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Leptin produced by obese adipose stromal/stem cells enhances proliferation and metastasis of estrogen receptor positive breast cancers.

Strong AL, Ohlstein JF, Biagas BA, Rhodes LV, Pei DT, Tucker HA, Llamas C, Bowles AC, Dutreil MF, Zhang S, Gimble JM, Burow ME, Bunnell BA - Breast Cancer Res. (2015)

Bottom Line: BCCs co-cultured with obASCs demonstrated enhanced expression of epithelial-to-mesenchymal transition (EMT) and metastasis genes (SERPINE1, MMP-2, and IL-6), while BCCs co-cultured with leptin shRNA obASCs did not display similar levels of gene induction.Knockdown of leptin significantly reduced tumor volume and decreased the number of metastatic lesions to the lung and liver.These results correlated with reduced expression of both SERPINE1 and MMP-2 in tumors formed with MCF7 cells mixed with leptin shRNA obASCs, when compared to tumors formed with MCF7 cells mixed with control shRNA obASCs.

View Article: PubMed Central - PubMed

Affiliation: Center for Stem Cell Research and Regenerative Medicine, Tulane University School of Medicine, 1430 Tulane Avenue, SL-99, New Orleans, LA, 70112, USA. alin1@tulane.edu.

ABSTRACT

Introduction: The steady increase in the incidence of obesity among adults has been paralleled with higher levels of obesity-associated breast cancer. While recent studies have suggested that adipose stromal/stem cells (ASCs) isolated from obese women enhance tumorigenicity, the mechanism(s) by which this occurs remains undefined. Evidence suggests that increased adiposity results in increased leptin secretion from adipose tissue, which has been shown to increased cancer cell proliferation. Previously, our group demonstrated that ASCs isolated from obese women (obASCs) also express higher levels of leptin relative to ASCs isolated from lean women (lnASCs) and that this obASC-derived leptin may account for enhanced breast cancer cell growth. The current study investigates the impact of inhibiting leptin expression in lnASCs and obASCs on breast cancer cell (BCC) growth and progression.

Methods: Estrogen receptor positive (ER+) BCCs were co-cultured with leptin shRNA lnASCs or leptin shRNA obASCs and changes in the proliferation, migration, invasion, and gene expression of BCCs were investigated. To assess the direct impact of leptin inhibition in obASCs on BCC proliferation, MCF7 cells were injected alone or mixed with control shRNA obASCs or leptin shRNA obASCs into SCID/beige mice.

Results: ER+ BCCs were responsive to obASCs during direct co-culture, whereas lnASCs were unable to increase ER(+) BCC growth. shRNA silencing of leptin in obASCs negated the enhanced proliferative effects of obASC on BCCs following direct co-culture. BCCs co-cultured with obASCs demonstrated enhanced expression of epithelial-to-mesenchymal transition (EMT) and metastasis genes (SERPINE1, MMP-2, and IL-6), while BCCs co-cultured with leptin shRNA obASCs did not display similar levels of gene induction. Knockdown of leptin significantly reduced tumor volume and decreased the number of metastatic lesions to the lung and liver. These results correlated with reduced expression of both SERPINE1 and MMP-2 in tumors formed with MCF7 cells mixed with leptin shRNA obASCs, when compared to tumors formed with MCF7 cells mixed with control shRNA obASCs.

Conclusion: This study provides mechanistic insight as to how obesity enhances the proliferation and metastasis of breast cancer cells; specifically, obASC-derived leptin contributes to the aggressiveness of breast cancer in obese women.

No MeSH data available.


Related in: MedlinePlus

Leptin inhibition reduces the expression of key regulatory genes involved in invasion and metastasis. Breast cancer cells (BCCs) were co-cultured CC with adipose stromal/stem cells isolated from lean women (lnASCs) or adipose stromal/stem cells isolated from obese women (obASCs) stably transfected with a control short hairpin RNA (control shRNA) or leptin shRNA. After 7 days, co-cultured cells were sorted by FACS and BCCs were collected for total RNA extraction and cDNA synthesis. Genes involved in invasion and metastasis were assessed. Bar represents ± standard error of the mean. *P <0.05, ***P <0.001 relative to BCCs cultured alone; ###P <0.01 for comparison between BCCs after co-culture with control shRNA lnASCs and control shRNA obASCs; ΦΦΦP <0.001 for comparison between BCCs after co-culture with control shRNA obASCs and leptin shRNA obASCs. MMP matrix metalloproteinase
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Fig4: Leptin inhibition reduces the expression of key regulatory genes involved in invasion and metastasis. Breast cancer cells (BCCs) were co-cultured CC with adipose stromal/stem cells isolated from lean women (lnASCs) or adipose stromal/stem cells isolated from obese women (obASCs) stably transfected with a control short hairpin RNA (control shRNA) or leptin shRNA. After 7 days, co-cultured cells were sorted by FACS and BCCs were collected for total RNA extraction and cDNA synthesis. Genes involved in invasion and metastasis were assessed. Bar represents ± standard error of the mean. *P <0.05, ***P <0.001 relative to BCCs cultured alone; ###P <0.01 for comparison between BCCs after co-culture with control shRNA lnASCs and control shRNA obASCs; ΦΦΦP <0.001 for comparison between BCCs after co-culture with control shRNA obASCs and leptin shRNA obASCs. MMP matrix metalloproteinase

