Abstract
Several studies have shown that brain and muscle aryl hydrocarbon receptor nuclear translocator-like 1 (BMAL1), an important molecule for maintaining circadian rhythms, inhibits the growth and metastasis of tumor cells in several types of cancer, including lung, colon, and breast cancer. However, its role in glioblastoma has not yet been established. Here, we addressed the function of BMAL1 in U87MG glioblastoma cells with two approaches—loss and gain of function. In the loss of function experiments, cell proliferation in U87MG cells transfected with small interfering RNA (siRNA) targeting BMAL1 was increased by approximately 24% (small interfering (si)-NC 0.91 ± 0.00 vs. si-BMAL1 1.129 ± 0.08) via upregulation of cyclin B1. In addition, cell migration and invasion of BMAL1 siRNA-treated glioblastoma cells were elevated by approximately 20% (si-NC 51.00 ± 1.53 vs. si-BMAL161.33 ± 0.88) and 209% (si-NC 21.28 ± 1.37 vs. si-BMAL1 44.47 ± 3.48), respectively, through the accumulation of phosphorylated-AKT (p-AKT) and matrix metalloproteinase (MMP)-9. Gain of function experiments revealed that adenovirus-mediated ectopic expression of BMAL1 in U87MG cells resulted in a 19% (Adenovirus (Ad)-vector 0.94± 0.03 vs. Ad-BMAL1 0.76 ± 0.03) decrease in cell proliferation compared with the control via downregulation of cyclin B1 and increased early and late apoptosis due to changes in the levels of BCL2-associated X protein (BAX), B-cell lymphoma 2 (BCL-2), and cleaved caspase-3. Likewise, cell migration and invasion were attenuated by approximately 24% (Ad-vector 55.00 ± 0.00 vs. Ad-BMAL1 41.83 ± 2.90) and 49% (Ad-vector 70.01 ± 1.24 vs. Ad-BMAL1 35.55 ± 1.78), respectively, in BMAL1-overexpressing U87MG cells following downregulation of p-AKT and MMP-9. Taken together, our results suggest that BMAL1 acts as an anti-cancer gene by altering the proliferation, migration, and invasion of glioblastoma cells. Therefore, the BMAL1 gene could be a potential therapeutic target in the treatment of glioblastoma.
Highlights
Glioblastoma is the most malignant type of tumor in the central nervous system, largely originating from astrocytes, and occurs at a rate of 3 per 100,000 people per year [1]
Our results suggest that the accumulation of cyclin B1 protein might contributed to the increased growth ofOBuMr AreLs1ulstsiRsNugAg-etsrtanthsfaetcttehde Uac8c7uMmuGlacteiollns. of cyclin B1 protein might contributed to the increased growth of BMAL1 small interfering RNA (siRNA)-transfected U87MG cells
BMAL1 Silencing Increased the Cell Migration and Invasion in BMAL1 Small Interfering RNA (SiRNA)-Treated U87MG Cells we investigated whether BMAL1 silencing could influence cell migration using a wound healing asNsaeyx.t,MwiegrinavtieosntigoafteBdMwAhLet1hseirRBNMAA-Ltr1asnilsefneccitnegdcUou8l7dMinGfluceenllcsewcealsl minicgrreaatisoendubsyinagpapwrooxuinmdately 20% cohapemapplrionaxgriemdaawstseialtyyh.2N0M%CiscgiorRmaNtpioAanr-etdroawfnsitBfheMcNtAeCdLs1ciRelNslisRA(Ns-tirA-aN-ntCrsaf5en1cst.f0eedc±tce1ed.l5ls3U(vs8is-7.NsMiC-GB5M1.0cAe±lLl1s1.563w1v.a3ss3.s±i-iBn0cM.8reA8a)Ls(e1Fdi6g1u.b3ry3e 3A)
Summary
Glioblastoma is the most malignant type of tumor in the central nervous system, largely originating from astrocytes, and occurs at a rate of 3 per 100,000 people per year [1]. TMZ alkylates or methylates the N-7 or O-6 positions, respectively, of guanine residues in DNA and triggers tumor cell death, and this drug is favored for the treatment of malignant brain tumors such as astrocytoma and glioblastoma. TMZ provides an anti-tumor effect by inducing the damage-associated molecular patterns from glioblastoma and enhancing the tumor-specific immune responses [2]. Glioblastoma is occasionally insensitive to TMZ as some tumor cells express O6-alkylguanine DNA alkyltransferase, which allows repair of this type of DNA damage [4]. The development of new innovative medicine for the treatment of glioblastoma with less toxicity and high efficacy is urgently required
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