OBJECTIVE: To investigate the effect and mechanism of Hyssopus cuspidatus Boriss. extract (HCE) in ovalbumin (OVA)-induced allergic asthma. METHODS: Components identification of HCE was conducted using ultra performance liquid chromatography-quadrupole-time of flight-mass spectrometry. Mice were sensitized with OVA to establish asthmatic model, and dexamethasone was used as positive control. Respiratory reactivity, white cells counting in bronchoalveolar lavage fluid and peripheral blood, cytokine level measurement in serum and lung tissue, and histologic examination were performed to evaluate the therapeutic effect of HCE on asthma. Network pharmacology approach was used for mechanism prediction. Western blotting and untargeted lipidomics method were applied for mechanism validation. RESULTS: Fifty-two compounds were identified in HCE, predominantly terpenoids and flavonoids. HCE markedly reduced airway resistance, the eosinophil infiltration in lung tissues, and the levels of immunoglobulin E, interleukin-4, interleukin-5, and interleukin-13. Network pharmacology analysis suggested phosphatidylinositol 3-kinases (PI3K), c-Jun N-terminal kinase (JNK), and p38 Mitogen-activated protein kinase (p38 MAPK) may be key proteins of HCE in the treatment of allergic asthma. Western blot results indicated that the levels of phosphorylated PI3K, JNK, and P38 were downregulated in HCE-treated group. Moreover, HCE significantly upregulated the levels of ceramide and sphingomyelin and downregulated the level of phosphatidylcholine. CONCLUSION HCE inhibited allergic asthma via PI3K/JNK/P38 signaling pathway and lipid homeostasis regulation.

Objective:To investigate the effect of applying the analog of miR-145(agomiR-145)to target a disintegrin and metallopro-tease 17(ADAM17)on the growth of triple-negative breast cancer(TNBC)xenograft tumors in nude mice and the underlying mecha-nism.Methods:A subcutaneous xenograft tumor model was established in nude mice with MDA-MB-231 cells(n=30).The 30 nude mice were randomly divided into agomiR group,agomiR-NC group,and control group to receive intratumoral injection of 100 μL of agomiR-145(0.33 g/L),agomiR-NC(0.33 g/L),and normal saline,respectively.We examined tumor tissue morphology with HE stain-ing;measured the expression of miR-145,ADAM17,and epidermal growth factor receptor(EGFR)in tumor tissues by real-time poly-merase chain reaction(RT-PCR);and determined the protein expression of ADAM17,EGFR,and p-EGFR in tumor tissues by immu-nohistochemistry and Western blot.Results:The tumor growth in the agomiR group was slow,with a significantly smaller tumor volume than those in the control group and the agomiR-NC group(P<0.05).The results of HE staining showed more severe necrosis and hem-orrhage within tumor tissues in the control group and the agomiR-NC group than in the agomiR group.RT-PCR results showed that the expression level of miR-145 in the agomiR group was significantly higher than those in the agomiR-NC and control groups(P<0.001);the agomiR group had a significantly lower mRNA expression level of ADAM17 than the other two groups(P<0.05);and there was no significant difference in EGFR mRNA expression between the three groups(P>0.05).Immunohistochemistry and Western blot detected significantly lower protein expression levels of ADAM17 and EGFR in the agomiR group than in the agomiR-NC group and the control group(P<0.05).Conclusion:AgomiR-145 inhibits the growth of TNBC xenografts in nude mice by targeted suppression of ADAM17-EGFR activity.

