I'm curious about this model of primary cells and want to compare normal to diabetic sourced cells. There seems to be a lack of sources in the literature using these cells for glucose uptake experiments. I'd prefer not to spend ~$1000 to see if the uptake conditions (insulin concentration and time) I use for rat L6 cells will work the same with these cells.
Thanks.
Derek,
I understand your reluctance in not spending a considerable sum of money just find out that the cells do not behave as you hoped. [Several years ago I ran into the same situation with various hepatoma cells as well as primary human hepatocytes supplied by more than one commercial outfits. From my perspective, they all proved useless.]
Have you tried to contact Lonza to provide you detailed information about the strain as to what they have done (in-house) or other investigators have done with their strains.
Hi Derek,
The short answer to your question is yes, it is possible to use Lonza's human skeletal muscle myoblasts for 2-deoxyglucose uptake experiments
I am in the process of finishing up a study that does just that. We differentiated the HSMM and used the differentiated cells for our assay. We used only the healthy cell line and not the diabetic sourced line from Lonza. Compared to the L6s we found that the absolute glucose uptake was much lower in the HSMM compared to the L6s, but the overall relative increase in insulin stimulated glucose uptake was still significant. We also found that the HSMM required a larger dose of insulin to stimulate maximal glucose uptake compared to the L6s, though the time for incubation was the same. I also found that the glucose uptake assay was not possible with out coating the plates prior to plating them (I used Cultrex BME) as the differentiated cells would flake off without it. I strongly recommend optimizing the protocol for your laboratory conditions prior to your experiments.
One of the big considerations with these cells is the fact that they are not fully immortalized and consequently you will need to perform your experiments at around the same passage number, as the early passage cell and late passage cells behave very differently in culture. These cells are definitely more finicky to work with compare the L6s and C2C12 cells.
@Tausif_Alam Thank you for your reply. I have used Lonza in the past for another primary cell line and had success with them, but the growth conditions and the assay conditions were simpler. I've looked at the publication database on the Lonza website for these cells, and they don't have anything listed. I did manage to find one paper through a Google search that had used the cells. I'm still on the fence.
@Katie Thank you for your detailed reply. You have confirmed some of my concerns regarding these cells. I was curious about the coating and have considered using a coating for the L6 cells as well, as I have mixed results with the cells lifting during heavy washing protocols. I find the L6 and C2C12 cells very difficult to work with, and am sort of at a loss as to why I seem to have such trouble with them compared to the rest of the scientific community.
Having the primary human cells would add an extra level of importance and novelty to my current projects. I will take a look at the coating you suggest and go from there. The paper I mentioned in the reply to Tausif, and an old paper looking at human primary myoblasts from fresh isolation, also suggested that these cells require higher doses of insulin. These papers also used a longer time point of insulin treatment than I currently use.
Best of luck finishing your study.
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