Objective:To investigate effect of cedrol on radiosensitivity of prostate cancer(PCa)cells and its mechanism.Methods:mRNA and protein expressions of cGAS and STING in PCa tissues and cells were detected by qRT-PCR and Western blot,respectively.Human PCa cells PC-3 were divided into control group,radiation group,cedrol group,combination group and inhibitor group.Radiosensitivity of cells in each group was detected by plate cloning test;apoptosis,migration and invasion were detected by flow cytometry,scratch test and Transwell chamber,respectively;γ-H2AX immunofluorescence staining was used to analyze effect of cedrol on DNA damage repair;after PC-3 cells and CD8+T cells were co-cultured,cells were divided into T cell group,co-culture group,cedrol group and inhibitor group,MTT and flow cytometry were used to detect proliferation and apoptosis of CD8+T cells,IFN-γ,IL-2 and TNF-α levels in supernatant of CD8+T cells were detected by ELISA;Western blot was used to detect prolife-rating cell nuclear antigen(PCNA),Bcl-2 associated protein(Bax),programmed death recepter ligand 1(PD-L1)and cGAS-STING signal pathway protein.Results:cGAS and STING protein and mRNA expressions in PCa tissues and cells were decreased(P<0.05),which were lowest in PC-3 cells.Compared with control group,colony formation rate,cell survival rate,scratch healing rate and cell inva-sion rate of PC-3 cells in radiation group and cedrol group were decreased obviously,apoptosis rate and number of γ-H2AX focus were increased obviously(P<0.05);compared with radiation group and cedrol group,combined group inhibited proliferation,migration and invasion of PC-3 cells,and induced DNA damage and apoptosis(P<0.05);inhibitor group attenuated inhibitory effect of com-bined group on proliferation,migration and invasion of PC-3 cells,and promoted DNA damage and apoptosis(P<0.05);compared with T cell group,A490,levels of IFN-γ,IL-2 and TNF-α in CD8+T cells in co-culture group were decreased,apoptosis rate was in-creased(P<0.05);compared with co-culture group,A490,levels of IFN-γ,IL-2 and TNF-α in CD8+T cells in cedrol group were in-creased,apoptosis rate was decreased(P<0.05);compared with cedrol group,A490,levels of IFN-γ,IL-2 and TNF-α in CD8+T cells in inhibitor group were decreased,apoptosis rate was increased(P<0.05).Compared with control group,expressions of PCNA and PD-L1 proteins in PC-3 cells in cedrol group were reduced obviously,expressions of Bax,cGAS and STING proteins were increased obviously(P<0.05);inhibitors of cGAS-STING pathway could reverse effect of cedrol on above proteins(P<0.05).Conclusion:Cedrol may enhance radiosensitivity of PCa cells by activating cGAS-STING signal pathway and inhibiting immune escape.

