Background: Spent culture medium (SCM) as a source of DNA for preimplantation genetic tests aneuploidy (PGT-A) has been widely discussed. Methods: Seventy-five blastocysts that were donated for research provided a unique possibility in which multiple specimens, including trophectoderm (TE) biopsy, SCM, and paired corresponding whole blastocyst (WB) specimens from the same blastocyst source, could be utilized for the purpose of this preclinical validation. Results: To conduct a validation ploidy concordance assessment, we evaluated the full chromosomal concordance rates between SCM and WB (SCM-to-WB), and between TE and WB (TE-to-WB) as well as sensitivity, specificity and overall diagnostic accuracy. 78.67% (59/75) of NGS results in the SCM group were interpretable, a significantly lower percentage than their corresponding TE and WB groups. This discrepancy manifests itself in intrinsically low quantity and poor integrity DNA from SCM. Subsequently, remarkable differences in full concordance rates (including mosaicism, and segmental aneuploidies) are seen as follows: 32.2% (SCM-to-WB, 19/59) and 69.33% (TE-to-WB, 52/75), (p < 0.001). In such cases, full concordance rates were 27.27% (15/55) in SCM-to-WB, and, 76% (57/75) in TE-to-WB (p < 0.001). Collectively, the NGS data from SCM also translated into lower sensitivities, Positive Predictive Value (PPV), Negative Predictive Value (NPV), overall diagnostic accuracies, and higher Negative Likelihood Ratio (NLR). Conclusions: Our study reveals that DNA is detectable in the majority of SCM samples. Individual chromosomal aberration, such as segmental aneuploidy and mosaicism, can be quantitatively and qualitatively measured. However, TE still provides a more accurate and reliable high-throughput methodology for PGT-A. Meanwhile, cell-free DNA in SCM reporting lacks uniform diagnostic interpretations. Considering that this test is meant to determine which embryos are relegated to be discarded, PGT-A with cell-free DNA in SCM should not be permitted to be applied in routine clinical settings for diagnosis purpose.

Background: Toll-like receptor 2 (TLR2) is essential for the immune response to tuberculosis (TB). The goal of the present study was to investigate whether the guanine-thymine (GT) repeat microsatellite polymorphism in intron 2 of the TLR2 gene might be correlated with susceptibility to TB in Han Chinese. Methods: The number of (GT)n repeats was determined by gene scanning from 244 patients with TB and 233 control subjects. The expression of TLR2 on CD14+ peripheral blood mononuclear cells was determined using flow cytometry. Results: No association in allelic polymorphism between control subjects and patients with TB was found. However, the S/M genotype of the microsatellite polymorphism was more frequent in TB patients than in healthy controls (p=0.01). The S/L genotype was more popular in controls than in patients with TB (p=0.007). TLR2 expression was higher in subjects with the S/L genotype than in those with the S/M genotype (p<0.05). Conclusions: Our data suggest that the S/M genotype of the microsatellite (GT)n polymorphisms in intron 2 of the TLR2 gene may increase susceptibility to TB in Chinese, and the S/L genotype may act as a negative risk factor. Clin Chem Lab Med 2010;48:785–9.

Atopic dermatitis (AD) is a prevalent inflammatory skin disease. Ample evidence has shown that non-coding RNAs play important roles in the progression of AD, however, the function of plasma microRNAs in AD is poorly understood. To identify key plasma microRNAs and explore their potential roles in AD. Plasma microRNAs from five children with AD and five control children were sequenced by microRNA sequencing (miRNA-seq) and five differentially expressed microRNAs were verified by RT-qPCR in 30 AD and 15 control children. The most differentially expressed microRNA, hsa-miR-194-5p, was selected for further analysis. Human epidermal keratinocytes were subjected to RNA sequencing following over-expression of hsa-miR-194-5p, and down-regulated genes were detected by RT-qPCR. The diagnostic potential of hsa-miR-194-5p in AD was evaluated based on receiver operating characteristic (ROC) curve analysis. Further hsa-miR-194-5p-regulated expression and gene-hsa-miR-194-5p interactions were evaluated by western blotting and luciferase assays, respectively. We identified 40 differentially expressed microRNAs, 26 up-regulated and 14 down-regulated, in children with AD compared with controls. Among the five verified plasma microRNAs, the most significant change was down-regulated hsa-miR-194-5p, which was shown to potentially serve as a biomarker for AD based on ROC analysis. Twenty-two down-regulated genes were observed following hsa-miR-194-5p over-expression, which were significantly associated with keratinocyte differentiation and establishment of the skin barrier. Moreover, HS3ST2 protein expression was down-regulated following over-expression of hsa-miR-194-5p, and the 3'-UTR of HS3ST2 was shown to bind to hsa-miR-194-5p. Hsa-miR-194-5p might be involved in the pathogenesis of AD by regulating HS3ST2 expression.

Background. This study is aimed at investigating the repairing effect of mesenchymal stem cells and their exosomes from different sources on ovarian granulosa cells damaged by chemotherapy drugsphosphoramide mustard (PM). Methods. In this study, we choose bone marrow mesenchymal stem cells (BMSCs) and human placental mesenchymal stem cells (HPMSCs) for research. Then, they were cocultured with human ovarian granulosa cells (SVOG) injured by phosphoramide mustard (PM), respectively. β-Galactosidase staining, flow cytometry, and Western blot were used to detect the changes in the senescence and apoptosis of SVOG cells before and after their coculture with the above two types of MSCs. Subsequently, exosomes from these two types of MSCs were extracted and added to the culture medium of SVOG cells after PM injury to test whether these two types of exosomes played a role similar to that of MSCs in repairing damaged SVOG cells. Results. PM treatment-induced apoptotic SVOG cells were significantly decreased after HPMSCs and BMSCs as compared with control group. After coculturing with these two types of MSCs, PM-treated SVOG cells showed significantly reduced senescence and apoptosis proportions as well as cleaved-Caspase 3 expression, and HPMSCs played a slightly stronger role than BMSCs in repairing SVOG cells in terms of the above three indicators. In addition, the ratios of senescent and apoptotic SVOG cells were also significantly reduced by the two types of exosomes, which played a role similar to that of MSCs in repairing cell damages. Conclusions. The results indicated that BMSCs, HPMSCs, and their exosomes all exerted a certain repair effect on SVOG cells damaged by PM, and consistent repair effect was observed between exosomes and MSCs. The repair effect of exosomes secreted from BMSCs and HPMSCs on the SVOG cells was studied for the first time, and the results fully demonstrated that exosomes are the key carriers for MSCs to play their role.