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Characterization of an Ex Vivo Equine Endometrial Tissue Culture Model Using Next-Generation RNA-Sequencing Technology.

ABSTRACT: Persistent mating-induced endometritis is a major cause of poor fertility rates in the mare. Endometritis can be investigated using an ex vivo equine endometrial explant system which measures uterine inflammation using prostaglandin F_2α as a biomarker. However, this model has yet to undergo a wide-ranging assessment through transcriptomics. In this study, we assessed the transcriptomes of cultured endometrial explants and the optimal temporal window for their use. Endometrium harvested immediately post-mortem from native pony mares (n = 8) were sampled (0 h) and tissue explants were cultured for 24, 48 and 72 h. Tissues were stored in RNALater, total RNA was extracted and sequenced. Differentially expressed genes (DEGs) were defined using DESeq2 (R/Bioconductor). Principal component analysis indicated that the greatest changes in expression occurred in the first 24 h of culture when compared to autologous biopsies at 0 h. Fewer DEGs were seen between 24 and 48 h of culture suggesting the system was more stable than during the first 24 h. No genes were differentially expressed between 48 and 72 h but the low number of background gene expression suggested that explant viability was compromised after 48 h. ESR1, MMP9, PTGS2, PMAIP1, TNF, GADD45B and SELE genes were used as biomarkers of endometrial function, cell death and inflammation across tissue culture timepoints. STRING assessments of gene ontology suggested that DEGs between 24 and 48 h were linked to inflammation, immune system, cellular processes, environmental information processing and signal transduction, with an upregulation of most biomarker genes at 24 h. Taken together our observations indicated that 24-48 h is the optimal temporal window when the explant model can be used, as explants restore microcirculation, perform wound healing and tackle inflammation during this period. This key observation will facilitate the appropriate use of this as a model for further research into the equine endometrium and potentially the progression of mating-induced endometritis to persistent inflammation between 24 and 48 h.

SUBMITTER: Monteiro de Barros MR

PROVIDER: S-EPMC8300099 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundWith a high incidence rate and mortality rate, tuberculosis (TB) is a major public health concern worldwide. There is an urgent need to develop the rapid, reliable and affordable diagnostic test for the detection of Mycobacterium tuberculosis (Mtb). Metagenomic next-generation sequencing (mNGS) showed promising values in the rapid diagnosis of clinical TB, while studies on the application of mNGS in pulmonary and extrapulmonary TB remain scarce.MethodsIn this study, we collected the results of both mNGS and culture of 70 specimens from suspected TB patients at the Shanghai Pulmonary Hospital of Tongji University. Results of GeneXpert MTB/RIF assay (Xpert) were obtained from 19 patients. We also assessed the diagnostic performance, relationship and consistency between mNGS, traditional culture method, and Xpert.ResultsOverall, 36 of 70 suspected patients were finally diagnosed with TB. The sensitivity, specificity, positive predictive value, negative predictive value and the Youden index of mNGS in all clinical TB specimens were calculated as 66.7% (95% CI: 0.489-0.809), 97.1% (95% CI: 0.829-0.998), 96.0% (95% CI: 0.777-0.998), 73.3% (95% CI: 0.578-0.849), 63.8%, respectively. The sensitivity of mNGS was much superior to that of conventional culture method (66.7%, 95% CI: 0.489-0.809 vs. 36.1%, 95% CI: 0.213-0.538) and Xpert (76.9%, 95% CI: 0.460-0.938 vs. 61.5%, 95% CI: 0.323-0.849). In pulmonary TB cases, mNGS, culture, and Xpert all demonstrated better diagnostic capacity for TB when compared with extrapulmonary TB cases. Among all methods and sample classifications, the Youden index of parallel diagnostic test in pulmonary samples was outstanding (mNGS/culture 88.2%; mNGS/Xpert 100%). Furthermore, the correlation and concordance between mNGS and culture method in all types of specimens was statistically significant (both P<0.001).ConclusionsWith high sensitivity and specificity, mNGS showed remarkable diagnostic performance in various samples from suspected TB patients. Combining mNGS and culture or Xpert method had the potential for clinical application in TB.

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Characterization of an Ex Vivo Equine Endometrial Tissue Culture Model Using Next-Generation RNA-Sequencing Technology.

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