Objective The pathophysiological process of acute high-altitude hypoxia-induced lung injury affects protein expression levels,which are mainly evaluated by Western blot.No systematic study has investigated changes in internal control proteins as calibration loading amounts.Methods Lung injury at an altitude of 6000 m was induced in a low-pressure,low-oxygen chamber for 8,24,and 72 h using C57BL/6J mice.Establishment of the model was confirmed by hematoxylin and eosin staining.Expression levels of various internal control proteins,including vinculin,α-tubulin,eukaryotic translation initiation factor 5(EIF5),β-actin,and glyceraldehyde 3-phosphate dehydrogenase(GAPDH)were detected by Western blot,and total protein expression was detected by Coomassie blue staining.Furthermore,the lung injury model in vitro was establised by using,Bronchial epithelial cell(BZAS-2B)andhunman umbilical vein endothelial cells(HUVECS)confirmed by TUNEL staining.Expression levels of internal control proteins were detected by Western blot,and total protein expression was detected by Coomassie Blue staining.Results Acute 8,24,and 72 h hypoxic models were successfully established in lung tissue,demonstrating consistent total protein expression and stable levels of the internal reference proteins vinculin,α-tubulin,EIF5,andβ-actin.GAPDH expression was elevated in the HH8 h,HH24 h,and HH72 h groups compared with the normoxia(Nor)group,but only the increase at HH72 h groups was significant.Similarly,8,24,and 48 h hypoxic models were successfully established in BEAS-2B cells and HUVECs,with consistent total protein expression.In BEAS-2B cells,expression levels of the internal reference proteins β-actin and GAPDH were consistent with the normoxic control(NC)group,while vinculin,α-tubulin,and EIF5 expression levels were significantly reduced under hypoxic conditions for up to 24 h.In HUVECs,vinculin and α-tubulin expression levels were also consistent with the NC group,while EIF5,β-actin,and GAPDH expression levels were significantly reduced at 8 h and increased at 48 h.Conclusions Acute hypoxia induces lung tissue injury,and protein expression levels of the internal reference proteins vinculin,α-tubulin,EIF5,and β-actin are stable,making them suitable internal references for Western blot.Additionally,Western blot detected differential expression levels of the internal reference proteins vinculin,α-tubulin,EIF5,β-actin,and GAPDH in BEAS-2B cells and HUVECs,as the most important in vitro lung tissue models of hypoxia-induced injury.

Hepatic stellate cells (HSCs) are the primary fibrogenic cells in the liver, and their activation plays a crucial role in the development and progression of hepatic fibrosis. Here, we report that retinoid X receptor-alpha (RXRα), a unique member of the nuclear receptor superfamily, is a key modulator of HSC activation and liver fibrosis. RXRα exerts its effects by modulating calcium/calmodulin-dependent protein kinase kinase β (CaMKKβ)-mediated activation of AMP-activated protein kinase-alpha (AMPKα). In addition, we demonstrate that K-80003, which binds RXRα by a unique mechanism, effectively suppresses HSC activation, proliferation, and migration, thereby inhibiting liver fibrosis in the CCl₄ and amylin liver NASH (AMLN) diet animal models. The effect is mediated by AMPKα activation, promoting mitophagy in HSCs. Mechanistically, K-80003 activates AMPKα by inducing RXRα to form condensates with CaMKKβ and AMPKα via a two-phase process. The formation of RXRα condensates is driven by its N-terminal intrinsic disorder region and requires phosphorylation by CaMKKβ. Our results reveal a crucial role of RXRα in liver fibrosis regulation through modulating mitochondrial activities in HSCs. Furthermore, they suggest that K-80003 and related RXRα modulators hold promise as therapeutic agents for fibrosis-related diseases.

Objective The pathophysiological process of acute high-altitude hypoxia-induced lung injury affects protein expression levels,which are mainly evaluated by Western blot.No systematic study has investigated changes in internal control proteins as calibration loading amounts.Methods Lung injury at an altitude of 6000 m was induced in a low-pressure,low-oxygen chamber for 8,24,and 72 h using C57BL/6J mice.Establishment of the model was confirmed by hematoxylin and eosin staining.Expression levels of various internal control proteins,including vinculin,α-tubulin,eukaryotic translation initiation factor 5(EIF5),β-actin,and glyceraldehyde 3-phosphate dehydrogenase(GAPDH)were detected by Western blot,and total protein expression was detected by Coomassie blue staining.Furthermore,the lung injury model in vitro was establised by using,Bronchial epithelial cell(BZAS-2B)andhunman umbilical vein endothelial cells(HUVECS)confirmed by TUNEL staining.Expression levels of internal control proteins were detected by Western blot,and total protein expression was detected by Coomassie Blue staining.Results Acute 8,24,and 72 h hypoxic models were successfully established in lung tissue,demonstrating consistent total protein expression and stable levels of the internal reference proteins vinculin,α-tubulin,EIF5,andβ-actin.GAPDH expression was elevated in the HH8 h,HH24 h,and HH72 h groups compared with the normoxia(Nor)group,but only the increase at HH72 h groups was significant.Similarly,8,24,and 48 h hypoxic models were successfully established in BEAS-2B cells and HUVECs,with consistent total protein expression.In BEAS-2B cells,expression levels of the internal reference proteins β-actin and GAPDH were consistent with the normoxic control(NC)group,while vinculin,α-tubulin,and EIF5 expression levels were significantly reduced under hypoxic conditions for up to 24 h.In HUVECs,vinculin and α-tubulin expression levels were also consistent with the NC group,while EIF5,β-actin,and GAPDH expression levels were significantly reduced at 8 h and increased at 48 h.Conclusions Acute hypoxia induces lung tissue injury,and protein expression levels of the internal reference proteins vinculin,α-tubulin,EIF5,and β-actin are stable,making them suitable internal references for Western blot.Additionally,Western blot detected differential expression levels of the internal reference proteins vinculin,α-tubulin,EIF5,β-actin,and GAPDH in BEAS-2B cells and HUVECs,as the most important in vitro lung tissue models of hypoxia-induced injury.

