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Integrin α6A splice variant regulates proliferation and the Wnt/β-catenin pathway in human colorectal cancer cells.

Groulx JF, Giroux V, Beauséjour M, Boudjadi S, Basora N, Carrier JC, Beaulieu JF - Carcinogenesis (2014)

Bottom Line: The α6A silencing was also found to be associated with a significant repression of a number of Wnt/β-catenin pathway end points.Moreover, it was accompanied by a reduction in the capacity of these cells to develop tumours in xenografts.Taken together, these results demonstrate that the α6A variant is a pro-proliferative form of the α6 integrin subunit in CRC cells and appears to mediate its effects through the Wnt/β-catenin pathway.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Intestinal Physiopathology, Department of Anatomy and Cell Biology and Department of Medicine, Faculty of Medicine and Health Sciences, Université de Sherbrooke, Sherbrooke, Quebec J1H 5N4, Canada.

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Knockdown of α6A variant in human CRC cells inhibits their growth in xenografts. (A and B) Tumour growth (mm3) following subcutaneous injection of 2×106 T84 and HT29 stably expressing shα6A and shctl cells into nude mice. Tumour volumes were determined by external measurement [V = (d2 × D)/2]. (C and D) Weight (g) of tumours from T84 and HT29 shctl and shα6A cells at the time of killing. Statistical analysis between shctrl and shα6A: *P ≤ 0.05, **P ≤ 0.01, t-test, n = 4. (E) Representative haematoxylin and eosin staining images for T84, HT29 and DLD-1 shctl and shα6A xenograft tumours. Scale bars = 200 μm for main panels and 50 μm for inserts. Hash symbols denote necrosis/oedema regions. (F) qPCR analyses for the expression of α6A and α6B transcript levels in xenograft tumours from T84, HT29 and DLD-1 shctl and shα6A cells. Data are expressed by α6A normalized to α6B levels. **P ≤ 0.01, §P = 0.0783, t-test, n = 4.
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Figure 4: Knockdown of α6A variant in human CRC cells inhibits their growth in xenografts. (A and B) Tumour growth (mm3) following subcutaneous injection of 2×106 T84 and HT29 stably expressing shα6A and shctl cells into nude mice. Tumour volumes were determined by external measurement [V = (d2 × D)/2]. (C and D) Weight (g) of tumours from T84 and HT29 shctl and shα6A cells at the time of killing. Statistical analysis between shctrl and shα6A: *P ≤ 0.05, **P ≤ 0.01, t-test, n = 4. (E) Representative haematoxylin and eosin staining images for T84, HT29 and DLD-1 shctl and shα6A xenograft tumours. Scale bars = 200 μm for main panels and 50 μm for inserts. Hash symbols denote necrosis/oedema regions. (F) qPCR analyses for the expression of α6A and α6B transcript levels in xenograft tumours from T84, HT29 and DLD-1 shctl and shα6A cells. Data are expressed by α6A normalized to α6B levels. **P ≤ 0.01, §P = 0.0783, t-test, n = 4.

Mentions: The capacity of α6A knockdown cells to form tumours in vivo was next evaluated by subcutaneous injection of nude mice with T84, HT29 and DLD-1 cells. Caco-2/15 cells were not included in this assay because of the long latency period required to observe tumour formation in nude mice with this cell line. Interestingly, we found that the latency period for the detection of palpable tumours was significantly delayed for T84/shα6A cells compared with T84/shctl (36 days versus 12 days) (Figure 4A), whereas this was not so for HT29 and DLD-1 cells. However, abolition of α6A in T84 and HT29 strongly diminished their growth capacity as tumours in nude mice (Figure 4A and B), resulting in a significant reduction of the tumour weight at the time of the killing (Figure 4C and D). The decrease in proliferation rate observed in DLD-1 shα6A cells in vitro was not transposed into a significant reduction in tumour growth and weight (data not shown). However, histological haematoxylin and eosin analysis showed that DLD-1 shctl xenograft tumours displayed large necrosis/oedema regions, a feature not observed in DLD-1 shα6A xenograft tumours (Figure 4E). This observation could explain the lack of difference in tumour size development observed, despite the decrease in proliferation in DLD-1 shα6A cells. On the other hand, no histological difference was observed between shctl and shα6A xenograft tumours from T84 and HT29 cells (Figure 4E). As shown in Figure 4F, qPCR analysis of α6A and α6B in tumours confirmed that the α6A knockdown is retained in HT29 and DLD-1 and tends to be retained in T84 xenogafts even after 50 days (P < 0.01 for HT29 and DLD-1; P < 0.08 for T84). Taken together, these results demonstrate that the α6A variant can regulate tumour growth in at least a subset of CRC cell lines in xenografts, confirming the pro-proliferative effect of the α6A variant.


