BACKGROUND: P16 inactivation is frequently accompanied by telomerase reverse transcriptase ( TERT ) amplification in human cancer genomes. P16 inactivation by DNA methylation often occurs automatically during immortalization of normal cells by TERT . However, direct evidence remains to be obtained to support the causal effect of epigenetic changes, such as P16 methylation, on cancer development. This study aimed to provide experimental evidence that P16 methylation directly drives cancer development. METHODS: A zinc finger protein-based P16 -specific DNA methyltransferase (P16-Dnmt) vector containing a "Tet-On" switch was used to induce extensive methylation of P16 CpG islands in normal human fibroblast CCD-18Co cells. Battery assays were used to evaluate cell immortalization and transformation throughout their lifespan. Cell subcloning and DNA barcoding were used to track the diversity of cell evolution. RESULTS: Leaking P16-Dnmt expression (without doxycycline-induction) could specifically inactivate P16 expression by DNA methylation. P16 methylation only promoted proliferation and prolonged lifespan but did not induce immortalization of CCD-18Co cells. Notably, cell immortalization, loss of contact inhibition, and anchorage-independent growth were always prevalent in P16-Dnmt&TERT cells, indicating cell transformation. In contrast, almost all TERT cells died in the replicative crisis. Only a few TERT cells recovered from the crisis, in which spontaneous P16 inactivation by DNA methylation occurred. Furthermore, the subclone formation capacity of P16-Dnmt&TERT cells was two-fold that of TERT cells. DNA barcoding analysis showed that the diversity of the P16-Dnmt&TERT cell population was much greater than that of the TERT cell population. CONCLUSION P16 methylation drives TERT -mediated immortalization and transformation of normal human cells that may contribute to cancer development.
Humans ; Telomerase/genetics* ; DNA Methylation/physiology* ; Fibroblasts/cytology* ; Cyclin-Dependent Kinase Inhibitor p16/metabolism* ; Cell Line ; Cell Transformation, Neoplastic/genetics*

Objective: To explore the network structure of dietary intake and physical fitness subtypes among junior and senior high school students, so as to provide precise and efficient intervention guidance for improving abnormal physical health issues in adolescents. Methods: Based on the 2020-2021 Database of Youth Health (DYH), a total of 9 730 junior and senior high school students in Shandong Province were included for analysis. Dietary intake was assessed using a modified Chinese Dietary Quality Questionnaire, and physical fitness was evaluated according to the "2014 National Student Physical Fitness Standard". The Kmeans clustering algorithm was employed to identify potential subtypes of physical fitness in junior and high school students. Network analysis was used to construct a network linking dietary intake and physical fitness symptoms and to identify clusters of dietary behaviors and physical fitness symptoms among adolescents of different genders. Results: The physical fitness among junior and senior high school students of different genders were categorized into two subtypes: the baseline steady type (3 427 male students, 4 039 female students) and the morphological deviation type (1 294 male students, 970 female students). Statistically significant differences were observed in network strength and network connectivity strength among different genders and physical fitness subtypes (all P<0.05). Connections between food intake and physical health symptom clusters across different genders and physical health types among junior and senior high school students were primarily achieved through instant noodle intake and physical fitness (males of morphological deviation type, weight=0.06), fruit intake and physical fitness (males of baseline steady type, weight=-0.07), potato intake and vital capacity (females of morphological deviation type, weight=0.09), and processed meat intake and vital capacity (females of baseline steady type, weight=0.05). Conclusions Dietary intake serves as a significant modifiable risk factor for the physical fitness of junior and high school students. Interventions should focus on promoting healthy eating habits.