Mentions: In contrast, investigation of EMT and metastatic genes previously identified by us to be altered in MCF7 cells after co-culturing with obASCs showed uniform changes in all three BCC lines. Of the genes assessed, BCCs overexpressed SERPINE1, IL-6, and MMP-2 following direct co-culture with obASCs. Furthermore, inhibition of leptin in obASCs diminished the expression of these genes in BCCs (relative to obese ASCs; P <0.05; Fig. 4; Additional files 3–5). No statistically significant differences in gene expression were observed in BCCs after co-culture with control shRNA lnASCs or leptin shRNA lnASCs (Fig. 4; Additional files 3–5). SERPINE1 expression was significantly decreased in MCF7, ZR75, and T47D after leptin knockdown 393.9-fold, 21.7-fold, and 12.6-fold, respectively (P <0.05; Fig. 4; Additional files 3−5). Additionally, leptin inhibition in obASCs limited the induction of IL-6 expression in BCC by the obASCs, with a 2.3-fold decrease in MCF7 cells, 2.0-fold decrease in ZR75, and 1.5-fold decrease in T47D (P <0.05; Fig. 4; Additional files 3–5). Moreover, inhibition of leptin in obASCs resulted in a 27.4-fold, 179.2-fold, and 843.5-fold decrease in MMP-2 expression in MCF7, ZR75, and T47D cells, respectively (P <0.05; Additional files 3–5). Together, these results indicate that obASCs increase the mRNA expression of key metastatic genes in ER+ BCC lines and suggest that obASCs may play a significant role in EMT and metastasis.Fig. 4


Leptin produced by obese adipose stromal/stem cells enhances proliferation and metastasis of estrogen receptor positive breast cancers.

Strong AL, Ohlstein JF, Biagas BA, Rhodes LV, Pei DT, Tucker HA, Llamas C, Bowles AC, Dutreil MF, Zhang S, Gimble JM, Burow ME, Bunnell BA - Breast Cancer Res. (2015)

Leptin inhibition reduces the expression of key regulatory genes involved in invasion and metastasis. Breast cancer cells (BCCs) were co-cultured CC with adipose stromal/stem cells isolated from lean women (lnASCs) or adipose stromal/stem cells isolated from obese women (obASCs) stably transfected with a control short hairpin RNA (control shRNA) or leptin shRNA. After 7 days, co-cultured cells were sorted by FACS and BCCs were collected for total RNA extraction and cDNA synthesis. Genes involved in invasion and metastasis were assessed. Bar represents ± standard error of the mean. *P <0.05, ***P <0.001 relative to BCCs cultured alone; ###P <0.01 for comparison between BCCs after co-culture with control shRNA lnASCs and control shRNA obASCs; ΦΦΦP <0.001 for comparison between BCCs after co-culture with control shRNA obASCs and leptin shRNA obASCs. MMP matrix metalloproteinase
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Fig4: Leptin inhibition reduces the expression of key regulatory genes involved in invasion and metastasis. Breast cancer cells (BCCs) were co-cultured CC with adipose stromal/stem cells isolated from lean women (lnASCs) or adipose stromal/stem cells isolated from obese women (obASCs) stably transfected with a control short hairpin RNA (control shRNA) or leptin shRNA. After 7 days, co-cultured cells were sorted by FACS and BCCs were collected for total RNA extraction and cDNA synthesis. Genes involved in invasion and metastasis were assessed. Bar represents ± standard error of the mean. *P <0.05, ***P <0.001 relative to BCCs cultured alone; ###P <0.01 for comparison between BCCs after co-culture with control shRNA lnASCs and control shRNA obASCs; ΦΦΦP <0.001 for comparison between BCCs after co-culture with control shRNA obASCs and leptin shRNA obASCs. MMP matrix metalloproteinase
Mentions: In contrast, investigation of EMT and metastatic genes previously identified by us to be altered in MCF7 cells after co-culturing with obASCs showed uniform changes in all three BCC lines. Of the genes assessed, BCCs overexpressed SERPINE1, IL-6, and MMP-2 following direct co-culture with obASCs. Furthermore, inhibition of leptin in obASCs diminished the expression of these genes in BCCs (relative to obese ASCs; P <0.05; Fig. 4; Additional files 3–5). No statistically significant differences in gene expression were observed in BCCs after co-culture with control shRNA lnASCs or leptin shRNA lnASCs (Fig. 4; Additional files 3–5). SERPINE1 expression was significantly decreased in MCF7, ZR75, and T47D after leptin knockdown 393.9-fold, 21.7-fold, and 12.6-fold, respectively (P <0.05; Fig. 4; Additional files 3−5). Additionally, leptin inhibition in obASCs limited the induction of IL-6 expression in BCC by the obASCs, with a 2.3-fold decrease in MCF7 cells, 2.0-fold decrease in ZR75, and 1.5-fold decrease in T47D (P <0.05; Fig. 4; Additional files 3–5). Moreover, inhibition of leptin in obASCs resulted in a 27.4-fold, 179.2-fold, and 843.5-fold decrease in MMP-2 expression in MCF7, ZR75, and T47D cells, respectively (P <0.05; Additional files 3–5). Together, these results indicate that obASCs increase the mRNA expression of key metastatic genes in ER+ BCC lines and suggest that obASCs may play a significant role in EMT and metastasis.Fig. 4

