitoneally for 15 consecutive days.Behavioral changes in mice were assessed using open field,rotarod,pole-climbing,and balance beam tests.Immunofluorescence(IF)assay to detect the expression of tyrosine hydroxylase(TH)neurons,content of TH-positive fibers in the striatum and to detect the activation status of nigrostriatal microglia in the mouse midbrain;Real-time PCR was employed to measure the levels of interleukin(IL)-6,IL-1β,and tumor necrosis factor-α(TNF-α)in the substantia nigra of the mouse brain.Western blotting was used to measure the protein levels of TH,nuclear factor-κB(NF-κB)p65,NF-κB inhibitor α(IκBα),and phosphorylated IκBα(p-IκBα)in the substantia nigra of the mouse brain.Results Compared to the control group and the solvent control group,the rotenone-induced PD model group exhibited motor impairments in behavioral tests,a decrease in the number of TH positive neurons in the substantia nigra(P＜0.0001),decreased levels of TH-positive fibers in the striatum,activation of midbrain substantia microglia,and elevated levels of IL-6,IL-1β,TNF-α,p-IκBα,and NF-κB p65 expression.ACT001 significantly improved the behavioral impairments and substantia nigra damage in PD mice,increased the number of TH-positive neurons in the substantia nigra,increased levels of TH-positive fibers in the striatum,inhibition of microglial cell activation in the midbrain substantia nigra,and elevated the protein expression levels of IκBα while reducing the levels of IL-6,IL-1β,TNF-α,p-IκBα,and NF-κB p65 in the substantia nigra(P＜0.05).At a dose of 5 mg/kg,ACT001 significantly improved behavioral impairments in rotenone-induced PD mice,reduced the loss of dopaminergic neurons,and its mechanism may be related to the inhibition of the NF-κB signaling pathway and the suppression of inflammation.In summary,the intervention of ACT001 in the rotenone-induced PD mouse model inhibited the inflammatory response in the midbrain,increased the number of TH-positive neurons,and augmented the population of dopaminergic neurons in the substantia nigra,exerting a protective effect on neurons.Conclusion ACT001 significantly improves behavioral deficits in ROT-induced PD mice,ameliorates of dopaminergic neuron loss from the midbrain substantia nigra and striatum,inhibits the activation of nigrostriatal microglia in the midbrain,and suppresses inflammatory responses by inhibiting the activation of the NF-κB signaling pathway.

Objective To study the influence of laser diffraction ultrasound parameters on the determination of particle size and particle size distribution of budesonide suspension for inhalation,and explore the rationality of ultrasound parameter settings of the laser diffraction method.Methods The Mastersizer 3000 was set to measure the particle size and particle size distribution of budesonide suspension for inhalation from 6 companies at different ultrasonic times,and the FlowCam was used to test the samples simultaneously.Results The ultrasound parameters have a significant impact on the particle size and particle size distribution of budesonide suspension for inhalation in some companies,and the FlowCam test results show that particles in some samples exhibit aggregation,which can be dispersed by ultrasound and affect the results.Conclusion It is necessary to optimize the ultrasound parameters of laser diffraction to determine the particle size and particle size distribution of suspension for inhalation,and verify the rationality of the ultrasound parameter settings through microscopic techniques.