Objective:To establish a complete system for the isolation, purification, identification, and culture of Kaposiform hemangioendothelioma-derived endothelial cells (KHE-ECs), to analyze the phenotype of KHE-ECs, and to explore the possibility of establishing a KHE-EC bank.Methods:A novel digestion solution for KHE tumors (patent number: CN202410500224.2) was formulated using collection fluid, Liberase TM and dispase stock solutions, and was used to process tumor tissues to obtain cells. High-purity KHE-ECs were purified using CD31 + immunomagnetic beads. The EGM-2 complete medium containing 10% fetal bovine serum and 2% penicillin-streptomycin solution was employed for cell culture. To verify the characteristics of KHE-ECs, immunofluorescence assay was conducted to determine the expression of endothelial cell-specific markers CD31 and CD34, KHE disease markers podoplanin (D2-40), prospero-related homeobox 1 (Prox-1), and lymphatic vessel endothelial hyaluronan receptor 1 (LYVE1), as well as an infantile hemangioma-specific diagnostic marker glucose transporter 1 (GLUT-1). Human umbilical vein endothelial cells (HUVECs) served as controls for the phenotype analysis of KHE-ECs, including cell viability, cytoskeleton, proliferation, migration, invasion, tube formation, and sprouting ability. Results:Primary cells were successfully isolated from KHE tumor tissues, and high-purity KHE-ECs were obtained by using CD31 + immunomagnetic beads. The cells exhibited typical spindle-shaped morphology and an adherent growth pattern. Immunofluorescence assay showed that KHE-ECs expressed CD31, CD34, D2-40, Prox-1, and LYVE1, but did not express GLUT-1. There were significant differences in cell morphology, cell viability, and cytoskeletal structures between KHE-ECs and HUVECs. Additionally, the KHE-EC group showed significantly increased percentages of proliferative cells (29.1% ± 2.5%), numbers of migratory cells (114.3 ± 9.4) and invasive cells (110.0 ± 6.1), tube length (32 121.0 ± 892.0 μm), and number of sprouting cells (25.0 ± 3.6) compared with the HUVEC group (13.0% ± 2.2%, 38.0 ± 3.6, 35.3 ± 2.3, 25 345.0 ± 448.1 μm, 5.0 ± 1.0, respectively, all P ≤ 0.001) . Conclusion:An innovative digestion solution specifically for KHE tumors was formulated for the first time, and high-purity and well-growing KHE-EC strains were successfully isolated and purified by using the novel digestion solution in combination with CD31 + immunomagnetic beads, providing a stable and reliable cell source for subsequent experimental studies on KHE and laying the foundation for establishing a KHE-EC bank.

Objective:To establish a complete system for the isolation, purification, identification, and culture of Kaposiform hemangioendothelioma-derived endothelial cells (KHE-ECs), to analyze the phenotype of KHE-ECs, and to explore the possibility of establishing a KHE-EC bank.Methods:A novel digestion solution for KHE tumors (patent number: CN202410500224.2) was formulated using collection fluid, Liberase TM and dispase stock solutions, and was used to process tumor tissues to obtain cells. High-purity KHE-ECs were purified using CD31 + immunomagnetic beads. The EGM-2 complete medium containing 10% fetal bovine serum and 2% penicillin-streptomycin solution was employed for cell culture. To verify the characteristics of KHE-ECs, immunofluorescence assay was conducted to determine the expression of endothelial cell-specific markers CD31 and CD34, KHE disease markers podoplanin (D2-40), prospero-related homeobox 1 (Prox-1), and lymphatic vessel endothelial hyaluronan receptor 1 (LYVE1), as well as an infantile hemangioma-specific diagnostic marker glucose transporter 1 (GLUT-1). Human umbilical vein endothelial cells (HUVECs) served as controls for the phenotype analysis of KHE-ECs, including cell viability, cytoskeleton, proliferation, migration, invasion, tube formation, and sprouting ability. Results:Primary cells were successfully isolated from KHE tumor tissues, and high-purity KHE-ECs were obtained by using CD31 + immunomagnetic beads. The cells exhibited typical spindle-shaped morphology and an adherent growth pattern. Immunofluorescence assay showed that KHE-ECs expressed CD31, CD34, D2-40, Prox-1, and LYVE1, but did not express GLUT-1. There were significant differences in cell morphology, cell viability, and cytoskeletal structures between KHE-ECs and HUVECs. Additionally, the KHE-EC group showed significantly increased percentages of proliferative cells (29.1% ± 2.5%), numbers of migratory cells (114.3 ± 9.4) and invasive cells (110.0 ± 6.1), tube length (32 121.0 ± 892.0 μm), and number of sprouting cells (25.0 ± 3.6) compared with the HUVEC group (13.0% ± 2.2%, 38.0 ± 3.6, 35.3 ± 2.3, 25 345.0 ± 448.1 μm, 5.0 ± 1.0, respectively, all P ≤ 0.001) . Conclusion:An innovative digestion solution specifically for KHE tumors was formulated for the first time, and high-purity and well-growing KHE-EC strains were successfully isolated and purified by using the novel digestion solution in combination with CD31 + immunomagnetic beads, providing a stable and reliable cell source for subsequent experimental studies on KHE and laying the foundation for establishing a KHE-EC bank.