Objective:To investigate the facilitators and hindrances of smoking cessation in patients with chronic obstructive pulmonary disease (COPD), and to provide a basis for developing individualized smoking cessation intervention strategies for COPD patients.Methods:Based on the health ecology theory and using a phenomenological approach in qualitative research, purposive sampling was used to select 15 COPD patients with smoking history who were admitted to Tianjin Fourth Central Hospital from March to May 2023 and face-to-face semi-structured interviews were conducted. Colaizzi seven-step analysis was used to analyze the interview content.Results:A total of 15 COPD patients were interviewed, including 13 males and 2 females, aged 61-75 years old. The facilitators of smoking cessation in COPD patients included complications, sequelae of novel coronavirus infection, fear of death, smoking cessation counseling by medical staff, heavy family financial burden, and smoke-free environment. The hindrances of smoking cessation in COPD patients included milder disease symptoms of COPD, higher levels of nicotine dependence, false disease-related perceptions, family supervision and control, and occupational experience.Conclusions:Smoking cessation in COPD patients is influenced by five factors: personal characteristics, behavioral characteristics, interpersonal network, living and working conditions, and policy environment. Medical staff and relevant national institutions should formulate corresponding smoking cessation strategies according to address the facilitators and hindrances of smoking cessation in COPD patients, so as to further reduce the smoking prevalence of COPD patients, promote the health of patients and reduce the burden of disease.

OBJECTIVE To explore the action mechanism of kushenol F （KSCF） in treating ulcerative colitis （UC） in mice. METHODS The potential targets of KSCF intervening in UC were predicted with network pharmacology and molecular docking. C57BL/6J mice were randomly divided by body weight into model group， positive control group （sulfasalazine， 703 mg/kg）， KSCF group （100 mg/kg）， and normal group， with 6 mice per group. The UC model of mice was induced by dextran sulfate sodium solution. During the modeling period， the mice were given relevant medicine intragastrically， once a day， for 7 consecutive days. After the last administration， the disease activity index （DAI） of the mice was scored； the length of the mice’s colon was measured； pathological changes in the colon tissue of mice were observed； the levels of lipopolysaccharide （LPS） in serum， myeloperoxidase （MPO）， nitric oxide （NO） and superoxide dismutase （SOD） in the colon were detected in mice； the expression levels of occludin and ZO-1 in colon tissue of mice were detected； the proportions of CD3+T， CD4+T， and CD8+T lymphocytes in the spleen and the ratio of CD4+/CD8+ were detected； changes in colonic microbiota were analyzed by 16S rDNA sequencing. RESULTS Results of network pharmacology indicated that KSCF may treat UC by regulating signaling pathways such as phosphatidylinositol-3 kinase/protein kinase B （PI3K/AKT） and nuclear factor kappa B （NF- κB）. Molecular docking results showed that KSCF bound most stably with NF-κB p65 protein. Animal experiment results demonstrated that， compared with the model group， the pathological characteristics of colon tissue in mice were improved in KSCF group. DAI scores， serum levels of LPS， the levels of MPO，NF-κB p65 phosphorylation and NLRP3 protein expression in the colon， and the proportion of CD8+T lymphocytes in the spleen were reduced significantly （P＜0.05）. Body weight， SOD levels， expression levels of occludin and ZO-1 in the colon， proportions of CD3+T and CD4+T lymphocytes， and the CD4+/CD8+ ratio in the spleen were significantly increased （P＜0.05）； the abundance of Firmicutes， Actinobacteria， Akkermansia， and Lactobacillus genera were increased， while Proteobacteria decreased； the microbial community structure tended towards that of the normal group. CONCLUSIONS KSCF alleviates UC by restoring intestinal microbial imbalance， enhancing immune response， and inhibiting colonic inflammatory responses， thereby improving intestinal barrier integrity.