Integrin α6A splice variant regulates proliferation and the Wnt/β-catenin pathway in human colorectal cancer cells.

Groulx JF, Giroux V, Beauséjour M, Boudjadi S, Basora N, Carrier JC, Beaulieu JF - Carcinogenesis (2014)

Knockdown of α6A variant in human CRC cells inhibits their growth in xenografts. (A and B) Tumour growth (mm3) following subcutaneous injection of 2×106 T84 and HT29 stably expressing shα6A and shctl cells into nude mice. Tumour volumes were determined by external measurement [V = (d2 × D)/2]. (C and D) Weight (g) of tumours from T84 and HT29 shctl and shα6A cells at the time of killing. Statistical analysis between shctrl and shα6A: *P ≤ 0.05, **P ≤ 0.01, t-test, n = 4. (E) Representative haematoxylin and eosin staining images for T84, HT29 and DLD-1 shctl and shα6A xenograft tumours. Scale bars = 200 μm for main panels and 50 μm for inserts. Hash symbols denote necrosis/oedema regions. (F) qPCR analyses for the expression of α6A and α6B transcript levels in xenograft tumours from T84, HT29 and DLD-1 shctl and shα6A cells. Data are expressed by α6A normalized to α6B levels. **P ≤ 0.01, §P = 0.0783, t-test, n = 4.
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Figure 4: Knockdown of α6A variant in human CRC cells inhibits their growth in xenografts. (A and B) Tumour growth (mm3) following subcutaneous injection of 2×106 T84 and HT29 stably expressing shα6A and shctl cells into nude mice. Tumour volumes were determined by external measurement [V = (d2 × D)/2]. (C and D) Weight (g) of tumours from T84 and HT29 shctl and shα6A cells at the time of killing. Statistical analysis between shctrl and shα6A: *P ≤ 0.05, **P ≤ 0.01, t-test, n = 4. (E) Representative haematoxylin and eosin staining images for T84, HT29 and DLD-1 shctl and shα6A xenograft tumours. Scale bars = 200 μm for main panels and 50 μm for inserts. Hash symbols denote necrosis/oedema regions. (F) qPCR analyses for the expression of α6A and α6B transcript levels in xenograft tumours from T84, HT29 and DLD-1 shctl and shα6A cells. Data are expressed by α6A normalized to α6B levels. **P ≤ 0.01, §P = 0.0783, t-test, n = 4.
Mentions: The capacity of α6A knockdown cells to form tumours in vivo was next evaluated by subcutaneous injection of nude mice with T84, HT29 and DLD-1 cells. Caco-2/15 cells were not included in this assay because of the long latency period required to observe tumour formation in nude mice with this cell line. Interestingly, we found that the latency period for the detection of palpable tumours was significantly delayed for T84/shα6A cells compared with T84/shctl (36 days versus 12 days) (Figure 4A), whereas this was not so for HT29 and DLD-1 cells. However, abolition of α6A in T84 and HT29 strongly diminished their growth capacity as tumours in nude mice (Figure 4A and B), resulting in a significant reduction of the tumour weight at the time of the killing (Figure 4C and D). The decrease in proliferation rate observed in DLD-1 shα6A cells in vitro was not transposed into a significant reduction in tumour growth and weight (data not shown). However, histological haematoxylin and eosin analysis showed that DLD-1 shctl xenograft tumours displayed large necrosis/oedema regions, a feature not observed in DLD-1 shα6A xenograft tumours (Figure 4E). This observation could explain the lack of difference in tumour size development observed, despite the decrease in proliferation in DLD-1 shα6A cells. On the other hand, no histological difference was observed between shctl and shα6A xenograft tumours from T84 and HT29 cells (Figure 4E). As shown in Figure 4F, qPCR analysis of α6A and α6B in tumours confirmed that the α6A knockdown is retained in HT29 and DLD-1 and tends to be retained in T84 xenogafts even after 50 days (P < 0.01 for HT29 and DLD-1; P < 0.08 for T84). Taken together, these results demonstrate that the α6A variant can regulate tumour growth in at least a subset of CRC cell lines in xenografts, confirming the pro-proliferative effect of the α6A variant.

Bottom Line: The α6A silencing was also found to be associated with a significant repression of a number of Wnt/β-catenin pathway end points.Moreover, it was accompanied by a reduction in the capacity of these cells to develop tumours in xenografts.Taken together, these results demonstrate that the α6A variant is a pro-proliferative form of the α6 integrin subunit in CRC cells and appears to mediate its effects through the Wnt/β-catenin pathway.

View Article: PubMed Central - PubMed

Affiliation: Laboratory of Intestinal Physiopathology, Department of Anatomy and Cell Biology and Department of Medicine, Faculty of Medicine and Health Sciences, Université de Sherbrooke, Sherbrooke, Quebec J1H 5N4, Canada.

Show MeSH
Related in: MedlinePlus