Objective:To analyze the impact of cecal ligation and puncture (CLP)-induced sepsis on the proliferation and differentiation of intestinal epithelial cells.Methods:① Animal experiment: sixteen male C57BL/6 mice were divided into sham operation group (Sham group) and CLP-induced sepsis model group (CLP group) by random number table method, with 8 mice in each group. After 5 days of operation, the jejunal tissues were taken for determination of leucine-rich-repeat-containing G-protein-coupled receptor 5 (LGR5) and intestinal alkaline phosphatase (IAP) by polymerase chain reaction (PCR). The translation of LGR5 was detected by Western blotting. The expression of proliferating cell nuclear antigen (Ki67) was analyzed by immunohistochemistry. IAP level was detected by modified calcium cobalt staining and colorimetry. Immunofluorescence staining was used to detect the expression of Paneth cell marker molecule lysozyme 1 (LYZ1) and goblet cell marker molecule mucin 2 (MUC2). ② Cell experiment: IEC6 cells in logarithmic growth stage were divided into blank control group and lipopolysaccharide (LPS) group (LPS 5 μg/mL). Twenty-four hours after treatment, PCR and Western blotting were used to analyze the transcription and translation of LGR5. The proliferation of IEC6 cells were detected by 5-ethynyl-2'-deoxyuridine (EdU) staining. The transcription and translation of IAP were detected by PCR and colorimetric method respectively.Results:① Animal experiment: the immunohistochemical results showed that the positive rate of Ki67 staining in the jejunal tissue of CLP group was lower than that of Sham group [(41.7±2.5)% vs. (48.7±1.4)%, P = 0.01]. PCR and Western blotting results showed that there were no statistical differences in the mRNA and protein expressions of LGR5 in the jejunal tissue between the CLP group and Sham group (Lgr5 mRNA: 0.7±0.1 vs. 1.0±0.2, P = 0.11; LGR5/β-actin: 0.83±0.17 vs. 0.68±0.19, P = 0.24). The mRNA (0.4±0.1 vs. 1.0±0.1, P < 0.01) and protein (U/g: 47.3±6.0 vs. 73.1±15.3, P < 0.01) levels of IAP in the jejunal tissue were lower in CLP group. Immunofluorescence saining analysis showed that the expressions of LYZ1 and MUC2 in the CLP group were lower than those in the Sham group. ②Cell experiment: PCR and Western blotting results showed that there was no significant difference in the expression of LGR5 between the LPS group and the blank control group (Lgr5 mRNA: 0.9±0.1 vs. 1.0±0.2, P = 0.33; LGR5/β-actin: 0.71±0.18 vs. 0.69±0.04, P = 0.81). The proliferation rate of IEC6 cells in the LPS group was lower than that in the blank control group, but there was no significant difference [positivity rate of EdU: (40.5±3.8)% vs. (46.5±3.6)%, P = 0.11]. The mRNA (0.5±0.1 vs. 1.0±0.2, P < 0.01) and protein (U/g: 15.0±4.0 vs. 41.2±10.4, P < 0.01) of IAP in the LPS group were lower than those in the blank control group. Conclusion:CLP-induced sepsis inhibits the proliferation and differentiation of intestinal epithelial cells, impairing the self-renewal ability of intestinal epithelium.

Objective To analyze the red blood cell lifespan and its influencing factors in patients with acute leukemia (AL) under different disease states.Methods A total of 142 cases of patients with AL admit-ted to the department of hematology in the First Hospital of Lanzhou University from January 2022 to June 2023 were selected as the research subjects,and their red blood cell lifespan and other clinical data were col-lected.The red blood cell lifespan were compared among patients with AL under different disease states. Spearman correlation analysis was used to determine the correlation between the red blood cell lifespan with age and other laboratory indicators,and univariate analysis and multiple linear regression analysis were used to explore the influencing factors of red blood cell lifespan in patients with AL.Results Among the 142 cases of AL,there were 33 newly diagnosed cases,19 relapsed cases,8 cases of partial response+no response,and 82 cases of complete response.The red blood cell lifespan was 33.0 (9.0-147.0),52.0 (15.0-115.0),20.5 (12.0-46.0),50.0 (11.0-186.0) d,respectively.The red cell lifespan of newly diagnosed patients and pa-tients with partial response+no response was shorter than that of patients with complete response,and the differences were statistically significant (Z=-3.933,P<0.001;Z=-3.586,P=0.002).Fifteen newly di-agnosed AL patients achieved complete response after treatment,and the red blood cell lifespan was signifi-cantly prolonged compared with that at initial diagnosis[42 (14-101) d vs. 27 (9-68) d,Z=-2.179,P=0.029].The results of correlation analysis showed that the red blood cell lifespan was positively correlated with white blood cell count,red blood cell count,hemoglobin level and platelet count (P<0.05),and nega-tively correlated with blast cell count and erythropoietin level (P<0.05).The results of multiple linear re-gression analysis showed that the chromosomal abnormalities in karyotypes,FMS-like tyrosine kinase 3 (FLT3),RUNX1 gene mutations and blood transfusion were the influencing factors of red blood cell lifespan in patients with AL (B=-11.151,-24.969,-30.838,-18.784,P<0.05).Conclusion The red blood cell lifespan in patients with AL is shortened under different disease states,which could obtain a certain degree of recovery after achieving complete response after treatment (but still below the normal reference range).The red blood cell lifespan in patients with AL is related to chromosomal abnormalities in karyotypes,FLT3,RUNX1 gene mutations and short term blood transfusion.

