Laryngopharyngeal reflux (LPR) induces a differential damage effect on several anatomic sites within the larynx and hypopharynx; therefore, an in vitro model is needed for each anatomic site. This study aimed to establish a primary culture method for human laryngeal and hypopharyngeal epithelial cells derived from multiple anatomic sites. Surgical mucosa specimens were treated with a two-step enzymatic strategy to establish a primary culture. Of the 46 samples, primary cultivation was achieved successfully with 36 samples, and the positive ratio was 78.3%. In addition, flow cytometry revealed that these primary cells were epithelial cells with a purity of 94.9%. The proliferative ability was confirmed by positive staining for Ki-67. Laryngeal and hypopharyngeal epithelial cells from multiple sites exhibited similar epithelial morphology and positive cytokeratin expression. These cells can be cultured to passage 4. In summary, we successfully established the in vitro epithelial model of larynx and hypopharynx subsites, which may potentially be used as a platform for reflux research, especially for site-specific damage effect.
Bacterial leaf blight (BLB) has caused severe yield losses in cantaloupe (Cucumis melo L.) in the major melon-growing regions of China since the beginning of the twentieth century. Historically, Pseudomonas syringae pv. lachrymans was considered to be the causal agent of BLB of cantaloupe and angular leaf spot of cucumber. In the process of characterizing bacteria isolated from cantaloupe, we observed that putative P. syringae pv. lachrymans yielded negative results in P. syringae pv. lachrymans-specific PCR assays. This suggested that the P. syringae pv. lachrymans-like strains from cantaloupe were distinct from those recovered from cucumber. To investigate the differences between P. syringae pv. lachrymans-like strains isolated from cantaloupe and cucumber, 13 P. syringae strains isolated from cantaloupe [12 from China and 1 from Zimbabwe (NCPPB2916)] and 7 additional P. syringae reference strains were analyzed by catabolic profiling, phylogenetic analysis by multilocus sequence analysis (MLSA) and pathogenicity tests on cantaloupe leaflets. Catabolic profiling and MLSA based on 10 housekeeping genes and 2 hypersensitive response and pathogenicity (hrp) genes allowed us to differentiate strains isolated from cantaloupe and cucumber. Pseudomonas syringae pv. lachrymans strains isolated from cucumber clustered with genomospecies 2, and 13 P. syringae strains isolated from cantaloupe belonged to genomospecies 1. While all cantaloupe strains were closely related to P. syringae pv. aptata, they could be differentiated from this pathovar based on metabolic tests and MLSA. Pathogenicity tests showed that all strains isolated from cantaloupe and cucumber were only pathogenic on their original hosts. Based on these observations we conclude that P. syringae pv. lachrymans strains recovered from cantaloupe in China represent a novel phylotype.
Objective The role of lifestyle habits in patients with laryngopharyngeal reflux disease (LPRD) is comparatively underexplored. We aim to examine the specific lifestyle habits in patients with LPRD. METHODS Systematic sampling was applied to select respondents aged 18 through 80 years in otorhinolaryngology-head and neck surgery (OHNS) clinics in Nan Fang Hospital during August 2017–July 2018, 1658 eligible participants were included by a systematic sampling method. Subjects with RSI score>13 were considered as LPRD patients. The risk of reflux symptoms was estimated and multivariate calculated as odds ratios in relation to exposure to tobacco smoking, alcohol, coffee, tea, carbonated drinks, chocolate, spicy food, night sleep time, dinner-to-bed time, subjective sleep quality, and physical exercise. Results There was a significant dose-response association between carbonated beverage and LPRD. Among people who had drinking carbonated drinks the odds ratio was 1.76 (OR 1.77, 95% CI 1.24–2.50, P = .002) compared with non-carbonated drinker. A similar positive association was found for poor subjective sleep quality and shorter night sleeping time, the odds ratio for reflux was 1.58 (95% CI 1.14 to 2.18) among those who always have poor subjective sleep quality compared with those whose have good subjective sleep quality. The odds ratio for reflux was 2.29 (95% CI 1.23–4.28, P = .015) among those who always sleep 3–5 hours every night compared with those who sleep more than 8 hours every night. Beyond that, we found high BMI may have a negative correlation with LPRD, the odds ratio for reflux was .61 (95% CI 0.39 to .95, P = .054) among those whose BMI >25 kg/m2 compared with those BMI ≤ 20 kg/m2. Conclusions Patients with LPRD may have certain lifestyle habits, avoid carbonated beverage, poor subjective sleep quality, and lack of sleep should be advised in treatment of LPRD.
