Objective To investigate the expression of dipeptidyl peptidase 4 (DPP4) and angiotensin-converting enzyme 2 (ACE2) in lung tissues of patients with four different diseases including coronavirus disease 2019 (COVID-19), chronic obstructive pulmonary disease (COPD), pulmonary sarcoidosis and pulmonary bullae, and to find out the potential risk factors affecting COVID-19. Methods This study retrospectively analyzed the clinical data of 40 patients admitted to Renmin Hospital of Wuhan University with COVID-19 (COVID-19 group), COPD (COPD group), pulmonary sarcoidosis (pulmonary sarcoidosis group) and pulmonary bullae (pulmonary bullae group) and surgically resected paraffin-embedded pathological lung tissues were obtained from their lung tissue pathological specimens after surgery and paraffin embedding. The GEO database (https://www.ncbi.nlm.nih.gov/geo/) was used for bioinformatics analysis to explore the expression difference of DPP4 and ACE2 mRNA in COVID-19, COPD, pulmonary sarcoidosis and normal lung tissues. Immunohistochemistry method was used to detect the expression of DPP4 and ACE2 protein in lung tissues of each group and the average optical density was measured by image analysis software. Results The results of GEO database analysis showed that compared with pulmonary bullae group, the expression level of DPP4 mRNA had no significant difference in the COPD group and pulmonary sarcoidosis group (both P>0.05), but it was increased in the COVID-19 group (P<0.05); There was no significant difference in the expression level of ACE mRNA in the pulmonary sarcoidosis group (P>0.05), but it was increased in the lung tissue of COVID-19 group and COPD group (both P<0.05). The results of immunohistochemistry showed that DPP4 and ACE2 proteins were lowly expressed in the pulmonary sarcoidosis group and pulmonary bullae group, while their expression level was high in COVID-19 and COPD groups without significant difference (P>0.05). The expression of DPP4 and ACE2 proteins in COVID-19 group was not related to the patient’s gender and age (P>0.05), but was related to smoking and long smoking duration (P<0.05), and there was a positive correlation between DPP4 and ACE2 expression (P<0.05). Conclusions DPP4 and ACE2 proteins are lowly expressed in the pulmonary sarcoidosis group and pulmonary bullae group, while their expression level is high in COVID-19 and COPD groups. There is no significant difference in the expression level of DPP4 and ACE2 protein in the COVID-19 and COPD lung tissues. There may be a positive correlation between DPP4 and ACE2 proteins expression in lung tissue, and smoking may be a potential risk factor for COVID-19.
Objective In this study, we present a living biobank of patient-derived tumoroids from advanced colorectal cancer (CRC) patients and show examples of how these tumoroids can be used to simulate cancer behavior ex vivo and provide more evidence for tumoroids could be utilized as a predictive platform during chemotherapy to identify the chemotherapy response. Methods The tumor tissues of CRC patients were collected to isolate and culture tumoroids, and the histomorphology of tumoroids was evaluated. Further, tumoroids were treated with drugs of different chemotherapy schemes, and the drug sensitivity of tumoroids was evaluated by using CellTiter-GIo 3D cell viability assay, and the clinical efficacy was compared with that of patients. Results The tumoroids were still highly consistent with the original tumor histomorphology after continuous passage. The consistency between the drug sensitivity of tumoroids from different patients and the clinical efficacy of corresponding CRC patients was 91.18% (31/34). The drug inhibition rate of tumoroids was positively correlated with the progression free survival (PFS) of CRC patients (rs=0.412, P=0.016), while the area under the cell activity drug concentration curve of tumoroids was negatively correlated with the PFS of CRC patients (rs=–0.479, P=0.004). Conclusion This study established a biological sample bank of tumoroids for CRC patients, and suggested that tumoroids had the potential to be used as preclinical experimental models and predict the chemotherapy effect of CRC patients.