Objective　To explore the relationship between serum cystatin C (Cys-C),uric acid (UA)，CD4+T cells and CD8+T cells with intracranial artery stenosis and cerebral infarction and analyze their influencing factors.Methods 　The clinical data of 124 patients with cerebral infarction who were hospitalized in our hospital from June 2020 to September 2021 were collected.All patients were given Brain MRA or /and CTA and routine biochemical examinations.According to the degree of proximal stenosis of intracranial large vessels，patients were divided into no stenosis group (36cases),mild stenosis group (43 cases),and moderate to severe stenosis group (45 cases).The clinical data of the three groups of patients and the differences in serum Cys-C,UA,and T cell subsets were compared.The factors affecting intracranial artery stenosis in cerebral infarction were analyzed by multivariate Logistic regression.The ROC curve was used to evaluate the value of serum CysC,UA and T cell subsets in patients with the cerebral infarction intracranial artery stenosis narrow.Results　The serum levels of Cys-C,UA,CD4+T cells were lower in no stenosis group than those in mild stenosis group and those in medium-severe stenosis group,and positively correlated with cerebral infarction intracranial artery stenosis.The serum levels of CD8+T cells were higher in no stenosis group than those in mild stenosis group and those in medium-severe stenosis group,and were negatively correlated with cerebral infarction intracranial artery stenosis.Conclusion　The increase of serum Cys- C,UA,CD4+ T cells and the decrease of CD8+T cells are risk factors for intracranial artery stenosis in cerebral infarction.As the degree of intracranial artery stenosis worsens,Cys-C,UA,CD4+T cells increase,and CD8+T cells decrease.

Objective: To explore the role and mechanism of long noncoding RNA(lncRNA) AC005062.1 in colorectal cancer(CRC) cell proliferation and apoptosis. Methods: The analysis of GSE84983 and GSE 104364 was used to indicate the expression level lncRNA AC005062.1 in CRC. The tumor tissues(tumor group) and adjacent tissues(control group) of patients undergoing CRC surgery in the hospital tumor surgery department were collected. Eight pairs of tissues were randomly selected, and qPCR was used to detect the expression of lncRNA AC005062.1 in CRC tissues and paired adjacent tissues in the two groups. After using siRNA to down-regulate the expression of lncRNA AC005062.1 in CRC cells, CCK-8 assay was used to detect cell proliferation in two groups, flow cytometry was used to detect cell cycle and apoptosis in two groups, wound scratch test was used to detect cell migration in two groups, and the lncRNA was determined. The database was used to compare the localization of lncRNA AC005062.1 and MACC1 genes on staining. The expression levels of MACC1 transcription and protein levels in the control group and tumor group were detected by qPCR and Western blot, respectively. The lncRNA AC005062.1 in CRC cells was down-regulated, and the expression levels of MACC1 transcription and protein levels in the two groups of cells were detected by qPCR and Western blot, respectively. Results: The database analysis of GSE84983 and GSE104364 and the detection results of the two groups indicated that the lncRNA AC005062.1 was highly expressed in CRC. After down-regulation of lncRNA AC005062.1, the results of CCK-8 experiments showed that the proliferation rate of HT29 cells in the down-regulation group decreased. Flow cytometry showed that the number of apoptosis of HT29 cells in the down-regulated group increased, the proportion of G1 phase increased, and the proportion of S phase and G2 phase decreased. Western blot experiments showed that the expressions of cleaved-caspase3 and Bax in the down-regulated HT29 cells increased, while the expression of Bcl-2 decreased. Wound scratch experiments showed that the migration rate of HT29 cells in the down-regulated group was lower than that in the control group. The analysis of UCSC to compare the location of lncRNA ac005062.1 and MACC1 on chromosomes showed that they were located very close together. MACC1 was highly expressed in CRC and down-regulated lncRNA AC005062.1 expression in HT29 cells could reduce the expression of MACC1. Conclusion lncRNA AC005062.1 can inhibit the proliferation, cycle and migration of HT29 cells, and promote its apoptosis by regulating the expression of MACC1 in CRC.

