Objective To investigate the role of the O-6-methylguanine-DNA methyltransferase (MGMT) gene-3′ untranslated region (UTR) microRNA-associated single nucleotide polymorphism (miRSNP) (rs7896488 G>A) in affecting miR-4297-targeted modulation of MGMT in senescence of lung cells with polymorphic genotypes induced by fractionated low dose ionizing radiation (LDIR). Methods i) MiRSNPs were predicted and screened using bioinformatics, and DNA from two types of lung cells, A549 cells and human bronchial epithelioid cells (HBE cells), was extracted for target gene sequencing. After co-transfection of pGL3c-MGMT-3′UTR-rs7896488 G>A reporter gene recombinant plasmid, pRL-TK Vector with micrON mimic NC #22 or micrON hsa-miR-4297 mimic (set up as the mimic NC group and the miR-4297 mimic group) in these two types of lung cells, dual luciferase reporter gene assay was performed. The relative expression of MGMT mRNA was detected by real-time fluorescence quantitative polymerase chain reaction, and the relative expression of MGMT protein was detected by Western blotting. ii) These two types of lung cells were randomly divided into the control group and irradiation group, which received either 0 or 100 mGy X-rays irradiation seven times. After irradiation, the cells were transfected with either micrON mimic NC #22 or micrON hsa-miR-4297 mimic, resulting in mimic NC + control group, miR-4297 mimic + control group, mimic NC + irradiation group, and miR-4297 mimic + irradiation group. Cells were collected for senescence-associated-β-galactosidase (SA-β-Gal) staining, and the relative expression of matrix metalloproteinase-9 (MMP-9) and chemokine (C-X-C motif) ligand-1 (CXCL-1) proteins was detected via Western blotting. Results i) The rs7896488 G>A was the miRSNP located in the conserved binding region targeted by miR-4297 in the MGMT gene 3′UTR. A549 cells were the rs7896488 GG wild-type homozygous genotype, while HBE cells were the rs7896488 GA heterozygous mutant genotype. In the miR-4297 mimic group, A549 and HBE cells carrying the rs7896488 G allele showed significantly lower dual-luciferase activity compared with that in the mimic NC group (both P<0.01). However, there was no significant difference in dual-luciferase activity between the two groups in both A549 and HBE cells carrying the rs7896488 A allele (both P>0.05). The relative expression levels of MGMT mRNA and MGMT protein of A549 cells in the miR-4297 mimic group were lower than those in the mimic NC group (both P<0.05). However, there was no significant difference in MGMT mRNA and MGMT protein of HBE cells between these two groups (both P>0.05). ii) The relative activity of SA-β-Gal and the relative expression of MMP-9 and CXCL-1 proteins of A549 cells in the miR-4297 mimic+irradiation group were higher than those in the mimic NC + control group, the miR-4297 mimic + control group, and the mimic NC + irradiation group (all P<0.05). The relative activity of SA-β-Gal and the relative expression of MMP-9 and CXCL-1 proteins of HBE cells in the miR-4297 mimic + irradiation group were higher than those in the mimic NC + control group and the miR-4297 mimic + control group (all P<0.05), while there was no significant difference compared with those in the mimic NC + irradiation group (all P>0.05). Conclusion MGMT-3′UTR-miRSNP rs7896488 G>A plays a role in LDIR-induced senescence of lung cells with different polymorphic genotypes by affecting miR-4297-targeted regulation of MGMT.

