Figure 6

Hypoxia promotes EMT of LIM1863 human CRC cells. A) EMT of LIM1863 human CRC cells cultured in either normoxia (20% O₂) or hypoxia (1% O₂) in the presence of 0.2 ng/ml TGFβ. Positive control cells underwent EMT in the presence of 2 ng/ml TGFβ. *p = 0.02 for the difference from normoxic cells cultured in the presence of 0.2 ng/ml TGFβ (n = 3). B) Dual immunofluorescence for E-cadherin (red) and 15-PGDH (green) on LIM1863 human CRC cells six days after addition of TGFβ. DAPI was used for nuclear visualisation (blue). LIM 1863 human CRC cells were cultured in the presence of 0.2 ng/ml TGFβ for six days (normoxia) or were transferred to hypoxic conditions (1% O2) on day two. In normoxic conditions, E-cadherin-negative cells at the edge of the colony did not express 15-PGDH (white arrows). However, E-cadherin-low cells at the edge of hypoxic colonies were still 15-PGDH-positive (yellow arrows). C) Immunohistochemistry for 15-PGDH and E-cadherin on sequential sections of a representative central region of CRCLM. Images taken from representative areas of stained sections show an inverse relationship between E-cadherin and 15-PGDH immunoreactivity such that 15-PGDH expression was high in E-cadherin-low cells and vice versa.