Objective:To investigate the image characteristics of woven coronary artery (WCA)on intravascular ultrasound(IVUS) and optical coherence tomography(OCT).Methods:Thirty-seven patients suspected of WCA on coronary angiography were enrolled from Teaching Hospital of Chengdu University of Traditional Chinese Medcine, Zhengzhou Cardiovascular Disease Hospital and Zhongshan Hospital of Fudan University from January 2013 to July 2020. The intraluminal imaging features of WCA were analyzed using IVUS and OCT.Results:Of the 37 patients admitted at the cardiology service, 9 patients had WCA. All the patients underwent coronary angiography, IVUS and OCT, of which 6 lesions were located on the right coronary artery, 2 lesions were located on the left anterior descending artery and 1 patient had WCA on the circumflex artery. The mean length of WCA lesions was 2.2 cm(ranged from 1.2 cm to 4.5 cm). The angiographic appearance of WCA was numerous small tortious channels origined form the main lumen. The channels appeared to be " doughnut" like pattern and they merged to normal artery again after the anomalous segment. Flow limitation was rare unless there was coronary atherosclerosis. OCT and IVUS showed multiple spiral channels in the anomalous segment, which were independent of each other and each channels had a relatively complete three-layers vascular structure.Conclusions:With typical image characteristics, IVUS and OCT are able to screen out WCA and guide the treatment decision making.

Objective To study the relationships between pathological grade of Henoch-Schonlein purpura nephritis (HSPN) and levels of serum transforming growth factor-beta 1 (TGF-β1),monocyte chemoattractant protein 1 (MCP-1),interleukin (IL)-17 and prognosis in adults.Methods 98 HSPN patients treated in our hospital from June 2015 to December 2017 were selected as the study group,65 IgA nephritis patients were selected as the IgA nephritis group,and 60 healthy people who came to our hospital for physical examination during the same period were selected as the control group.The levels of TGF-β1,MCP-1 and IL-17 in serum of the three groups were detected,and the Cox regression analysis was used to analyze the risk factors affecting the patient's condition.Results The levels of serum TGF-β1,MCP-1 and IL-17 in the study group were significantly higher than those in IgA nephritis group and control group (P ＜ 0.05).The levels of serum TGF-β1,MCP-1 and IL-17 in IgA nephritis group were significantly higher than those in control group (P ＜ 0.05).There was no significant difference in age,sex,hemoglobin,albumin,urinary protein and renal phenotype among groups (P ＜ 0.05).Platelets of type Ⅲ were significantly lower than those of type Ⅱ (P ＜0.05);C-reactive protein (CRP) level of type Ⅳ,Ⅴ and Ⅵ was significantly higher than that of type Ⅱ (P ＜ 0.05).The degree of glomerulosclerosis in patients with type Ⅲ,Ⅳ,Ⅴ and Ⅵ was significantly higher than that in patients with type Ⅱ,and the degree of glomerulosclerosis in patients with type Ⅴ and Ⅵ was also significantly higher than that in patients with type Ⅲ (P ＜ 0.05).The formation of crescents in patients with type Ⅲ,Ⅳ,Ⅴ and Ⅵ was significantly higher than that in patients with type Ⅱ,and the formation of crescents in patients with type Ⅳ,Ⅴ and Ⅵ was also significantly higher than that in patients with type Ⅲ (P ＜ 0.05).The levels of serum TGF-β1,MCP-1 and IL-17 were the lowest in type Ⅱ patients and the highest in type Ⅴ and Ⅵ patients.The levels of TGF-β1,MCP-1 and IL-17 in type Ⅲ,Ⅳ,Ⅴ and Ⅵ were significantly higher than those in type Ⅱ (P ＜0.05),and the level of TGF-β1 in type Ⅳ,Ⅴ and Ⅵ was significantly higher than that in type Ⅲ (P ＜ 0.05);Serum IL-17 level of type Ⅴ and Ⅵ was significantly higher than that of type Ⅲ (P ＜ 0.05).Cox regression analysis showed that TGF-β1 and IL-17 were risk factors for pathological grading.Conclusions The higher the pathological grade of Henoch-Schonlein purpura nephritis in adults,the higher the levels of serum TGF-β1 and IL-17.TGF-β1 and IL-17 are the risk factors affecting the pathological grade of Henoch-Schonlein purpura nephritis.