Objective:To investigate effect of cedrol on radiosensitivity of prostate cancer(PCa)cells and its mechanism.Methods:mRNA and protein expressions of cGAS and STING in PCa tissues and cells were detected by qRT-PCR and Western blot,respectively.Human PCa cells PC-3 were divided into control group,radiation group,cedrol group,combination group and inhibitor group.Radiosensitivity of cells in each group was detected by plate cloning test;apoptosis,migration and invasion were detected by flow cytometry,scratch test and Transwell chamber,respectively;γ-H2AX immunofluorescence staining was used to analyze effect of cedrol on DNA damage repair;after PC-3 cells and CD8+T cells were co-cultured,cells were divided into T cell group,co-culture group,cedrol group and inhibitor group,MTT and flow cytometry were used to detect proliferation and apoptosis of CD8+T cells,IFN-γ,IL-2 and TNF-α levels in supernatant of CD8+T cells were detected by ELISA;Western blot was used to detect prolife-rating cell nuclear antigen(PCNA),Bcl-2 associated protein(Bax),programmed death recepter ligand 1(PD-L1)and cGAS-STING signal pathway protein.Results:cGAS and STING protein and mRNA expressions in PCa tissues and cells were decreased(P<0.05),which were lowest in PC-3 cells.Compared with control group,colony formation rate,cell survival rate,scratch healing rate and cell inva-sion rate of PC-3 cells in radiation group and cedrol group were decreased obviously,apoptosis rate and number of γ-H2AX focus were increased obviously(P<0.05);compared with radiation group and cedrol group,combined group inhibited proliferation,migration and invasion of PC-3 cells,and induced DNA damage and apoptosis(P<0.05);inhibitor group attenuated inhibitory effect of com-bined group on proliferation,migration and invasion of PC-3 cells,and promoted DNA damage and apoptosis(P<0.05);compared with T cell group,A490,levels of IFN-γ,IL-2 and TNF-α in CD8+T cells in co-culture group were decreased,apoptosis rate was in-creased(P<0.05);compared with co-culture group,A490,levels of IFN-γ,IL-2 and TNF-α in CD8+T cells in cedrol group were in-creased,apoptosis rate was decreased(P<0.05);compared with cedrol group,A490,levels of IFN-γ,IL-2 and TNF-α in CD8+T cells in inhibitor group were decreased,apoptosis rate was increased(P<0.05).Compared with control group,expressions of PCNA and PD-L1 proteins in PC-3 cells in cedrol group were reduced obviously,expressions of Bax,cGAS and STING proteins were increased obviously(P<0.05);inhibitors of cGAS-STING pathway could reverse effect of cedrol on above proteins(P<0.05).Conclusion:Cedrol may enhance radiosensitivity of PCa cells by activating cGAS-STING signal pathway and inhibiting immune escape.