Objective:To characterize the longitudinal and dynamic high-density lipoprotein (HDL) trajectories in critically ill children and explore their correlation with clinical outcomes.Methods:Retrospective cohort study.All critically ill children admitted to the Pediatric Intensive Care Unit (PICU) of West China Hospital, Sichuan University from January 1, 2015 to October 1, 2020 were included in this retrospective study.Group-based trajectory modeling (GBTM) was applied to characterize the HDL trajectories in days 0-6 post-PICU admission and develop HDL trajectory groups.The in-hospital mortality rate was reported as frequency (%) and then compared by the Chi-square test or Fisher′s exact test between HDL trajectory groups.The length of stay (LOS) in the PICU was described by M( Q1, Q3), and its difference between HDL trajectory groups was evaluated by the Kruskal Wallis test.Logistic regression and multiple linear regression were used to determine the correlation between HDL trajectories and clinical outcomes.The primary outcome was in-hospital mortality rate, and the secondary outcome was LOS in the PICU. Results:A total of 4 384 critically ill children were ultimately enrolled in the study, and 6 HDL trajectory groups were developed based on GBTM analyses: group 1 (758 cases), the lowest HDL group; group 2 (1 413 cases), the low HDL group; group 3 (74 cases), the low-to-high HDL group; group 4 (621 cases), the medium HDL group; group 5 (1 371 cases), the high HDL group; and group 6 (147 cases), the highest HDL group.Logistic regression analysis showed that compared with critically ill children in group 1, those belonging to groups 2, 3, 4, 5, and 6 were at lower risks of in-hospital mortality with odds ratio ( OR): 0.475, 95%confidence interval ( CI): 0.352-0.641, P<0.001; OR: 0.093, 95% CI: 0.013-0.679, P=0.019; OR: 0.322, 95% CI: 0.208-0.479, P<0.001; OR: 0.263, 95% CI: 0.185-0.374, P<0.001, and OR: 0.142, 95% CI: 0.044-0.454, P=0.001, respectively.Multiple linear regression analysis revealed that compared with critically ill children in group 1, those belonging to groups 4, 5, and 6 had the trend of shorter LOS in PICU, and the β value and 95% CI were β: -4.332, 95% CI: -5.238- -3.426, P<0.001; β: -3.053, 95% CI: -3.809--2.297, P<0.001; β: -6.281, 95% CI: -7.842--4.721, P<0.001, respectively. Conclusions:The dynamic HDL trajectories during 0-6 days after PICU admission are associated with in-hospital mortality rate of critically ill children.The HDL trajectory at a persistently low level is associated with higher mortality, while the HDL trajectory at a persistently high level or with the trend from a low level rising to a high level shows a lower risk of mortality.It is suggested that the HDL trajectory model may become an indicator to predict the condition and prognosis of critically ill children.

