MALDI-TOF-MS for Rapid Screening and Typing of ß-Globin Variant and ß-Thalassemia through Direct Measurements of Intact Globin Chains.

BACKGROUND: Traditional phenotype-based screening for ß-globin variant and ß-thalassemia using hematological parameters is time-consuming with low-resolution detection. Development of a MALDI-TOF-MS assay using alternative markers is needed. METHODS: We constructed a MALDI-TOF-MS-based approach for identifying various ß-globin disorders and classifying thalassemia major (TM) and thalassemia intermedia (TI) patients using 901 training samples with known HBB/HBA genotypes. We then validated the accuracy of population screening and clinical classification in 2 separate cohorts consisting of 16 172 participants and 201 ß-thalassemia patients. Traditional methods were used as controls. Genetic tests were considered the gold standard for testing positive specimens. RESULTS: We established a prediction model for identifying different forms of ß-globin disorders in a single MALDI-TOF-MS test based on δ- to ß-globin, γ- to α-globin, γ- to ß-globin ratios, and/or the abnormal globin-chain patterns. Our validation study yielded comparable results of clinical specificity (99.89% vs 99.71%), and accuracy (99.78% vs 99.16%) between the new assay and traditional methods but higher clinical sensitivity for the new method (97.52% vs 88.01%). The new assay identified 22 additional abnormal hemoglobins in 69 individuals including 9 novel ones, and accurately screened for 9 carriers of deletional hereditary persistence of fetal hemoglobin or δß-thalassemia. TM and TI were well classified in 178 samples out of 201 ß-thalassemia patients. CONCLUSIONS: MALDI-TOF-MS is a highly accurate, predictive tool that could be suitable for large-scale screening and clinical classification of ß-globin disorders.

miR­302d­3p regulates the viability, migration and apoptosis of breast cancer cells through regulating the TMBIM6­mediated ERK signaling pathway.

MicroRNAs (miRs/miRNAs) play important roles in the occurrence, metastasis and prognosis of multiple types of cancers. However, the specific role of miR­302d­3p and its underlying mechanism in breast cancer (BC) have not yet been reported. The present study aimed to identify the role of miR­302D­3p in BC and its potential mechanism using BC cell lines MCF7 and MDA­MB­231 and normal breast epithelial cell MCF­10A. Cancer and paracancerous tissue from patients with BC were also used. Reverse transcription­quantitative PCR was performed to detect the expression of miR­302d­3p and transmembrane Bax inhibitor motif containing 6 (TMBIM6). Dual­luciferase reporter assays verified the binding sites of miR­302d­3p and TMBIM6. Immunohistochemistry was used to measure the expression of TMBIM6. Cell transfection techniques were used to overexpress or interfere with miR­302d­3p and TMBIM6. A Cell Counting Kit­8 assay was performed to detect cell viability, and migration was measured using a wound healing assay. Apoptosis was detected by flow cytometry. The expression levels of apoptosis­related proteins and pathway­related proteins were detected by western blotting. The expression of miR­302d­3p in BC cell lines was found to be downregulated. It was also demonstrated that miR­302d­3p could inhibit cell viability and migration and promote apoptosis. The expression of TMBIM6 in BC cell lines and tissues was upregulated. Upregulated miR­302d­3p was shown to inhibit viability and migration, and promote apoptosis by targeting TMBIM6, during which extracellular signal­regulated kinase (ERK) and its phosphorylation were inhibited in the ERK signaling pathway in cells. Overall, the present study demonstrated that miR­302d­3p could regulate the viability, migration and apoptosis of BC cells through regulating TMBIM6­mediated ERK signaling pathway.