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Hsa-miR-125a-3p and hsa-miR-125a-5p are downregulated in non-small cell lung cancer and have inverse effects on invasion and migration of lung cancer cells.

Jiang L, Huang Q, Zhang S, Zhang Q, Chang J, Qiu X, Wang E - BMC Cancer (2010)

Bottom Line: Furthermore, the results from the Spearman correlation test showed a negative relationship between hsa-miR-125a-3p expression and pathological stage or lymph node metastasis and an inverse relationship between hsa-miR-125a-5p expression and pathological stage or lymph node metastasis.In vitro gain-of-function experiments indicated that hsa-miR-125a-3p and hsa-miR-125a-5p function in an opposing manner, suppressing or enhancing cell migration and invasion in A549 and SPC-A-1 cell lines, respectively.These opposing functions were further validated by suppression of hsa-miR-125a-3p and hsa-miR-125a-5p expression in loss-of-function experiments.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Pathology, the First Affiliated Hospital of China Medical University, Heping District, Shenyang, Liaoning 110001, China.

ABSTRACT

Background: Two mature microRNAs (miRNAs), hsa-miR-125a-3p and hsa-miR-125a-5p (collectively referred to as hsa-miR-125a-3p/5p), are derived from 3' and 5' ends of pre-miR-125a, respectively. Although impaired regulation of hsa-miR-125a-5p has been observed in some tumors, the role of this miRNA in invasion and metastasis remains unclear, and few studies have examined the function of hsa-miR-125a-3p. In order to characterize the functions of hsa-miR-125a-3p/5p in invasion and metastasis of non-small cell lung cancer (NSCLC), we investigated the relationships between hsa-miR-125a-3p/5p expression and lymph node metastasis in NSCLC tissues. We also explored the impact of expression of these miRNAs on invasive and migratory capabilities of lung cancer cells.

Methods: Expression of hsa-miR-125a-3p/5p in NSCLC tissues was explored using real-time PCR. The relationships between hsa-miR-125a-3p/5p expression and pathological stage or lymph node metastasis were assessed using the Spearman correlation test. For in vitro studies, lung cancer cells were transfected with sense and antisense 2'-O-methyl oligonucleotides for gain-of-function and loss-of-function experiments. Transwell experiments were performed to evaluate cellular migration and invasion.

Results: Expression of hsa-miR-125a-3p/5p was lower in NSCLC tissues than in adjacent normal lung tissues (LAC). Furthermore, the results from the Spearman correlation test showed a negative relationship between hsa-miR-125a-3p expression and pathological stage or lymph node metastasis and an inverse relationship between hsa-miR-125a-5p expression and pathological stage or lymph node metastasis. In vitro gain-of-function experiments indicated that hsa-miR-125a-3p and hsa-miR-125a-5p function in an opposing manner, suppressing or enhancing cell migration and invasion in A549 and SPC-A-1 cell lines, respectively. These opposing functions were further validated by suppression of hsa-miR-125a-3p and hsa-miR-125a-5p expression in loss-of-function experiments.

Conclusion: Hsa-miR-125a-3p and hsa-miR-125a-5p play distinct roles in regulation of invasive and metastatic capabilities of lung cancer cells, consistent with the opposing correlations between the expression of these miRNAs and lymph node metastasis in NSCLC. These results provide new insights into the roles of miR-125a family members in the development of NSCLC.

