1. What are miRNAs, and how do they regulate gene expression?
Micro RNAs are small RNA molecules that are non-coding and that are existent in plants, viruses, and animals and that silence RNA. The regulation occurs cleavage of a messenger RNA strand into double pieces (Ma et al., 2018). Additionally, the messenger RNA poly-A tail shortens, and the translation into protein becomes inefficient.
2. How does miR-10b regulate metastasis? What is the role of Twist?
MiR-10B regulates metastasis by inhibiting the translation of messenger RNA encoding homeobox protein. This inhibition causes increased repression of RHOC, which is a pro-metastatic gene (Ma et al., 2018). Twist orchestrates epithelial-mesenchymal transitions and causes multiple characteristics of high-grade malignancy to carcinoma cells.
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3. Explain how it is possible to inhibit miR-10b in cells specifically.
The inhibition of miR-10B in cells is possible by culturing or treating cells with antagomir-10b. This culturing ensures the silencing of miR-10b in the cells, which reduces motility in the cells (Ma et al., 2018).
4. Why did the authors examine Hox10 expression? How does inhibition of the miR-10b affect cultured 4T1 cells?
The purpose of examining Hox10 was to analyze the behavior of the miR-10b, which had been taken by cells in the quickly developing tumors. Hence, this way, it would be easy to understand in what manner the miR-10b acts (Ma et al., 2018). The inhibition enhances change in metastasis-suppressing effect in the cultured 4TI cells reducing the suppressing impact.
5. Why did the authors examine tumor incidence in the lungs if they are studying breast cancer cells?
The examination of incidence in lungs was to use lungs as a control subject to understand the correlation of levels of miRNA in primary tumors and pulmonary metastasis population in the two groups of mice used. Therefore, the use of the lungs and liver was for comparison to the incidence in mammary cells.
6. How did they determine that miR-10b expression was indeed inhibited in mice? Is it possible that nonspecific effects caused the change in Hox10 expression levels? Use data to support your answers.
The inhibition was determined through comparison with the levels of miR-10b from PBS –treated mice (Ma et al., 2018). Essentially, there is no possibility that nonspecific effects were the cause of the change in Hox10 expression levels. This fact is because the mutant miR-10b antagomir was synthesized. Substantially, the miR-10b antagomir did not match a particular sequence in the genome of mice.
7. Does inhibition of miR-10b expression affect primary tumors? Use data to support your answer.
Significantly, the inhibition of miR-10b does not affect the primary tumors. In the experiment, the miR-10b expression was reduced by 62% as a result of sponge expression (Ma et al., 2018). However, despite this action, the primary mammary tumors formed by the sponge that had infection remained the same. There was absolutely no change in the size of these tumors. The controlling or reduction of the miR-10b was affected through the silencing of the miR-10b in tumor cells.
8. The authors claim that inhibition of miR-10b expression does not adversely affect healthy mice. What evidence supports or refutes this claim?
The inhibition of the expression of miR-10b does not in any way affect the normal mice. This information can be subject to assessment through the test carried out to analyze the effect of miR-10b antagomir (Ma et al., 2018). The normal mice were introduced to antagomir in a similar amount, as explained in the metastasis study.
Mostly, miR-10b saw a reduction of 72% in the liver of the mice as a result of exposure to antagomir. At the end of the test, there was no notable change in all the groups of mice, including the healthy group (Ma et al., 2018). The behavior of the mice was still normal regarding the levels of activity and grooming, which were always constant. The weights of the lungs, heart, and body were still the same throughout the procedure. There were also no lobular or portal inflammations, and there was no steatosis in the livers. Despite a slight change in the number of white cells in comparison with the first number, the normal mice were healthy.
Reference
Ma L, Reinhardt F, Pan E, Soutschek J, Bhat B, Marcusson EG, Teruya-Feldstein J, Bell GW, and Weinberg RA. (2010)Therapeutic silencing of miR-10b inhibits metastasis in a mouse mammary tumor model. Nature Biotechnology, 28: 341-7.