Objective To develop and validate a nomogram model that integrates systemic inflammatory markers to predict the likelihood of pelvic lymph node metastasis(PLNM)in prostate cancer patients prior to surgery.Methods This study retrospectively analyzed the clinical data and preoperative inflammatory markers—including neutrophil-to-lymphocyte ratio(NLR),platelet-to-lymphocyte ratio(PLR),systemic immune-inflammation index(SII),and monocyte-to-lymphocyte ratio(MLR)—of patients diagnosed with prostate cancer.Univariate and multi-variate logistic regression analyses were conducted to identify markers that were significantly associated with PLNM.Based on the results of the multivariate analysis,a nomogram was developed and its predictive accuracy was assessed using receiver operating characteristic curves(ROC)and calibration plots.Results Among the 334 enrolled patients with prostate cancer,107 were identified with PLNM.Univariate analysis revealed statistically significant differences in free prostate-specific antigen(fPSA),Gleason score,NLR,PLR,MLR,and SII between the PLNM and non-pelvic lymph node metastasis(NPLNM)groups(P<0.05).Multivariate analysis confirmed that fPSA,Gleason score,and SII were independent predictors of PLNM(P<0.05).A nomogram incorporating these predic-tors exhibited strong discriminative ability,with an area under the ROC curve(AUC)of 0.79(95%CI:0.73～0.84).Calibration analysis further demonstrated good consistency between the predicted and observed probabilities of PLNM.Conclusions This study successfully developed a nomogram model based on systemic inflammatory markers for preoperative prediction of pelvic lymph node metastasis in prostate cancer.Owing to its user-friendly design and high predictive accuracy,this tool may serve as a valuable complementary method to conventional imaging techniques,thereby supporting personalized treatment decision-making.

BACKGROUND: The potential impact of pre-existing coronary artery stenosis (CAS) on right ventricular (RV) function during acute pulmonary embolism (PE) episodes remains underexplored. This study aimed to investigate the association between pre-existing CAS and RV dysfunction in patients with acute PE. METHODS: In this multicenter, case-control study, 89 cases and 176 controls matched for age were enrolled at three study centers (Peking Union Medical College Hospital, Fuwai Hospital, and the Second Affiliated Hospital of Harbin Medical University) from January 2016 to December 2020. The cases were patients with acute PE with CAS, and the controls were patients with acute PE without CAS. Coronary artery assessment was performed using coronary computed tomographic angiography. CAS was defined as ≥50% stenosis of the lumen diameter in any coronary vessel >2.0 mm in diameter. Conditional logistic regression analysis was used to evaluate the association between CAS and RV dysfunction. RESULTS: The percentages of RV dysfunction (19.1% [17/89] vs. 44.6% [78/176], P <0.001) and elevated systolic pulmonary artery pressure (sPAP) (19.3% [17/89] vs. 39.5% [68/176], P = 0.001) were significantly lower in the case group than those in the control group. In the multivariable logistic regression model, CAS was independently and negatively associated with RV dysfunction (adjusted odds ratio [OR]: 0.367; 95% confidence interval [CI]: 0.185-0.728; P = 0.004), and elevated sPAP (OR: 0.490; 95% CI: 0.252-0.980; P = 0.035), respectively. CONCLUSIONS Pre-existing CAS was significantly and negatively associated with RV dysfunction and elevated sPAP in patients with acute PE. This finding provides new insights into RV dysfunction in patients with acute PE with pre-existing CAS.
Humans ; Pulmonary Embolism/complications* ; Case-Control Studies ; Male ; Ventricular Dysfunction, Right/physiopathology* ; Female ; Middle Aged ; Aged ; Coronary Stenosis/complications* ; Logistic Models ; Adult

