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. 2023 Mar 6;28(5):2412.
doi: 10.3390/molecules28052412.

LncRNA JHDM1D-AS1 Is a Key Biomarker for Progression and Modulation of Gemcitabine Sensitivity in Bladder Cancer Cells

Isadora Oliveira Ansaloni Pereira  1 , Glenda Nicioli da Silva  1 , Tamires Cunha Almeida  2 , Ana Paula Braga Lima  1 , André Luiz Ventura Sávio  3   4 , Katia Ramos Moreira Leite  5 , Daisy Maria Fávero Salvadori  6
Affiliations

LncRNA JHDM1D-AS1 Is a Key Biomarker for Progression and Modulation of Gemcitabine Sensitivity in Bladder Cancer Cells

Isadora Oliveira Ansaloni Pereira et al. Molecules. .

Abstract

Long non-coding RNAs are frequently found to be dysregulated and are linked to carcinogenesis, aggressiveness, and chemoresistance in a variety of tumors. As expression levels of the JHDM1D gene and lncRNA JHDM1D-AS1 are altered in bladder tumors, we sought to use their combined expression to distinguish between low-and high-grade bladder tumors by RTq-PCR. In addition, we evaluated the functional role of JHDM1D-AS1 and its association with the modulation of gemcitabine sensitivity in high-grade bladder-tumor cells. J82 and UM-UC-3 cells were treated with siRNA-JHDM1D-AS1 and/or three concentrations of gemcitabine (0.39, 0.78, and 1.56 µM), and then submitted to cytotoxicity testing (XTT), clonogenic survival, cell cycle progression, cell morphology, and cell migration assays. When JHDM1D and JHDM1D-AS1 expression levels were used in combination, our findings indicated favorable prognostic value. Furthermore, the combined treatment resulted in greater cytotoxicity, a decrease in clone formation, G0/G1 cell cycle arrest, morphological alterations, and a reduction in cell migration capacity in both lineages compared to the treatments alone. Thus, silencing of JHDM1D-AS1 reduced the growth and proliferation of high-grade bladder-tumor cells and increased their sensitivity to gemcitabine treatment. In addition, the expression of JHDM1D/JHDM1D-AS1 indicated potential prognostic value in the progression of bladder tumors.

Keywords: JHDM1D-AS1; bladder cancer; long non-coding RNAs.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
Expression of the JHDM1D gene (a) and JHDM1D-AS1 lncRNA (b) in patients with low- and high-grade bladder tumors. (c) Moderately positive correlation between JHDM1D and JHDM1D-AS1 expression (r = 0.7204) in low- and high-grade bladder tumor samples (Spearman correlation analysis). (d) Receiver operating characteristics (ROC) curves using the combination between JHDM1-AS1 and JHDM1D to distinguish between low- and high-grade bladder tumors. * p < 0.05.
Figure 2
Figure 2
Percentage of viability in J82 (a) and UM-UC-3 (b) cell lines after treatment with gemcitabine, siRNA-JHDM1D-AS1, or gemcitabine combined with siRNA-JHDM1D-AS1. Control: untreated cells; gem: gemcitabine. a: p < 0.05 compared to control; b: p < 0.05 compared to siRNA; c: p < 0.05 compared to respective gemcitabine concentration alone.
Figure 3
Figure 3
Morphologies of J82 (a) and UM-UC-3 (b) cells after treatment with gemcitabine, siRNA-JHDM1D-AS1, or gemcitabine combined with siRNA-JHDM1D-AS1. Control: untreated cells; gem: gemcitabine. Black arrows: elongated cells; white arrows: round cells; red arrows: dead cells. Phase-contrast microscope, ×200.
Figure 4
Figure 4
Percentages of J82 (a) and UM-UC-3 (b) colonies after treatment with gemcitabine, siRNA JHDM1D-AS1, or gemcitabine combined with siRNA JHDM1D-AS1. Control: untreated cells; gem: gemcitabine. a: p < 0.05 compared to control; b: p < 0.05 compared to siRNA; c: p < 0.05 compared to respective gemcitabine concentration alone.
Figure 5
Figure 5
Representative histograms and percentages of J82 cells in each phase after treatment with gemcitabine, siRNA JHDM1D-AS1, or gemcitabine combined with siRNA JHDM1D-AS1. Control: untreated cells; gem: gemcitabine; IP: propidium iodide. * p < 0.05 compared to control.
Figure 6
Figure 6
Representative histograms and percentages of UM-UC-3 cells in each phase after treatment with gemcitabine, siRNA JHDM1D-AS1, or gemcitabine combined with siRNA JHDM1D-AS1. Control: untreated cells; gem: gemcitabine; IP: propidium iodide. * p < 0.05 compared to control.
Figure 7
Figure 7
Photographs (a) and quantification (b,c) of cell migration in J82 cells after treatment with gemcitabine, siRNA JHDM1D-AS1, or gemcitabine combined with siRNA JHDM1D-AS1. Control: untreated cells; gem: gemcitabine. a: p < 0.05 compared to control; b: p < 0.05 compared to siRNA; c: p < 0.05 compared to respective gemcitabine concentration alone.
Figure 8
Figure 8
Photographs (a) and quantification (b,c) of cell migration in UM-UC-3 cells after treatment with gemcitabine, siRNA JHDM1D-AS1, or gemcitabine combined with siRNA JHDM1D-AS1. Control: untreated cells; gem: gemcitabine. a: p < 0.05 compared to control; b: p < 0.05 compared to siRNA; c: p < 0.05 compared to respective gemcitabine concentration alone.
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