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. 2021 Jun 10;107(4):909–920. doi: 10.3324/haematol.2021.278469

Figure 8.

Figure 8.

ADAM10 inhibitors and brentuximab-vedotin induce caspase-3 activation in Hodgkin lymphoma cells and reduce glucose consumption in repopulated scaffolds. AviteneTM repopulated scaffolds (LN-MSC23274+L428 cells), were either untreated (A) and nil (C to F) or exposed to brentuximab-vedotin (BtxVed) (20 mg/mL or 2 mg/mL as indicated) (B to D) or 10 mM LT4 or MN8 (C to F) or BtxVed plus one of the inhibitors as indicated (C to F). (A and B) After 96 hours (h), scaffolds were fixed and 5 mm serial sections were stained in immunohistochemistry (IHC) with the rabbit monoclonal anti-caspase-3 antibody. Images were acquired with the Aperio AT2 Digital Pathology Scanner and data analyzed with the Genie software to identify and count caspase-3+cells. Subpanel a: sections of the whole scaffold; subpanel b: enlargements of squares in subpanel a; subpanel c: Hodgkin lymphoma (HL) cells, identified by morphology (blue), and caspase-3+ cells recognized by the Genie software (red) (C and D) percentage of caspase-3+ cells (C: L428, D: L540) counted in serial sections (3 every 15 mm/each scaffold) by the Genie software, are reported as the mean ± standard deviation (SD) of three serial sections analyzed from three different experiments in duplicate (2 scaffolds/experiment). (C) **P<0.0005 and *P<0.05 vs. nil; (D) **P<0.05 vs. BtxVed and *P<0.05 vs. nil. (E and F) Glucose evaluation in the supernatant (SN) recovered from LNMSC23274+ L428 (E) or LN-MSC23274+L540 (F) repopulated scaffolds exposed for the indicated time periods to either 2 mg/mL BtxVed, 10 mM LT4 or MN8, or LT4+BtxVed or MN8+BtxVed. Green symbols: medium containing dimethyl sulfoxide (DMSO) 1:1,000. Glucose was measured with the specific kit (Megazyme) in culture SN recovered every 24 h and data are expressed as percentage glucose consumpion referred to the content in fresh culture medium; mean ± SD from three experiments performed in duplicate (2 scaffolds/experiment). (E) *P<0.02 and **P<0.001 vs. nil; ***P<0.0001 vs. nil and vs. BtxVed. (F) *P<0.01, **P<0.001 and ***P<0.0001 vs. nil and vs. BtxVed.