Macrophages are vital in maintaining tissue homeostasis in the lungs by modulating and regulating immune responses.Based on different origins and anatomical locations,macrophages in the lungs are categorized into alveolar macrophages,interstitial macrophages,perivascular macrophages,and inflammatory macrophages.Alveolar macrophages are located in the alveolar spaces and are primarily responsible for maintaining alveolar surfactant homeostasis,defending against pathogens and regulating immune responses.Interstitial macrophages can maintain homeostasis,regulate immunity and anti-inflammation in the lung tissue.Perivascular macrophages play a crucial role in inhibiting lung inflammation,improving pulmonary fibrosis,and regulating lung tumor progression due to antigen-presenting and immunomodulatory effects.Inflammatory macrophages,which are differentiated from monocytes during inflammation,regulate the inflammatory process.This article reviews the origins of various subpopulations of macro-phages in the lung tissue and their physiological and pathological functions as well as discusses the underlying mechanisms and potential therapeutic targets.

Objective:To investigate the interactive effect of mismatch repair (MMR) protein status and adverse clinicopathological features on prognosis of stage Ⅰ-Ⅲ colon cancer.Methods:The retrospective cohort study was conducted. The clinicopathological data of 1 650 patients with colon cancer of stage Ⅰ-Ⅲ who were admitted to 7 hospitals in China from January 2016 to December 2017 were collected. There were 963 males and 687 females, aged 62(53,71)years. Patients were classified as 230 cases of MMR deficiency (dMMR) and 1 420 cases of MMR proficiency (pMMR) based on their MMR protein status. Observation indicators: (1) comparison of clinicopathological characteristics between patients of different MMR protein status; (2) analysis of factors affecting the survival outcomes of patients of dMMR; (3) analysis of factors affecting the survival outcomes of patients of pMMR; (4) interaction analysis of MMR and adverse clinicopathological features on survival outcomes. Measurement data with normal distribution were represented as Mean± SD, and comparison between groups was conducted using the independent t test. Measurement data with skewed distribution were represented as M( Q1, Q3), and comparison between groups was conducted using the Mann-Whitney U test. Count data were described as absolute numbers, and comparison between groups was conducted using the chi-square test or Fisher exact probability. Comparison of ordinal data was conducted using the Mann-Whitney U test. The random forest interpolation method was used for missing values in data interpolation. Univariate analysis was conducted using the COX proportional risk regression model, and multivariate analysis was conducted using the COX stepwise regression with forward method. The coefficient of multiplication interaction effect was obtained using the interaction term coefficient of COX proportional risk regression model. Evaluation of additive interaction effects was conducted using the relative excess risk due to interaction ( RERI). Results:(1) Comparison of clinicopathological characteristics between patients of different MMR protein status. There were significant differences in age, T staging, the number of lymph node harvest, the number of lymph node harvest <12, high grade tumor between patients of dMMR and pMMR ( P<0.05). (2) Analysis of factors affecting the survival outcomes of patients of dMMR. Results of multivariate analysis showed that T staging, N staging, the number of lymph node harvest <12 were independent factors affecting the disease-free survival (DFS) of colon cancer patients of dMMR ( hazard ratio=3.548, 2.589, 6.702, 95% confidence interval as 1.460-8.620, 1.064-6.301, 1.886-23.813, P<0.05). Age and N staging were independent factors affecting the overall survival (OS) of colon cancer patients of dMMR ( hazard ratio=1.073, 10.684, 95% confidence interval as 1.021-1.126, 2.311-49.404, P<0.05). (3) Analysis of factors affecting the survival outcomes of patients of pMMR. Results of multivariate analysis showed that age, T staging, N staging, vascular tumor thrombus were independent factors affecting the DFS of colon cancer patients of pMMR ( hazard ratio=1.018, 2.214, 2.598, 1.549, 95% confidence interval as 1.006-1.030, 1.618-3.030, 1.921-3.513, 1.118-2.147, P<0.05). Age, T staging, N staging, high grade tumor were independent factors affecting the OS of colon cancer patients of pMMR ( hazard ratio=1.036, 2.080, 2.591, 1.615, 95% confidence interval as 1.020-1.052, 1.407-3.075, 1.791-3.748, 1.114-2.341, P<0.05). (4) Interaction analysis of MMR and adverse clinicopathological features on survival outcomes. Results of interaction analysis showed that the multiplication interaction effect between the number of lymph node harvest <12 and MMR protein status was significant on DFS of colon cancer patients ( hazard ratio=3.923, 95% confidence interval as 1.057-14.555, P<0.05). The additive interaction effects between age and MMR protein status, between high grade tumor and MMR protein status were significant on OS of colon cancer patients ( RERI=-0.033, -1.304, 95% confidence interval as -0.049 to -0.018, -2.462 to -0.146). Conclusions:There is an interaction between the MMR protein status and the adverse clinicopathological features (the number of lymph node harvest <12, high grade tumor) on prognosis of colon cancer patients of stage Ⅰ-Ⅲ. In patients of dMMR, the number of lymph node harvest <12 has a stronger predictive effect on poor prognosis. In patients of pMMR, the high grade tumor has a stronger predictive effect on poor prognosis.