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Effects of loss-of-function of hsa-miR-125a-3p/5p on migration and invasion of lung cancer cells. Migration assay results showed that the number of A549 cells that migrated through the microporous membrane was significantly increased in the antisense-3p group (p < 0.001). However, the number of migratory SPC-A-1 cells was significantly decreased in the antisense-5p group (p < 0.001, bar = 20 μm). Invasion assay results showed that the number of A549 cells that invaded through the Matrigel was significantly increased in the antisense-3p group (p < 0.001). In contrast, the number of invasive SPC-A-1 cells was significantly decreased in the antisense-5p group (p < 0.001, bar = 20 μm). The results are representative of three independent experiments.
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Figure 8: Effects of loss-of-function of hsa-miR-125a-3p/5p on migration and invasion of lung cancer cells. Migration assay results showed that the number of A549 cells that migrated through the microporous membrane was significantly increased in the antisense-3p group (p < 0.001). However, the number of migratory SPC-A-1 cells was significantly decreased in the antisense-5p group (p < 0.001, bar = 20 μm). Invasion assay results showed that the number of A549 cells that invaded through the Matrigel was significantly increased in the antisense-3p group (p < 0.001). In contrast, the number of invasive SPC-A-1 cells was significantly decreased in the antisense-5p group (p < 0.001, bar = 20 μm). The results are representative of three independent experiments.

Mentions: To further examine whether endogenous hsa-miR-125a-3p and hsa-miR-125a-5p regulate migration and invasion, we adopted a loss-of-function approach that blocked the function of endogenous hsa-miR-125a-3p and hsa-miR-125a-5p using antisense 2'-O-methyl oligonucleotides [23-26]. We then analyzed the migratory and invasive capabilities of these cells using the methods described above. In the hsa-miR-125a-3p migration assay, the number of A549 cells in the untreated group that migrated through the microporous membrane was 31.00 ± 1.54. There was no difference between the untreated and scramble-3p-transfected cells (32.00 ± 0.58, p = 0.347). The number of migrating cells was significantly increased when A549 cells were transfected with the antisense-3p oligonucleotide (46.00 ± 3.16, p < 0.001). For the hsa-miR-125a-5p migration assay, the number of SPC-A-1 cells in the untreated group that migrated through the microporous membrane was 22.60 ± 2.07. No difference was observed between untreated and scramble-5p-transfected cells (22.40 ± 1.14, p = 0.855). The number of migrating cells was significantly decreased when SPC-A-1 cells were transfected with the antisense-5p oligonucleotide (15.00 ± 1.58, p < 0.001, Fig 8).


Hsa-miR-125a-3p and hsa-miR-125a-5p are downregulated in non-small cell lung cancer and have inverse effects on invasion and migration of lung cancer cells.

Jiang L, Huang Q, Zhang S, Zhang Q, Chang J, Qiu X, Wang E - BMC Cancer (2010)

Effects of loss-of-function of hsa-miR-125a-3p/5p on migration and invasion of lung cancer cells. Migration assay results showed that the number of A549 cells that migrated through the microporous membrane was significantly increased in the antisense-3p group (p < 0.001). However, the number of migratory SPC-A-1 cells was significantly decreased in the antisense-5p group (p < 0.001, bar = 20 μm). Invasion assay results showed that the number of A549 cells that invaded through the Matrigel was significantly increased in the antisense-3p group (p < 0.001). In contrast, the number of invasive SPC-A-1 cells was significantly decreased in the antisense-5p group (p < 0.001, bar = 20 μm). The results are representative of three independent experiments.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC2903529&req=5

Figure 8: Effects of loss-of-function of hsa-miR-125a-3p/5p on migration and invasion of lung cancer cells. Migration assay results showed that the number of A549 cells that migrated through the microporous membrane was significantly increased in the antisense-3p group (p < 0.001). However, the number of migratory SPC-A-1 cells was significantly decreased in the antisense-5p group (p < 0.001, bar = 20 μm). Invasion assay results showed that the number of A549 cells that invaded through the Matrigel was significantly increased in the antisense-3p group (p < 0.001). In contrast, the number of invasive SPC-A-1 cells was significantly decreased in the antisense-5p group (p < 0.001, bar = 20 μm). The results are representative of three independent experiments.
Mentions: To further examine whether endogenous hsa-miR-125a-3p and hsa-miR-125a-5p regulate migration and invasion, we adopted a loss-of-function approach that blocked the function of endogenous hsa-miR-125a-3p and hsa-miR-125a-5p using antisense 2'-O-methyl oligonucleotides [23-26]. We then analyzed the migratory and invasive capabilities of these cells using the methods described above. In the hsa-miR-125a-3p migration assay, the number of A549 cells in the untreated group that migrated through the microporous membrane was 31.00 ± 1.54. There was no difference between the untreated and scramble-3p-transfected cells (32.00 ± 0.58, p = 0.347). The number of migrating cells was significantly increased when A549 cells were transfected with the antisense-3p oligonucleotide (46.00 ± 3.16, p < 0.001). For the hsa-miR-125a-5p migration assay, the number of SPC-A-1 cells in the untreated group that migrated through the microporous membrane was 22.60 ± 2.07. No difference was observed between untreated and scramble-5p-transfected cells (22.40 ± 1.14, p = 0.855). The number of migrating cells was significantly decreased when SPC-A-1 cells were transfected with the antisense-5p oligonucleotide (15.00 ± 1.58, p < 0.001, Fig 8).