Objective To explore the influence and mechanism of mechanistic target of rapamycin kinase (mTOR)/ receptor of advanced glycation end products (RAGE) pathway mediated-ferritinophagy on high glucose consumption promoting invasion and migration of colorectal cancer (CRC). Methods① Patients and tissue samples. Clinical data and tissues were collected from CRC patients underwent surgery and completed the dietary questionnaire in the Second Affiliated Hospital of Harbin Medical University between October 2022 and October 2023. Real-time quantitative reverse transcription PCR (qRT-PCR) was used to analyzed the expression of nuclear receptor coactivator 4 (NCOA4) and ferritin in CRC and para-carcinoma tissues.② Cell culture and treatment. The HT29 and HCT116 cells were treated by RPMI1640 medium containing 0, 35, 70, 105, 140 mmol/L glucose, and cell counting kit-8 (CCK-8) and lactate dehydrogenase (LDH) activity analysis were performed to confirm 105 mmol/L glucose was the optimal concentration in the current study. Then the HT29 and HCT116 cells were randomly divided into: control group, glucose group; control group, glucose group, si-RAGE group, and glucose+si-RAGE group; control group, glucose group, rapamycin group, and glucose+rapamycin group. Untreated HT29 and HCT116 cells were considered as control group. The cells in glucose group were treated with 105 mmol/L glucose for 48 h. The CRC cells in si-RAGE group were transfected with si-RAGE for 6 h. The CRC cells in rapamycin group were treated with 10 nmol/L rapamycin for 48 h. The CRC cells in glucose+si-RAGE group were treated with 105 mmol/L glucose for 48 h combination transfected with si-RAGE for 6 h. The CRC cells in glucose+rapamycin group were treated with 105 mmol/L glucose for 48 h combination treated with 10 nmol/L rapamycin for 48 h. Then electron microscopy and western blot, wound healing assay and transwell assay were exhibited, respectively.③ Azoxymethane (AOM)-induced CRC rat model. The effects of glucose consumption on malignant behavior and ferritinophagy mediated by mTOR/RAGE pathway were evaluated in AOM-induced CRC rat models. A total of 16 rats were randomly divided into control group and glucose group, the colorectal tumor number was record and HE staining of colorectal tumor tissues was further performed. The expression of RAGE, mTOR, NCOA4, and ferritin in colorectal tissues of rats from each group was detected by qRT-PCR. Results① More lymphatic node metastasis and TNM Ⅲ/Ⅳ stages was observed in CRC patients from high glucose consumption group (P=0.004, P=0.004). Moreover, we confirmed that NCOA4 expression was significantly decreased (P<0.001) while ferritin was significantly increased (P<0.001) in CRC tissues especially in the CRC tissues from patients with positive lymph nodes metastasis. Additionally, high glucose consumption of CRC patients was negatively correlated with ferritinophagy flux. ② High glucose treatment significantly decreased autophagosomes in HT29 and HCT116 cells while si-RAGE transfection increased autophagic vacuoles compared to the control group. When compared with the glucose group, autophagosomes were increased in the glucose+si-RAGE group. Moreover, compared to the control group, the expression of RAGE, p-mTOR, and ferritin was increased (P<0.001) while the expression of NCOA4 was decreased (P<0.001) in glucose group, but the expression of RAGE, p-mTOR and ferritin was decreased (P<0.001) while the expression of NCOA4 was increased (P<0.001) in si-RAGE group; when compared with the glucose group, the expression of RAGE, p-mTOR and ferritin was downregulated (P<0.001) while the expression of NCOA4 was upregulated (P<0.001) in HT29 and HCT116 cells from the glucose+siRAGE group. Compared to the control group, the HT29 and HCT116 cells in the glucose group performed enhanced wound scratch healing and migration, invasion viability (P<0.05); but the HT29 and HCT116 cells in the si-RAGE group performed impaired wound scratch healing and migration, invasion viability (P<0.05). When compared with the glucose group, the HT29 and HCT116 cells in the glucose+si-RAGE group performed impaired wound scratch healing and migration, invasion viability (P<0.05).③ Rapamycin treatment significantly inhibited the expression of RAGE, p-mTOR and ferritin (P<0.05) but induced the expression of NCOA4 (P<0.05) compared to the control group. When compared with the glucose group, the expression of RAGE, p-mTOR and ferritin was downregulated (P<0.05) while the expression of NCOA4 was upregulated (P<0.05) in HT29 and HCT116 cells from the glucose+rapamycin group. Additionally, compared to the control group, rapamycin treatment performed inhibited effect on wound scratch healing and migration, invasion viability in the HT29 and HCT116 cells (P<0.05); while the HT29 and HCT116 cells in the glucose+rapamycin group performed impaired wound scratch healing and migration, invasion viability (P<0.05) when compared with the glucose group.④ In the AOM induced CRC rat model, we found the more colorectal tumors, aggravated cellular pleomorphism and upregulate expression of RAGE, p-mTOR, ferritin (P<0.05) while downregulated expression of NCOA4 (P<0.05) in the control group than those of the glucose group. ConclusionHigh glucose consumption promote invasion and migration in CRC through suppressing ferritinophagy via activating the mTOR/RAGE pathway.