Objective To evaluate the efficacy of excimer laser atherectomy (ELA) combined with drug-coated balloon (DCB) in the treatment of lower limb arteriosclerosis obliterans.Methods From Sep 2016 to Dec 2018 42 patients (42 limbs) underwent ELA combined with DCB therapy.Results There were 27 males and 15 females.The mean age was (65.7 ±9.7) years.The average lesion length was (14.3 ±7.5) cm.The technical success rate was 100％ and the remission rate of clinical symptoms was 92.9％.Ankle-brachial index was significantly improved from (0.45 ± 0.15) pre-operation to (0.85 ± 0.13) postoperation (P =0.001).The mean follow-up was 10.9 months.At 3,6,and 12 months follow-up,the primary patency rate was 95.2％,92.8％ and 84.9％.The 1-year limb salvage rate was 95.2％.Conclusion ELA combined with DCB has a good 1-year results as our data reveal.

Objective To investigate the safety and effectiveness of endovascular treatment and surgical clipping for the treatment of intracranial ruptured aneurysms. Methods From January 2012 to December 2017, patients with ruptured intracranial aneurysm treated at the Department of Neurosurgery, the Second People's Hospital of Taizhou were enrolled retrospectively. The demographics, baseline clinical data,outcomes, and complications were compared between the endovascular treatment group and the surgical clip group. Results A total of 220 patients were enrolled, they aged 55. 1 ±11. 8 years. There were 117 patients in the endovascular treatment group and 103 in the surgical clipping group. There were no significant differences in perioperative complications (26. 2％ vs. 19. 6％; χ2 = 1. 340, P = 0. 247), in-hospital mortality (6. 0％vs. 4. 9％; χ2 = 0. 135, P = 0. 713), and good outcomes at discharge (85. 5％ vs. 81. 6％; χ2 =0. 614, P = 0. 433) between the two groups. Multivariate logistic regression analysis showed that age (odds ratio [OR] 1. 072, 95％ confidence interval [CI] 1. 025-1. 124; P < 0. 001), smoking (OR 6. 325, 95％ CI 2. 367-16. 901; P < 0. 001 ), and high World Federation of Neurosurgery Societies (WFNS) grade (OR 5. 218, 95％ CI 1. 881-14. 449; P < 0. 001) had significant independent correlation with the poor clinical outcome at discharge. The imaging follow-up data in 155 aneurysms (81 in the endovascular treatment group and 74 in the surgical clipping group) were available. The follow-up time was 14. 3 ± 6. 9 months (range, 6-36 months); 20 aneurysms (12. 9％) had recurrence. There was no significant difference in the recurrence rate of the endovascular treatment group and surgical clipping group (17. 3％ vs. 8. 1％; χ2 =2. 900, P = 0. 089). The clinical follow-up data of 188 patients (95 in the endovascular treatment group and 93 in the surgical clipping group) were available. The follow-up time was 15. 5 ± 6. 8 months (range, 6- 36 months). There was no significant difference in the good outcome rate between the endovascular treatment group and surgical clipping group (95. 8％ vs. 90. 3％; χ2 = 2. 182, P = 0. 140 ). Multivariate logistic analysis showed that smoking (OR 4. 872, 95％ CI 1. 719-13. 872; P < 0. 001 ) and high WFNS grade (OR 3. 512, 95％ CI 1. 446-8. 583; P < 0. 001) were the independent risk factor for long-term poor outcome. Conclusion The efficacy and safety of surgical clipping for ruptured intracranial aneurysms were comparable to endovascular treatment. Age, smoking, and WFNS grade were the important factors affecting the outcomes of patients.