In combination with another mouse macrophage marker, F4/80 [32], we verified that these cells are macrophages and that a significant recruitment to the lungs appeared as early as 36 hours after introduction of the tumor cells into the circulation (Figure 3G).
We verified that these cells were ISCs by staining for Delta and with Nile red.
Importantly, the current study verified these experimental findings in cells obtained from lcSSc patients.
We therefore examined Fgf8 expression to verify that these cells are 'ectopic IHCs' as Fgf8 is a marker of IHCs of the organ of Corti [44].
We therefore optimized a dual-fluorescent MUC1(Alexa488 -EPISPOTexa488)-EPISPOtoto verify whether these cells were able to release both proteins simultaneously.
First, we injected 4 × 106 G2B-10A cells alone to verify that these cells were not tumorigenic, and indeed, G2B-10A cells did not form any tumors.
This sphere-derived monolayer culture is referred to as SAR-OS cells and was expanded and used in some subsequent studies to verify if these cells derived directly from sarcospheres acquired the same biological behavior as parental MNNG/HOS cells.
Thus, further studies are needed to explain the mechanisms underlying this recovery, verifying whether these cells are able to induce structural plasticity after ablation, and/or whether other mechanisms are lying beneath this therapeutic effect.
Indeed, in triple transgenic P2b::FRTCreTau1 FRTCre ; Tau GFPnLacZ embryos, brainstem expression of the Tau GFPnLacZ reporter was specific for periV and periVII cells, verifying that these cells are the only ones with a history of Phox2b and Atoh1 expression (hereafter Phox2b on /Atoh1 on cells).
While we have also sequenced the transcript from these cell pools to verify that these cells were indeed transfected with the said expression vectors (data not shown), this protein level data demonstrates that these cell pools were also producing the respective dhfr proteins with the engineered modifications.