Bottom Line: BCCs co-cultured with obASCs demonstrated enhanced expression of epithelial-to-mesenchymal transition (EMT) and metastasis genes (SERPINE1, MMP-2, and IL-6), while BCCs co-cultured with leptin shRNA obASCs did not display similar levels of gene induction.Knockdown of leptin significantly reduced tumor volume and decreased the number of metastatic lesions to the lung and liver.These results correlated with reduced expression of both SERPINE1 and MMP-2 in tumors formed with MCF7 cells mixed with leptin shRNA obASCs, when compared to tumors formed with MCF7 cells mixed with control shRNA obASCs.

View Article: PubMed Central - PubMed

Affiliation: Center for Stem Cell Research and Regenerative Medicine, Tulane University School of Medicine, 1430 Tulane Avenue, SL-99, New Orleans, LA, 70112, USA. alin1@tulane.edu.

ABSTRACT

Introduction: The steady increase in the incidence of obesity among adults has been paralleled with higher levels of obesity-associated breast cancer. While recent studies have suggested that adipose stromal/stem cells (ASCs) isolated from obese women enhance tumorigenicity, the mechanism(s) by which this occurs remains undefined. Evidence suggests that increased adiposity results in increased leptin secretion from adipose tissue, which has been shown to increased cancer cell proliferation. Previously, our group demonstrated that ASCs isolated from obese women (obASCs) also express higher levels of leptin relative to ASCs isolated from lean women (lnASCs) and that this obASC-derived leptin may account for enhanced breast cancer cell growth. The current study investigates the impact of inhibiting leptin expression in lnASCs and obASCs on breast cancer cell (BCC) growth and progression.

Methods: Estrogen receptor positive (ER+) BCCs were co-cultured with leptin shRNA lnASCs or leptin shRNA obASCs and changes in the proliferation, migration, invasion, and gene expression of BCCs were investigated. To assess the direct impact of leptin inhibition in obASCs on BCC proliferation, MCF7 cells were injected alone or mixed with control shRNA obASCs or leptin shRNA obASCs into SCID/beige mice.

Results: ER+ BCCs were responsive to obASCs during direct co-culture, whereas lnASCs were unable to increase ER(+) BCC growth. shRNA silencing of leptin in obASCs negated the enhanced proliferative effects of obASC on BCCs following direct co-culture. BCCs co-cultured with obASCs demonstrated enhanced expression of epithelial-to-mesenchymal transition (EMT) and metastasis genes (SERPINE1, MMP-2, and IL-6), while BCCs co-cultured with leptin shRNA obASCs did not display similar levels of gene induction. Knockdown of leptin significantly reduced tumor volume and decreased the number of metastatic lesions to the lung and liver. These results correlated with reduced expression of both SERPINE1 and MMP-2 in tumors formed with MCF7 cells mixed with leptin shRNA obASCs, when compared to tumors formed with MCF7 cells mixed with control shRNA obASCs.

Conclusion: This study provides mechanistic insight as to how obesity enhances the proliferation and metastasis of breast cancer cells; specifically, obASC-derived leptin contributes to the aggressiveness of breast cancer in obese women.

No MeSH data available.


Related in: MedlinePlus