Objective To investigate the effects of fractionated low-dose ionizing radiation (LDIR) on the ferroptosis in human bronchial epithelial (HBE) cells as well as the associated differentially expressed genes (DEGs), biological processes, and signaling pathways. Methods HBE cells were exposed to different single doses of X-ray irradiation (0, 25, 50, 75, and 100 mGy) for 24, 48, and 72 h, respectively. The change in cell viability was detected by MTT assay. Cells were irradiated with 0, 25, 50, and 100 mGy X-rays 5 times, with 48 h between each irradiation and a dose rate of 50 mGy/min. Cells were harvested 24 h after irradiation for the measurement of the expression of ferroptosis-related genes SLC7A11 and GPX4 at the mRNA and protein levels, cellular iron content, and the expression of FTH1 and FTL mRNAs. High-throughput sequencing was used to screen for the DEGs in each dose group, followed by Gene Ontology-Biological Process (GO-BP) analysis, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis, and Gene Set Enrichment Analysis (GSEA). Results Compared with the control group, single-dose LDIR significantly increased cell proliferation at 75 mGy after 24 h (P < 0.05), at 50, 75, and 100 mGy after 48 h (P < 0.05), and at 75 and 100 mGy after 72 h (P < 0.05). Compared with the control group, at the end of the fifth fractionated LDIR, SLC7A11 and GPX4 mRNAs decreased at all doses (P < 0.05), SLC7A11 protein decreased at all doses, GPX4 protein decreased at 25 and 100 mGy, iron content increased at all doses, and FTH1 and FTL mRNAs decreased at all doses (P< 0.05). Sequencing analysis identified 248, 30, and 291 DEGs and 10, 2, and 9 ferroptosis-associated genes at the three doses compared to the control. Gene Ontology-Biological Process analysis showed that DEGs were mainly enriched in biological processes such as response to lipids, cell death, and response to unfolded proteins. Kyoto Encyclopedia of Genes and Genomes analysis showed that DEGs were mainly enriched in the JAK-STAT signaling pathway, lipids and atherosclerosis, ferroptosis, protein processing in the endoplasmic reticulum, and FoxO signaling pathway. Gene set enrichment analysis showed that DEGs were mainly enriched in ferroptosis, fatty acid degradation, and glutathione metabolism. Conclusion Fractionated low-dose radiation induced ferroptosis in HBE cells, and DEGs were predominantly enriched in biological processes and signaling pathways related to inflammation, ferroptosis, and endoplasmic reticulum stress.

Background In recent years, the increasingly widespread application of nuclear and medical radiation technologies has resulted in a large number of occupational populations exposed to low-dose ionizing radiation (LDIR). At present, there is no consistent conclusion on the effects of long-term exposure to LDIR on the metabolic health of the occupational population. Objective To explore the association between long-term exposure to LDIR and metabolic syndrome (MetS) among medical radiologists. Methods A cross-sectional study was conducted to enroll 6431 medical radiologists who ordered occupational health checkups from January 2022 to December 2023, and basic information such as demographic characteristics, occupational characteristics, lifestyles, and dietary habits was collected through questionnaires. Physical examinations were conducted using a standardized protocol. Individual dose equivalents of occupational external exposure were monitored by dosimeters worn by medical radiologists. Logistic regression model was used for identifying influencing factors of MetS and sensitivity analysis, and restricted cubic spline model was used to explore the dose-response relationship between cumulative effective dose and MetS. Two-stage linear regression was used to evaluate threshold effect of association between cumulative effective dose and MetS. Results The positive rate of MetS was 13.6% (877/6431) among the enrolled 6431 study participants. The logistic regression showed that gender, age, monthly income, body mass index (BMI), cumulative effective dose, and smoking were influencing factors for MetS. The risk of MetS was higher in males (OR=1.511, 95%CI: 1.209, 1.888); using the < 30 years old group as reference, the risk of MetS was higher in the 30-39 years old (OR=1.509, 95%CI: 1.095, 2.079), 40-49 years old (OR=2.214, 95%CI: 1.551, 3.161), and ≥50 years old (OR=3.480, 95%CI: 2.338, 5.181) groups; using the <10000 yuan group as reference, the risk of MetS was lower in the group with a monthly income of 10000-14999 yuan (OR=0.715, 95%CI: 0.582, 0.878); the risk of MetS was higher in the BMI ≥ 24 kg·m⁻² group (OR=7.548, 95%CI: 6.223, 9.156); using the < 0.76 mSv group as reference, the risk of MetS was higher in the cumulative effective dose of 0.76-2.73 mSv group (OR=1.270, 95%CI: 1.017, 1.586); the risk of MetS was higher in the smoking group (OR=1.734, 95%CI: 1.428, 2.107). The results of restricted cubic spline showed that the cumulative effective dose was nonlinearly correlated with MetS (P _{non-linearity}=0.028), with an inflection point of 1.25 mSv. The risk of developing MetS increased by 34.1% for each unit increase in cumulative effective dose up to 1.25 mSv, whereas above 1.25 mSv, the statistical association was not significant. The sensitivity analysis results showed that after excluding the self-reported hypertensive and diabetic patients, the cumulative effective dose 0.76-2.73 mSv group still had an increased risk of developing MetS compared with the < 0.76 mSv group (P < 0.05), and the results were robust. Conclusion Among medical radiologists, male, age, BMI, and smoking are positively correlated with MetS, and monthly income is negatively correlated with MetS; cumulative effective dose is non-linearly correlated with MetS among medical radiologists.

