Dear Theodore,

Prednisolone works well. It diminishes many inflammatory cytokines including TNF-a and IL-1b without toxicicity or apoptosis.

Dear Theodore,

Prednisolone works well. It diminishes many inflammatory cytokines including TNF-a and IL-1b without toxicicity or apoptosis.

Prednisolone.

I would suggest an inhibitor of NF-kB or p3MAPK; very reliable and consistent. Examples would be parthenolide, TPCA-1, SB239063 (low micromolar range). Relatively inexpensive and easy to solubilise in DMSO. More specific than glucocorticoids.

I agree withj Keelan, v resukts in HK cells have been obtained with like parthenolide, TPCA-1 and SB239063. Howeevre the changes are very discrete and not very impressive to editors. We have used moclonal antibodies for specific cytokines and give very good results.

Hi all,

have you tested these chemicals on cell lines? And what would you think of more "traditional" drugs such as Dex?

I have used all the ones mentioned above and have found that it does depend on which cells you are trying to influence. For instance Dex works very well for T cells but does not effect antibody production from b cells. As a general block dex is best in my lab at least.

Dr. Kapellos,

Depending upon cell culture type, I guess I would try a couple of those anti-inflammatory drugs/agents listed by others in your system without stimulating to see what effects these have on cells. Dexamethasone/steroid may upregulate some anti-inflammatory markers on macrophage surface/modify differentiation state. The 38, NF-KB inhibitors may not as these act downstream from steroid? Also, how do you challenge cell lines to induce inflammation? What endpoints do you measure to evaluate effects of marker? Prostanoid production works well, is easy and inexpensive and for macrophages, there are no further metabolism of PGE2 as there might be in serum/urine. Apoptosis can easily be monitored as can proliferation (depends upon cell type).

I have the BV-2 microglia cell line and I want to reduce inflammation after LPS/IFNg treatment. I will measure NO levels, TNFa, IL1b, IL6, MIP-1a and MCP-1 expression to validate my system.

It will depend on you readout, but why not trying IL-10

Prednisone or dexamethasone. Good luck!

I would try celecoxib if you want COX2 inhibition, we tried it on tumor cells in vitro and also in vivo. If tou just want to set up a test for CX inhibitors you may try with Indomethacin it works well, it inhibits PGE2 and TNFa and other inflammatory cq as well.

I used dexamethasone, corticosterone and prednisone in the past to successfully block inflammatory cells, mainly lymphoid cells, in the rat paw edema and intradermal vascular permeability experimental models. In fact, the inflammatory reaction inhibited in leucopenic rats was restored by i.v. injection of lymphocytes collected from normal rat blood/thoracic lymphatic duct, but not from rats pretreated with such steroidal or several NSAIDs. Lymphocytes previously incubated with the same drugs and administered i.v. in leucopenic rats were also not able to revert the inhibited inflammatory reaction. A Lymphocyte pro-Inflammatory Factor (LpIF) was characterized and its role in non-immune acute inflammation was demonstrated. For more details see: Garcia Leme J, Bechara GH, Sudo LS, 1977. The pro-inflammatory function of lymphocytes in non-immune inflammation: Effect of steroidal and non-steroidal anti-inflammatory agents. British Journal of Experimental Pathology, 58:703-711.

If your intention is to test drugs that have also an immunosuppressive effect Dex and Betamethasone are fine. However, you should bear in mind that they act by interfering with multiple mechanisms. If you are testting NSAID-like drugs I do suggest that you use as your control ibuprofen that works well in vitro.