The study aimed to construct the second and third generation chimeric antigen receptor T cells (CAR-T) targeting the c-mesenchymal-epithelial transition factor (c-Met) protein, and observe their killing effect on human serous ovarian cancer cell SKOV-3. The expression of MET gene in ovarian serous cystadenocarcinoma, the correlation between MET gene expression and the abundance of immune cell infiltration, and the effect of MET gene expression on the tissue function of ovarian serous cystadenocarcinoma were analyzed by bioinformatics. The expression of c-Met in ovarian cancer tissues and adjacent tissues was detected by immunohistochemical staining. The second and third generation c-Met CAR-T cells, namely c-Met CAR-T(2G/3G), were prepared by lentivirus infection, and the cell subsets and infection efficiency were detected by flow cytometry. Using CD19 CAR-T and activated T cells as control groups and A2780 cells with c-Met negative expression as Non target groups, the kill efficiency on SKOV-3 cells with c-Met positive expression, cytokine release and cell proliferation of c-Met CAR-T(2G/3G) were explored by lactate dehydrogenase (LDH) release, ELISA and CCK-8 respectively. The results showed that MET gene expression was significantly up-regulated in ovarian cancer tissues compared with normal tissues, which was consistent with the immunohistochemistry results. However, in all pathological stages, there was no obvious difference in MET expression and no correlation between MET gene expression and the race and age of ovarian cancer patients. The second generation and third generation c-Met CAR-T cells were successfully constructed. After lentivirus infection, the proportion of CD8⁺ T cells in c-Met CAR-T(2G) was upregulated, while there was no significant change in the cell subsets of c-Met CAR-T(3G). The LDH release experiment showed that the kill efficiency of c-Met CAR-T(2G/3G) on SKOV-3 increased with the increase of effect-target ratio. When the effect-target ratio was 20:1, the kill efficiency of c-Met CAR-T(2G) reached (42.02 ± 5.17)% (P < 0.05), and the kill efficiency of c-Met CAR-T(3G) reached (51.40 ± 2.71)% (P < 0.05). ELISA results showed that c-Met CAR-T released more cytokine compared to CD19 CAR-T and activated T cells (P < 0.05). Moreover, the cytokine release of c-Met CAR-T(3G) was higher than c-Met CAR-T(2G) (P < 0.01). The CCK-8 results showed that after 48 h, the cell number of c-Met CAR-T(2G) was higher than that of c-Met CAR-T(3G) (P < 0.01). In conclusion, both the second and third generation c-Met CAR-T can target and kill c-Met-positive SKOV-3 cells, with no significant difference. c-Met CAR-T(2G) has stronger proliferative ability, and c-Met CAR-T(3G) releases more cytokines.
Humans ; Female ; Ovarian Neoplasms/immunology* ; Proto-Oncogene Proteins c-met/metabolism* ; Receptors, Chimeric Antigen/immunology* ; Cell Line, Tumor ; Cystadenocarcinoma, Serous/immunology* ; T-Lymphocytes/immunology*

Purpose: This study aimed to evaluate the contrast-enhanced ultrasound with Sonazoid (Sonazoid-CEUS) features of hepatocellular carcinoma (HCC) in patients with non-alcoholic fatty liver disease (NAFLD). Methods: In this retrospective study, patients who underwent surgical resection and were histopathologically diagnosed with NAFLD or cirrhosis-related HCC were included. All patients received Sonazoid-CEUS examinations within 1 week prior to hepatic surgery. The enhancement patterns of HCC lesions were evaluated and compared between the two groups according to the current World Federation for Ultrasound in Medicine and Biology guidelines. Multivariate logistic regression analysis was used to assess the correlations between Sonazoid-CEUS enhancement patterns and clinicopathologic characteristics. Results: From March 2022 to April 2023, a total of 151 patients with HCC were included, comprising 72 with NAFLD-related HCC and 79 with hepatitis B virus (HBV) cirrhosis–related HCC. On Sonazoid-CEUS, more than half of the NAFLD-related HCCs exhibited relatively early and mild washout within 60 seconds (54.2%, 39/72), whereas most HBV cirrhosis–related HCCs displayed washout between 60 and 120 seconds (46.8%, 37/79) or after 120 seconds (39.2%, 31/79) (P<0.001). In the patients with NAFLD-related HCC, multivariate analysis revealed that international normalized ratio (odds ratio [OR], 0.002; 95% confidence interval [CI], 0.000 to 0.899; P=0.046) and poor tumor differentiation (OR, 21.930; 95% CI, 1.960 to 245.319; P=0.012) were significantly associated with washout occurring within 60 seconds. Conclusion Characteristic Sonazoid-CEUS features are useful for diagnosing HCC in patients with NAFLD.