In recent years, the bacteriophage Φ29 (Phi29) DNA polymerase has garnered increasing attention due to its high-fidelity amplification capacity at constant temperatures. To advance the industrial application of this type of isothermal polymerases, this study mined and characterized new enzymes from the microbial metagenome based on the known Phi29 DNA polymerase sequence. The results revealed that a new enzyme, Php29 DNA polymerase, was identified in the microbial metagenome with plants as the hosts. This enzyme exhibited higher strand displacement activity, with a 59.5% similarity to bacteriophage Φ29. Experimental validation demonstrated that the enzyme had 3'→5' exonuclease activity, and its amplification products can serve as substrates for further catalytic reactions. The discovery and validation of Php29 DNA polymerase gives insights into the future industrial application of isothermal polymerases.
DNA-Directed DNA Polymerase/metabolism* ; Bacillus Phages/genetics* ; Metagenome

Bacterial therapy has attracted increasing attention due to its special mechanism and abundant applications. With the flourishing development of synthetic biology, therapeutic genes have been introduced into engineering bacteria to improve their antitumor efficacy. However, it is difficult to spatiotemporally control the expression of these therapeutic genes at the tumor site in vivo, thereby considerably limiting the application of engineered bacteria in tumor treatment. To resolve this problem, we constructed a temperature-responsive bacterial strain capable of triggering the expression of exogenous genes in a laser-controllable way. Noxa, a pro-apoptotic protein, is chosen to test the expression of exogenous protein and its anti-tumor effect in engineered bacteria upon laser irradiation. Firstly, Noxa was fused to the C-terminus of the bacterial outer membrane protein cytolysin A (ClyA), and then the recombinant gene fragment ClyA-Noxa was inserted into the temperature-sensitive plasmid pBV220 and the recombinant plasmid was transformed into non-pathogenic Escherichia coli MG1655. Thus, we constructed the engineering strain (TRB@Noxa) that could express Noxa on the bacterial surface. TRB@Noxa could target and colonize the tumor tissue without causing notable host toxicity. The bacterial infection triggered thrombosis in the tumor tissue, resulting in the darkness of tumor sites. In a xenograft mouse tumor model, our strategy demonstrated precise tumor targeting and strong tumor inhibition. In conclusion, we successfully constructed a new engineering bacterial strain TRB@Noxa. TRB@Noxa combined with photothermal therapy could arrest tumor growth in the absence of photosensitizers, which represents an appealing method for antitumor therapy in the future.
Escherichia coli/radiation effects* ; Animals ; Humans ; Lasers ; Mice ; Proto-Oncogene Proteins c-bcl-2/biosynthesis* ; Neoplasms/therapy* ; Genetic Engineering ; Cell Line, Tumor ; Escherichia coli Proteins/genetics*

Objective:To explore the interaction effect of diabetes and obesity on recurrence or anal fistula in patients with perianal abscess after simple incision and drainage.Methods:The clinical data of 163 patients with perianal abscess who underwent simple incision and drainage from Jun. 2021 to Jun. 2023 were analyzed retrospectively. The incidence of recurrence or anal fistula in 6 months after surgery was calculated. Multivariate Logistic regression model was used to analyze the influencing factors of postoperative recurrence or anal fistula. The multiplicative and additive models were used to analyze the interaction effect of diabetes and obesity on the risk of postoperative recurrence or anal fistula.Results:In 6 months after simple incision and drainage, the incidence of recurrence or anal fistula was 28.22% (46/163). Univariate analysis results showed that gender, obesity, and diabetes were related to recurrence of perianal abscess or incidence of anal fistula ( P<0.05). Multivariate Logistic regression analysis results showed that obesity ( OR=2.447, 95% CI: 1.320-4.538) and diabetes ( OR=2.162, 95% CI: 1.187-3.938) were independent risk factors for postoperative recurrence or anal fistula ( P<0.05). Interaction effect analysis found that after adjusting for confounding factors, diabetes and obesity had additive interaction effect on the risk of postoperative recurrence or anal fistula. The relative excess risk due to interaction (RERI), attribution percentage (AP), and interaction effect index (S) were 1.829 (95% CI: 0.605-3.007), 0.405 (95% CI: 0.143-0.597), and 2.098 (95% CI: 1.201-3.172), respectively. There was no multiplicative interaction effect between the two ( P>0.05) . Conclusions:Diabetes and obesity are independent risk factors for recurrence or anal fistula in patients with perianal abscess after simple incision and drainage. The two may have synergistic effect on the risk of postoperative recurrence or anal fistula.

