Breart et al Fig 3: Direct action of CTLs on individual tumor cells drives tumor regression
Breart et al, Fig. 3. Direct action of CTLs on
individual tumor cells drives tumor regression

Intravital microscopy — microscopic analysis, in real time, of processes within a living animal — has been used in immunology for maybe a decade now, but it hasn’t lost its cool factor yet. I don’t know that there have been any great intellectual breakthroughs arising from the work, but we have learned a fair bit about, say, interactions between T cells and their targets, and migration patterns, and so on. And of course there’s a huge help in visualization, which undoubtedly helps people understand what’s happening and, hopefully, develop other experimental approaches to test it.

Just as importantly, the “Awesome” factor of these things is absolutely off the scale. I pointed out Uli von Andrian’s collection of intravital videos the other day, and now the latest issue of Journal of Clinical Investigation has a paper from Phillipe Bousso’s group, showing 2-photon microscopy of cytotoxic T lymphocytes attacking a tumor.

The paper is:

Breart, B., LemaƮtre, F., Celli, S., Bousso, P. (2008). Two-photon imaging of intratumoral CD8+ T cell cytotoxic activity during adoptive T cell therapy in mice. Journal of Clinical Investigation, 118(4), 1390-1397. DOI: 10.1172/JCI34388

They used the EL4/EG7 tumor model in mice. These cells form solid tumors in C57BL/6 mice, and are not rejected by the immune system. The EG7 cells are derived from EL4; they have had a defined antigen introduced, and if you transfer activated T cells against the antigen, the tumor will be rejected. If you transfer naive T cells, and depend on them to be activated by the tumor, you’re out of luck; the tumor is not rejected. 1 They were able to watch all these things happening, in real time.

Here’s what happens with activated CTL (orange) around a tumor site (tumor cells in yellow/greenish). Watch the CTL zipping around merrily in areas where there are no tumor cells, and then screeching to a halt as they identify tumor antigen, engage their targets, and begin to kill:

(Embedded video! I’m so MySpace! I’m going to use tripple exclamation marks and mispell lot’s of words!!!)

The article is free access, I believe, so you should check it out for yourself; there are five videos to watch in the supplemental data. They show CTL engaging tumor cells and actively killing them, using indicators for cell death so they don’t have to guess what’s happening.

I think this is mainly a technical tour de force, and the amount of new information about tumor immunology is relatively small. But there are a couple things of interest. One is that naive T cells — the guys who do not reject the tumor — seem kind of indifferent to the whole thing. It’s not a question of the CTL entering the tumor, and then being turned off (which would have been my guess); rather, the naive cells never even entered the tumor in the first place:

Although CTL infiltration was quite variable in the different regions of the tumor (Figure 6A), EG7 patches were eliminated in CTL-rich areas, which was evidence that in vivo primed CTLs were not grossly impaired in their ability to kill target cells … Thus, the low level of CD8+ T cell infiltration, rather than a defect in the cytotoxic activity, appeared to be responsible for the inefficient response mounted by in vivo primed OT-I T cells.

Another surprising finding — which is so different from previous work in different systems that I’m hesitant to believe it — is the timing of cell killing. Previous studies (such as the von Andrian paper2 that produced this video) have suggested that CTL kill their targets in something under an hour; maybe 30 minutes or even less. Here. Bousso’s group find that the tumor cells take something like 6 hours to be killed. That’s such a large difference — and has such important implications for effectiveness of CTL killing — that, as I say, I’d like to see it confirmed before I take it to the bank.

Bousso’s web site has a bunch of other equally fascinating videos; check them all out.

  1. This is probably related to the ability of tumors to suppress immune responses, which I’ve talked about before.[]
  2. Mempel, T. R., Pittet, M. J., Khazaie, K., Weninger, W., Weissleder, R., von Boehmer, H., and von Andrian, U. H. (2006). Regulatory T cells reversibly suppress cytotoxic T cell function independent of effector differentiation. Immunity 25, 129-141.[]