Objective:To compare the clinical value of digital breast tomosynthesis (DBT) localization and stereotactic positioning biopsy of breast lesions.Methods:This study was a cross-sectional study. Totally of 250 patients who underwent breast biopsy at Shenzhen People′s Hospital, Luohu District People′s Hospital and Peking University Shenzhen Hospital between August 2021 to October 2023 was analyzed retrospectively, including 136 cases of DBT-guided biopsy (DBT-guided group) and 114 cases of stereotactic-guided biopsy (stereotactic-guided group). The stereotactic-guided biopsy methods included core needle biopsy (CNB) and wire positioning. The DBT-guided biopsy methods included CNB, wire positioning and vacuum-assisted breast biopsy (VABB). The χ2 test or Mann-Whitney U test was used to compare the puncture success rate, operation time, localization time, puncture time, number of first valid localization phases obtained, number of exposures, and complications of different biopsy methods between 2 groups. Results:In the wire positioning biopy, the puncture success rate was 100% (33/33) in DBT-guided group and 96% (48/50) in the stereotactic-guided group, with no statistically significant difference between the two groups ( P=0.515). Compared to the stereotactic-guided group, the operation time and localization time were shorter, and the number of first valid localization phases obtained, number of exposures were fewer in the DBT-guided group( P<0.05). The incidence of complications was lower in both the DBT-guided group and the stereotactic-guided group, with no statistically significant difference ( P=0.871). In CNB, both the DBT-guided group and the stereotactic-guided group had higher puncture success rates, with no statistically significant difference ( P=0.080). Compared to the stereotactic-guided group, the operation time, localization time and puncture time were shorter, and the number of first valid localization phases obtained, number of exposures were lower in the DBT-guided group, and the difference between the two groups were statistically significant ( P<0.05). The incidence of complications was lower in both the DBT-guided group and the stereotactic-guided group, with no statistically significant difference ( P=0.627). Twenty-one cases received DBT-guided VABB, with an operation time of (19.90±3.38) min, a localization time of 6.00 (6.00, 7.00) min, a puncture time of (13.42±3.28) min, the number of first effective localization phases obtained was 1.00 (1.00, 1.00) time, the number of exposures was 4.00 (3.50, 5.00) times, and one case experienced severe pain after puncture. Conclusion:Compared with stereotactic-guided biopsy, DBT-guided biopsy can reduce operation time and exposure times, and can target more types of breast lesions, with higher clinical application value.