Purpose: This study aimed to evaluate the contrast-enhanced ultrasound with Sonazoid (Sonazoid-CEUS) features of hepatocellular carcinoma (HCC) in patients with non-alcoholic fatty liver disease (NAFLD). Methods: In this retrospective study, patients who underwent surgical resection and were histopathologically diagnosed with NAFLD or cirrhosis-related HCC were included. All patients received Sonazoid-CEUS examinations within 1 week prior to hepatic surgery. The enhancement patterns of HCC lesions were evaluated and compared between the two groups according to the current World Federation for Ultrasound in Medicine and Biology guidelines. Multivariate logistic regression analysis was used to assess the correlations between Sonazoid-CEUS enhancement patterns and clinicopathologic characteristics. Results: From March 2022 to April 2023, a total of 151 patients with HCC were included, comprising 72 with NAFLD-related HCC and 79 with hepatitis B virus (HBV) cirrhosis–related HCC. On Sonazoid-CEUS, more than half of the NAFLD-related HCCs exhibited relatively early and mild washout within 60 seconds (54.2%, 39/72), whereas most HBV cirrhosis–related HCCs displayed washout between 60 and 120 seconds (46.8%, 37/79) or after 120 seconds (39.2%, 31/79) (P<0.001). In the patients with NAFLD-related HCC, multivariate analysis revealed that international normalized ratio (odds ratio [OR], 0.002; 95% confidence interval [CI], 0.000 to 0.899; P=0.046) and poor tumor differentiation (OR, 21.930; 95% CI, 1.960 to 245.319; P=0.012) were significantly associated with washout occurring within 60 seconds. Conclusion Characteristic Sonazoid-CEUS features are useful for diagnosing HCC in patients with NAFLD.

Immediate implant placement in the aesthetic zone has become increasingly widespread and has gradually evolved into a conventional techniques for implant procedures in the aesthetic region. To achieve favorable aesthetic and long-term outcomes, clinicians must possess extensive clinical experience as well as proficient surgical and restorative skills. This study summarizes the key factors influencing the long-term success of immediate implants in the aesthetic zone: strict adherence to the indications for immediate implant placement; thorough preoperative assessment of the patient’s systemic and local conditions, along with comprehensive evaluation of aesthetic risks; minimally invasive tooth extraction while preserving the integrity of the labial bone plate; selection of appropriately designed implants and their placement in an ideal three-dimensional position based on the implant’s characteristics; utilization of suitable bone and soft tissue augmentation techniques according to the patient’s specific hard and soft tissue anatomy, extent of bone defects, and periodontal phenotype; dynamic shaping of soft tissues through continuous adjustments in the emergence profile of provisional restorations; design of definitive restorations from the perspectives of health, function, and aesthetics; and implementation of regular follow-up and maintenance protocols after implant treatment, with increased emphasis on peri-implant care for patients who smoke, have diabetes, or undergo anti-osteoporosis therapy. This study proposes a decision-making framework to achieve long-term stable clinical outcomes with immediate implants in the aesthetic zone, providing a reference for clinicians in their clinical decision-making and treatment planning: ① for patients assessed as low aesthetic risk (e.g., thick gingival biotype, absence of hard and soft tissue defects, intact labial bone plate with thickness >1 mm, no acute infection), immediate implant placement after minimally invasive extraction is recommended, with the implant positioned in an ideal three-dimensional location, along with bone grafting in the gap between the implant and the labial bone plate and consideration of connective tissue grafting when required; ② for patients assessed as moderate aesthetic risk (e.g., thin gingival biotype, absence of soft tissue defects, intact labial bone plate but with thickness <1 mm or mild to moderate bone defects involving less than 50% height loss, chronic infection present), immediate implant placement with optimal three-dimensional positioning is feasible, accompanied by bone grafting in the implant-labial bone gap or external bone grafting on the labial aspect, with simultaneous or staged connective tissue grafting, or alternatively, use of the socket shield technique for immediate implant placement; ③ for patients assessed as high aesthetic risk (e.g., thin gingival biotype, presence of soft tissue defects, vertical bone deficiency, severe labial bone loss involving >50% height loss, acute infection present), ridge preservation followed by delayed implant placement is advised. By adhering to these treatment principles, immediate implant placement in the aesthetic zone can achieve reliable success rates and excellent aesthetic outcomes.