Bottom Line: Furthermore, the results from the Spearman correlation test showed a negative relationship between hsa-miR-125a-3p expression and pathological stage or lymph node metastasis and an inverse relationship between hsa-miR-125a-5p expression and pathological stage or lymph node metastasis.In vitro gain-of-function experiments indicated that hsa-miR-125a-3p and hsa-miR-125a-5p function in an opposing manner, suppressing or enhancing cell migration and invasion in A549 and SPC-A-1 cell lines, respectively.These opposing functions were further validated by suppression of hsa-miR-125a-3p and hsa-miR-125a-5p expression in loss-of-function experiments.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Pathology, the First Affiliated Hospital of China Medical University, Heping District, Shenyang, Liaoning 110001, China.

ABSTRACT

Background: Two mature microRNAs (miRNAs), hsa-miR-125a-3p and hsa-miR-125a-5p (collectively referred to as hsa-miR-125a-3p/5p), are derived from 3' and 5' ends of pre-miR-125a, respectively. Although impaired regulation of hsa-miR-125a-5p has been observed in some tumors, the role of this miRNA in invasion and metastasis remains unclear, and few studies have examined the function of hsa-miR-125a-3p. In order to characterize the functions of hsa-miR-125a-3p/5p in invasion and metastasis of non-small cell lung cancer (NSCLC), we investigated the relationships between hsa-miR-125a-3p/5p expression and lymph node metastasis in NSCLC tissues. We also explored the impact of expression of these miRNAs on invasive and migratory capabilities of lung cancer cells.

Methods: Expression of hsa-miR-125a-3p/5p in NSCLC tissues was explored using real-time PCR. The relationships between hsa-miR-125a-3p/5p expression and pathological stage or lymph node metastasis were assessed using the Spearman correlation test. For in vitro studies, lung cancer cells were transfected with sense and antisense 2'-O-methyl oligonucleotides for gain-of-function and loss-of-function experiments. Transwell experiments were performed to evaluate cellular migration and invasion.

Results: Expression of hsa-miR-125a-3p/5p was lower in NSCLC tissues than in adjacent normal lung tissues (LAC). Furthermore, the results from the Spearman correlation test showed a negative relationship between hsa-miR-125a-3p expression and pathological stage or lymph node metastasis and an inverse relationship between hsa-miR-125a-5p expression and pathological stage or lymph node metastasis. In vitro gain-of-function experiments indicated that hsa-miR-125a-3p and hsa-miR-125a-5p function in an opposing manner, suppressing or enhancing cell migration and invasion in A549 and SPC-A-1 cell lines, respectively. These opposing functions were further validated by suppression of hsa-miR-125a-3p and hsa-miR-125a-5p expression in loss-of-function experiments.

Conclusion: Hsa-miR-125a-3p and hsa-miR-125a-5p play distinct roles in regulation of invasive and metastatic capabilities of lung cancer cells, consistent with the opposing correlations between the expression of these miRNAs and lymph node metastasis in NSCLC. These results provide new insights into the roles of miR-125a family members in the development of NSCLC.

Show MeSH
Related in: MedlinePlus