Good Luck

Giuseppe Cirino

I have always considered DEX to the be gold standard as an overall anti-inflammatory agent. However, you need to address what inflammatory pathways your current drug is going to attenuate and then put them head to head. Compare apples to apples IMO.

how about Celastrol? it's a mighty proteasome inhibitor works at less than 1µM!

it became our most efficient anti-TNF for our HCS screening since our paper in BBRC

Pinna, G. F., Fiorucci, M., Reimund, J. M., Taquet, N., Arondel, Y. & Muller, C. D. (2004). Celastrol inhibits pro-inflammatory cytokine secretion in Crohn's disease biopsies. Biochemical and Biophysical Research Communications, Vol.322 N°3 (Sep 24), pp. 778-786

Moreira-Tabaka, H., Peluso, J., Vonesch, J. L., Hentsch, D., Kessler, P., Reimund, J. M., Dumont, S. & Muller, C. D. (2012). Unlike for human monocytes after LPS activation, release of TNF-alpha by THP-1 cells is produced by a TACE catalytically different from constitutive TACE. PLoS One, Vol.7 N°3 pp. e34184

Prednisolone and Dexamethasone can woork well.

Inflammation simply means the presence of white cells. Macrophages and other leukocytes can make markedly different types of "inflammatory" reactions depending on the circumstance (stimuli, etc.). For example, macs can make NO (killer molecule) or Ornithine (repair molecule) - both derived from arginine. So, it's not just about "inflammation" - its about the type. In this connection, my experience is that cortisol and related steriods kill leukocytes in vitro. So, careful if your inflammatory read out (whatever that is) goes down with your drug, it could just be cell death. This is much different than the ability of a drug to stop leukocytes from entering from the blood - an anti inflammatory property. Good luck

Perhaps this paper helps.

How about this? It's an oldie but a goodie; indomethacin. It has anti-inflammatory properties. By the way, you will need to determine whether your anti-inflammatory drugs or compounds work in a culture system or whether they exist as "pro-drugs" and need to be metabolised into active compounds. I was thinking also about cyclophosphamide. It is said to exist as a pro-drug and needs to be metabolised in the liver. However, there are papers out there indicating that it could exert its effects in vitro.

My main concern about prednisolone and dexametasone are their side-effects and genomic effects, thus we can conclude that they are not specific anti-inflammatory drugs.

We always use dexamethasone as the positive control to inhibit pro-inflammatory mediators in our lab. However, dexamethosone is a type of glucocorticoids which basically inhibits every single type of pro-inflammatory mediators. If you want something more specific, you may consider other specific inhibitors for example NF-kB or p38 MAPK inhibitor.

Theodore - what cells are you going to culture, macropphage, microglai, mixed glial? In macrophage or microglial cultures o switch off production of classic inflammatory mediators such as IL1B, COX2 and IL6 and increase anti-inflammatory markers (Arg1, IGF1 etc) treat with IL4 or IL10. This is known as inducing a M2 polarisation (see attache refs). Glucocorticoids will do what you want, but in terms of a strong positive control I would suggest IL-4 at dosages of 20ng/ml (for eg, Butovsky et al 2006; Mol Cell Neuro).

It depends on what inflammatory signaling pathway you are interested in. If NFkB, you may try some IKKbeta inhibitors for your screening. If STAT3, then JAK inhibitors...

Try CAPE to inhibit NF-kappa B. CAPE inhibits LPS-induced inflammation in macropahges. Here is the ref.

Caffeic acid phenethyl ester is a potent and specific inhibitor of ...www.pnas.org/content/93/17/9090.abstract

CAPE has been approved to be anti-HIV drug to inhibit LTR-signal and replication.

Would minocycline be useful for this?

Is there any way to specifically inhibit phagocytosis/endocytosis by macrophages (particularly microglia)?

I am not sure about Minocycline.

Pierre Bush - Why do you have doubts about Mino ?? I have been looking at the literature and cant find anything solid that shows Mino decreases activity in pure MG cultures. Have you got any experience with it? It is certainly anti-inflammatory in the whole brain, but with pure MG i wonder if it is has such strong effects, but maybe it is the culture conditions (ie., too strong inflammatory stimuli).

Hi everybody.

Would you suggest any specific prednisone dosis to treat a T cell line culture? I mean, a prednisone concentration.

Thanks in advance

Hi everybody.

I want to make inflammation cell model. I am thinking use mononuclear cells. how can I create inflamation cell model.