Primary laryngeal epithelial cells are essential to exploring the mechanisms of laryngeal and voice disorders; however, they are difficult to study and apply because of their limited life span. The purpose of this study was to develop a stable and reliable in vitro model for the comprehensive study of the pathogenesis of laryngeal and voice diseases. The pLVTHM-Bmi1 plasmid was constructed and used to immortalize primary laryngeal epithelial cells by lentiviral infection. The expressions of Bmi1, human telomerase reverse transcriptase (hTERT), p53, and pRB pathway proteins were detected by western blotting. Functional characteristics of the immortalized cell lines were verified by cell senescence β-galactosidase staining, 5-ethynyl-2′-deoxyuridine cell proliferation test, and flow cytometry. We successfully introduced Bmi into human subglottic (hSG) cells and human ventricle (hV) cells. Both the human immortalized subglottic Bmi1 (hSG-Bmi1) cell line and the human immortalized ventricle Bmi1 (hV-Bmi1) cell line maintained normal epithelial morphology and divided successfully after more than 20 culture passages. As Bmi1 was overexpressed in these cells, the expression of human telomerase reverse transcriptase (hTERT) and phosphorylated Rb increased while p16 and p21 decreased. Following Bmi1-mediated immortalization, cell senescence decreased significantly, and cell proliferation was accelerated. Tumor formation was not observed for hSG, hV, or hSG-Bmi1, and hV-Bmi1 cells in nude mice. hSG-Bmi1 cells dominated by stratified squamous epithelium and hV-Bmi1 cells dominated by columnar cells were established. The new cell lines lay a foundation for the study of the pathogenic mechanisms of laryngeal and voice diseases.
Mancozeb is a broad-spectrum fungicide frequently applied as foliar spray in tobacco fields to control fungal diseases. The response of tobacco phyllosphere microbiota toward mancozeb stress was assessed using high-throughput sequencing at four time points: before spraying, and 5, 10 and 15 days after fungicide application. Results showed that the foliar application of mancozeb had moderate but significant effect on fungal community composition of tobacco phyllosphere. In all samples, Ascomycota and Proteobacteria were the most abundant phyla, and Alternaria was the dominant fungal genus. Moreover, mancozeb significantly affected indigenous bacterial communities of tobacco leaves; Pseudomonas was predominant in untreated and before mancozeb treatment groups. An increase in abundance of Ascomycota was observed in diseased samples while healthy samples showed a reduction after mancozeb application. Increased abundance of Proteobacteria was observed in treated samples, which was much higher in diseased than in healthy groups. Increased abundance of Alternaria was observed until 15 days after mancozeb application, while successive reduction in abundance of the genus was observed in the untreated healthy group. Fungal alpha diversity indices in untreated groups increased from the first collection stage to the third, whereas a decrease in four fungal indices was observed at 5 d after mancozeb exposure. A significant difference between treated and untreated groups was observed in terms of fungal richness indices of similar samples from the previous stage. Bacterial diversity indices increased after mancozeb application while they decreased in untreated samples. Mancozeb was effective in altering the fungal community structure rather than bacterial community structure. After mancozeb treatment, the proportion of pathotroph-saprotroph-symbiotroph increased while it decreased in the untreated groups. The overall findings revealed ecological implications of the effects of mancozeb on tobacco phyllosphere microbiome; our results would provide a theoretical basis for future studies on microecological protection of phyllosphere.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.