Objective To investigate the mechanism of fractionated low-dose ionizing radiation (LDIR) in the induction of EA.hy926 cell senescence. Methods EA.hy926 cells were irradiated with X-ray at 0, 50, 100, and 200 mGy × 4, respectively, and cultured for 24, 48, and 72 h. Several indicators were measured, including the levels of cellular senescence-associated β-galactosidase (SA-β-gal) staining, mRNA levels of senescence-associated cell cycle protein-dependent kinase inhibitor genes CDKN1A and CDKN2A, reactive oxygen species (ROS), total antioxidant capacity (T-AOC), and phosphorylated H2A histone family member X (γ-H2AX). Results After 4 fractionated LDIR, compared with the control group, the treatment groups showed increased nucleus area, blurred cell edge, and increased SA-β-gal positive area (P < 0.05) at 24, 48 and 72 h. After 4 fractionated LDIR, the mRNA level of CDKN1A increased in the 100 and 200 mGy × 4 groups at 24 and 48 h (P < 0.05), and CDKN2A mRNA level increased in the 100 and 200 mGy × 4 groups at 48 and 72 h (P < 0.05). The fluorescence intensity of ROS increased in treatment groups at 24, 48, and 72 h after 4 fractionated LDIR (P < 0.05). After 4 fractionated LDIR, the T-AOC level increased in the 100 and 200 mGy × 4 groups at 24 h (P < 0.05), and T-AOC level increased in all treatment groups at 48 and 72 h (P < 0.05). After 4 fractionated LDIR, γ-H2AX fluorescence intensity increased in all treatment groups at 24 h (P < 0.05), and the fluorescence intensity increased in the 100 and 200 mGy × 4 groups at 48 and 72 h (P < 0.05). Conclusion Fractionated LDIR can induce cellular senescence in EA.hy926 cells by impacting the cellular oxidation-antioxidation and oxidative damage levels, and the effects were relatively evident at 100 and 200 mGy.

Objective To explore the differentially expressed mRNAs and related biological processes and pathways in fractional low-dose ionizing radiation (LDIR)-induced senescence of normal human bronchial epithelial (HBE) cells by high-throughput mRNA sequencing and bioinformatics techniques. Methods Senescence-associated β-galactosidase staining and senescence-associated secretion phenotype gene mRNA and protein expression levels were measured at 24 and 48 h after irradiating HBE cells 7 times at doses of 0, 50, 100, and 200 mGy, respectively. The differentially expressed genes were screened by high-throughput sequencing for Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses. Results The senescence-positive area of fractional low-dose irradiated HBE cells increased in a dose-dependent manner (P < 0.05). The mRNA levels and protein expression of transforming growth factor-β1(TGF-β1) and matrix metalloproteinase-9(MMP-9) genes were increased in the 100 mGy × 7 and 200 mGy × 7 groups at 24 and 48 h after the end of irradiation compared with the control group. High-throughput sequencing showed that there were 882, 475, and 1205 differentially expressed mRNAs in each dose group compared with the control group. GO analysis showed that the differentially expressed mRNAs in each dose group were mainly enriched in biological processes such as cell cycle regulation, regulation of nitrogen compound metabolic process, regulation of cell division and response to stimulus. KEGG analysis showed that the differentially expressed mRNAs were mainly enriched in the pathways of cell cycle, cell senescence, and ferroptosis. Conclusion Fractional LDIR induced senescence in HBE cells, and differentially expressed mRNA-associated biological processes and pathways in senescent cells are related to cell cycle and cell senescence.