Objective:To explore the value of deep learning technology based on mammography in differentiating for breast imaging reporting and data system (BI-RADS) category 3 and 4 lesions.Methods:The clinical and imaging data of 305 patients with 314 lesions assessed as BI-RADS category 3 and 4 by mammography were analyzed retrospectively in Shenzhen People′s Hospital and Shenzhen Luohu People′s Hospital from January to December 2020. All 305 patients were female, aged 21 to 83 (47±12) years. Two general radiologists (general radiologist A and general radiologist B) with 5 and 6 years of work experience and two professional breast imaging diagnostic radiologists (professional radiologist A and professional radiologist B) with 21 years of work experience and specialized breast imaging training were randomly assigned to read the imaging independently at a 1∶1 ratio, and then to read the imaging again in combination with the deep learning system. Finally, breast lesions were reclassified into BI-RADS category 3 or 4. The receiver operating characteristic curve and area under the curve (AUC) were used to evaluate the diagnostic performance, and the differences of AUCs were compared by DeLong method.Results:The AUC of general radiologist A combined with deep learning system to reclassify BI-RADS category 3 and 4 breast lesions was significantly higher than that of general radiologist A alone (AUC=0.79, 0.63, Z=2.82, P=0.005, respectively). The AUC of general radiologist B combined with deep learning system to reclassify BI-RADS category 3 and 4 breast lesions was significantly higher than that of general radiologist B (AUC=0.83, 0.64, Z=3.32, P=0.001, respectively). There was no significant difference in the AUCs between professional radiologist A combined with deep learning system and professional radiologist A, and professional radiologist B combined with deep learning system and professional radiologist B in reclassifying BI-RADS category 3 and 4 breast lesions ( P>0.05). Conclusion:The deep learning system based on mammography is more effective in assisting general radiologists to differentiate between BI-RADS category 3 and 4 lesions.

Objective:To establish the predictive models for the prognosis of ductal carcinoma in situ (DCIS) at different pathological stages, and to evaluate the predictive performance of the models.Methods:Complete data of 273 patients with confirmed DCIS at different pathological stages who underwent mammography examination in Shenzhen People′s Hospital, Peking University Shenzhen Hospital and Shenzhen Luohu People′s Hospital from November 2014 to December 2020 were retrospectively collected, including 110 cases in the DCIS+ductal carcinoma in situ with microinvasion (DCIS-MI) group and 163 cases in the invasive ductal carcinoma (IDC)-DCIS group. The clinical, imaging and pathological features were analyzed. Mammary Mammo AI fusion model and deep learning-based natural language processing (NLP) structured diagnostic report model were used for image feature extraction. Patients in each group were randomly divided into training set and validation set with a ratio of 6∶4, and the predictors were screened by univariate and multivariate logistic regression analysis. The lowest Akaike information criterion value of each group was selected to construct the final predictive model. The receiver operating characteristic (ROC) curve was drawn to evaluate the performance of each model.Results:Taking estrogen receptor (-) or human epidermal growth factor receptor 2 (3+) as the poor prognostic reference, there were 62 cases considered with poor prognosis and 48 cases with good prognosis in DCIS+DCIS-MI group; while in the IDC-DCIS group, taking the Nottingham prognostic index as the reference, 33 cases were considered with poor prognosis, 73 cases with moderate prognosis, and 57 cases with good prognosis. Four predictive factors were screened to construct the DCIS+DCIS-MI-group predictive model, including DCIS nuclear grade, calcification with suspicious morphology in mammography, DCIS pathologic subtype and DCIS with microinvasion. Five predictive factors were screened to construct the IDC-DCIS-group predictive model, including neural or vascular invasion, Ki67 level, DCIS subtype, DCIS component proportion and associated features in mammography. The area under curve (AUC) for predicting poor prognosis of DCIS+DCIS-MI was 0.92 (95%CI 0.84-1.00) in the training set and 0.90 (95%CI 0.82-0.99) in the validation set; while the AUC for predicting poor prognosis of IDC-DCIS was 0.84 (95%CI 0.76-0.93) in the training set and 0.78 (95%CI 0.64-0.91) in the validation set.Conclusion:The developed models based on deep learning combined with NLP can effectively predict the prognosis of DCIS at different pathological stages, which are beneficial to the risk stratification of patients with DCIS, providing a reference for clinical decision.