Objective:To investigate the clinical effectiveness of Piezocision combined with a microporous technique in accelerating periodontal tissue reconstruction during the anterior migration of mandibular molars in adults.Methods:A prospective, randomized, controlled study was conducted on 30 adult orthodontic patients recruited from Shaoxing Hospital of Traditional Chinese Medicine between January 2020 and September 2022. The inclusion criteria were patients who were unable to retain their first molars due to severe caries or long-term absence and were not suitable for implantation. Using the random number table method, the patients were randomly assigned to two groups: a simple orthodontic control group (Group A, n = 15) and a group that received Piezocision combined with a microporous technique (Group B, n = 15). After treatment, a comparison was made between the two groups in terms of mesial movement distance of the mandibular second molar, plaque index, gingival index, periodontal pocket depth, width of keratinized gingiva, gingival recession, clinical attachment loss, mesial root resorption of the mandibular second molar, alveolar bone height (measured as the distance from the center of the lower incisor to the anterior margin of the chin, referred to as the LM-AC distance), mandibular bone height (measured by the distance from the distal or mesial surface of the root to the alveolar bone margin, denoted as the CEJ-AC distance), and orthodontic satisfaction. Results:The mesial movement distances of the mandibular second molar in Group A patients were (0.86 ± 0.13) mm, (2.75 ± 0.24) mm, (3.54 ± 0.24) mm, and (4.67 ± 0.13) mm at 4, 6, 8, and 12 weeks, respectively. These values were significantly greater than those observed in Group B, which were (0.43 ± 0.06) mm, (1.27 ± 0.14) mm, (1.85 ± 0.53) mm, and (2.65 ± 0.06) mm ( t = 6.83, 14.13, 18.24, 23.78, all P < 0.001). Prior to treatment, there were no statistically significant differences in plaque index, gingival index, periodontal pocket depth, width of keratinized gingiva, gingival recession, or clinical attachment loss between the two groups (all P > 0.05). After treatment, Group A did not exhibit statistically significant differences in plaque index, gingival index, width of keratinized gingiva, and gingival recession compared with baseline values (all P > 0.05). However, in Group A, periodontal pocket depth and clinical attachment loss significantly increased compared with pretreatment levels ( t = -2.57, -7.50, both P < 0.05). After treatment, Group B exhibited significantly increased values for periodontal pocket depth, width of keratinized gingiva, gingival recession, and clinical attachment loss compared with baseline levels ( t = -8.66, -5.57, -45.33, -9.72, all P < 0.001). Furthermore, these values were significantly higher in Group B compared with those in Group A ( t = -4.28, -3.18, 10.00, 10.69, all P < 0.001). A statistically significant difference was also observed between the two groups in terms of mesial root resorption of the mandibular second molar ( t = 4.14, P < 0.001). However, there was no statistically significant difference in LM-AC distance between the two groups after treatment ( P > 0.05). Conclusion:The combination of Piezocision and a microporous technique can effectively accelerate the anterior migration of mandibular molars in adults while maintaining the health of periodontal tissues. This approach holds great potential for clinical promotion.

A 55-year-old male patient was treated with tacrolimus (2.5 mg in the morning and 2 mg at night) for 6 months after lung transplantation to prevent rejection. The blood trough concentration of tacrolimus was stable at 8.0-10.0 μg/L. The patient received antiviral treatment due to corona virus disease 2019 (nirmatrelvir/ritonavir 300 mg/100 mg twice daily orally for a total of 5 days). During the antiviral treatment, the patient continued the anti-rejection treatment. On the second day of antiviral treatment, the patient′s blood trough concentration of tacrolimus increased to >40.0 μg/L, which was considered to be caused by the interaction between nirmatrelvir/ritonavir and tacrolimus. Tacrolimus was withdrawn and antiviral therapy was continued. After discontinuation of tacrolimus for 8 days and nirmatrelvir/ritonavir for 3 days, the blood trough concentration of tacrolimus decreased to 25.7 μg/L. After re-giving tacrolimus at reducing dosage for 3 days, the blood trough concentration of tacrolimus was 8.3 μg/L. After 13 days of resuming administration of tacrolimus at the original dose and frequency, the patient′s blood trough concentration of tacrolimus was 9.2 μg/L. Since then, the blood trough concentration of tacrolimus in the patient was not abnormal again.

A 55-year-old male patient was treated with tacrolimus (2.5 mg in the morning and 2 mg at night) for 6 months after lung transplantation to prevent rejection. The blood trough concentration of tacrolimus was stable at 8.0-10.0 μg/L. The patient received antiviral treatment due to corona virus disease 2019 (nirmatrelvir/ritonavir 300 mg/100 mg twice daily orally for a total of 5 days). During the antiviral treatment, the patient continued the anti-rejection treatment. On the second day of antiviral treatment, the patient′s blood trough concentration of tacrolimus increased to >40.0 μg/L, which was considered to be caused by the interaction between nirmatrelvir/ritonavir and tacrolimus. Tacrolimus was withdrawn and antiviral therapy was continued. After discontinuation of tacrolimus for 8 days and nirmatrelvir/ritonavir for 3 days, the blood trough concentration of tacrolimus decreased to 25.7 μg/L. After re-giving tacrolimus at reducing dosage for 3 days, the blood trough concentration of tacrolimus was 8.3 μg/L. After 13 days of resuming administration of tacrolimus at the original dose and frequency, the patient′s blood trough concentration of tacrolimus was 9.2 μg/L. Since then, the blood trough concentration of tacrolimus in the patient was not abnormal again.