Objective:To explore the immunological characteristics of peripheral blood and genetic variations of 11 immunodeficiency virus(HIV)-negative children with Talaromyces marneffei(TM) infection, thus enhancing the diagnostic and therapeutic levels of TM infection in children. Methods:Clinical data of 11 HIV-negative children with TM infection who presented to Guangzhou Women and Children′s Medical Center, Guangzhou Medical University from January 2010 to December 2022 were retrospectively analyzed, including clinical characteristics, peripheral immune profile and genetic test results.Results:A total of 11 HIV-negative children with TM infections were recruited, involving 9 males and 2 females with a median age of 19 months.The main clinical manifestations were fever (10/11, 90.91%), cough (10/11, 90.91%) and hepatomegaly (7/11, 63.64%). Common severe complications included acute respiratory distress syndrome (7/11, 63.64%) and septic shock (5/11, 45.45%). Finally, 2 children died.Transient neutropenia occurred in 6 cases (6/11, 54.55%), and lymphocytopenia combined with serum immunoglobulin (Ig) G decrease was observed in 4 cases (4/11, 36.36%). IgA decrease, IgM decrease, IgE decrease, IgM increase and IgE increase were observed in 6 cases, 3 cases, 5 cases, 3 cases, and 2 cases, respectively.Both T-lymphocyte and B-lymphocyte counts decreases was observed in 1 case.Genetic testing was performed in all recruited children, and genetic variations were detected in all of them.Inborn errors of immunity (IEIs) were diagnosed in 8 cases, including 4 diagnosed as CD 40 ligand deficiency with CD40LG variation, 1 of severe combined immunodeficiency with IL2RG variation, 1 of Signal transduction and activator of transcription 3(STAT3)-hyper-IgE syndrome with STAT3 variation and 1 of familial candidiasis type 2 with CARD9 compound heterozygous mutations.In the other 3 cases, 2 carried genetic variations that were likely pathogenic, and 1 case was considered uncertain. Conclusions:The clinical manifestations of HIV-negative children with TM infection are atypical, which is characterized as serious complications and high mortality.Early identification and gene testing to detect potential IEIs can improve the prognosis of TM infection.

OBJECTIVE To optimize the steaming and processing technology of wine-steamed Taxillus chinensis， and to characterize its quality. METHODS Using the content of avicularin， quercitrin， quercetin and appearance traits as evaluation indicators， the analytic hierarchy process （AHP）-entropy weight method was used to determine the weights of each indicator， and the comprehensive scores of those indicators were used as response values. Box-Behnken response surface method was used to investigate the effects of solid-liquid ratio （g/mL）， soaking time， and steaming time on the processing technology of wine-steamed T. chinensis， optimize the best processing technology， and verify it. Fifteen batches of T. chinensis decoction pieces from different origins were used to prepare wine-steamed T. chinensis using the best processing technology， and their qualities were characterized. RESULTS The optimal processing technology for wine-steamed T. chinensis was to take 100 g of T. chinensis decoction pieces， add 20 mL of yellow wine， seal and moisten for 2 h， steam at normal pressure for 1 h， take out and dry at 50 ℃. The surface of wine-steamed T. chinensis prepared by the optimal processing technology was reddish brown or brownish， and its powder was dark brown， with a hard or brittle texture that was easy to break， and had a slight aroma of alcohol， and an astringent taste. Results of microscopic and thin-layer identification for the stem cross-section of wine-steamed T. chinensis were the same as those of raw T. chinensis. The contents of moisture， total ash and acid-insoluble ash were 3.92%-8.75%， 2.27%-5.08%， and 0.19%-0.82%， respectively； the contents of water-soluble extract were 11.28%-18.56%， and the contents of alcohol-soluble extract were 3.36%-8.58%； the contents of avicularin， quercitrin， and quercetin were 0.22-1.64， 0.26-2.45， and 0.01-0.38 mg/g， respectively. CONCLUSIONS This study successfully optimized the processing technology of wine-steamed T. chinensis and preliminarily characterized its quality， which can provide reference for the standardized processing and establishment of quality standards for wine-steamed T. chinensis decoction mail：wyl@sxtcm.edu.cn pieces.