I appriciate any answer.

Hi shokoufeh ,of course no. Cytokines are pleiotropic ... some Studies designed to determine the role of TNF in the animal models or cell culture system using pure recombinant molecules have revealed that TNF never operates by itself, but instead operates within a network of cytokines. Each cytokine has a matching cell -surface receptor. Upon binding, intracellular signaling and gene expression regulations are altered, leading to the production of other cytokines, surface receptors, or feedback inhibition.

But for a better and definitive answer, you can look at the path-way studio site.

I think it will be useful to you

The answer is most likely no. (Although you can differentiate the sigaling by looking at specific intracellular signaling proteins for each cytokine.)

Every cytokine does not have it's own signaling pathway. The pathways are usually grouped by receptor family.

And while the direct intracellular signaling events may be somewhat separated... depending on how close to the receptor you are looking at... There are a lot of major signaling molecules that cross receptor families. And a lot of stuff ends up activating NFkb.

Dear, most cytokines are pleiotropic in nature and work in synergistic manner to perform their functions.

Thank you so much Behnoush Nasr Zanjani, Shen-An Hwang, Emmanuel Ifeanyi Obeagu.

I have another question. I work with a co/culture model. I stimulated the cells with IL6, I checked the expression of TNF-a , INF-a, and IL-8, none of them has been upregulated . When I look at IL-6 signaling path is differed from TNF-a. I am thinking that's why didn't see the upregulation of TNF-a. I appreciate any answer from you.

Totally agree with fantastic answer of Zanjani.

Frida Gorreja hi dear , Pathway Studio İs an exhaustive resource of easily searchable data from biology articles describing interactions between molecules, cell processes, and diseases. It also helps biological researchers import and analyze their experimental data using statistical tools.

Firstly you can creatr an account then choose pathway mammalplus or plant . you can get valuable information

Pls fallow this link : https://www.pathwaystudio.com/

What are the cells in the co-culture?

Thank you very much for all answers. Dear Shen-An Hwang, the cell line are Caco2 and Hep G2.

Thanks a lots for your valuble knowledge that yoj are sharing with me . Still did not get a direct answer to my question. Can added IL6 to cell culture medium .upregulate TNF-a in cell culture model.

So... if you were using a macrophage/monocyte cell lines, I would have said yes, adding IL-6 will stimulate measureable TNFa production. Although this would also depend on the time point.

With these cell lines... I'm not too sure. It might not.

Dear Karimi,

Did you add soluble IL-6 receptor (sIL-6R) with IL-6 in CACO2 culture medium? If you have not done yet, I would like to recommend. IL-6 needs IL-6R expressed on the surface of cells or sIL-6R. In addition, you should also check the expression of IL-6R of CACO2.

All the best,

Kunihiko

Thank you very much Professor Kou Hayakawa for providing valuable knowledge.

Thanks Dr. Shen-An Hwang, do you have any suggestion why IL6 can stimulate TNF-a in macrophage and monocyte cell lines but not other cell lines?

Thank you Dr.Kunihiko Umekita.

No I have not added. But this cell lines express IL6, they might already have possibility of expression of the IL6R.What do you suggest?

Dr.Shen-An Hwang can you put the link for articles showing the upregulation of TNF-a in monocyte and macrophage cell lines after IL6 stimulation please?

Dear Karimi,

I would like to recommend to investigate the expression of IL-6 receptor or gp130 on your cell lines when you could not get any responses induced by IL-6.

Best wishes,

Kunihiko

Article Interleukin-6 trans-signaling increases the expression of ca...

In this article the presence of IL6R has been already identified.

I know that IL-6 is a pleiotropic cytokine , I dont know if it is what I see in my experiment, that cant upregulatet TNF-a, IL-8 and INF-a?

Dear Kunihiko Umekita, I hope I give you an proper snwer to your comment.

I take that back... I guess I must've remembered wrong. IL-6 can enhance TNF-a in the presence of something else (mostly LPS). That would explain why I kept thinking that macrophages might be more able to be stimulated with IL-6.

The only link I could find where using only IL-6 can stimulate TNFa is in vivo.

Article A Novel Small-Molecule Inhibitor Targeting the IL-6 Receptor...

Here's one in vitro... but apparently IL-6 may be more antagonistic to TNF production by itself without a primary stimulant.

https://ars.els-cdn.com/content/image/1-s2.0-S0925443917300108-gr4.jpg

Article IL-6 promotes M2 macrophage polarization by modulating purin...

Thanks Dr. Shen-An Hwang .

NO. IL-6 can induce TNF-a expression and vice versa. Even more other proinflammatory cytokines (e.g. IL-1) van also induce TNF-a and/or IL-6 expression nad chemokines were abloe to induce proinflammatory cytokine expression, too...and so on!

Dear Andreas Eisenreich Thank you. Can you send a link to a paper suggesting the fact above :IL6 can induce TNF-a.

e.g. involving RNA interaction :

Zhuang YT, Xu DY, Wang GY, Sun JL, Huang Y, Wang SZ.

Eur Rev Med Pharmacol Sci. 2017 Jan;21(2):302-309.

Other examples 4 an increasing activity:

-> Caiello I, Minnone G, Holzinger D, Vogl T, Prencipe G, Manzo A, De Benedetti F, Strippoli R. PLoS One. 2014 Oct 1;9(10):e107886. doi: 10.1371/journal.pone.0107886. eCollection 2014.

-> Ogawa H, Mukai K, Kawano Y, Minegishi Y, Karasuyama H.

Biochem Biophys Res Commun. 2012 Mar 30;420(1):114-8. doi: 10.1016/j.bbrc.2012.02.124. Epub 2012 Mar 3.

And:

Modulation of TNF-alpha mRNA production in rat C6 glioma cells by TNF-alpha, IL-1beta, IL-6, and IFN-alpha: in vitro analysis of cytokine-cytokine interactions.

Gayle D, Ilyin SE, Miele ME, Plata-Salamán CR.

Brain Res Bull. 1998 Oct;47(3):231-5.

Thank you so much Anders,

Really nice to see the involvement of IL-6 in inducing TNF-a.

I would like to give you more information that we have used IL-6 to stimulate the epithelial and liver cells and then we checked the TNF-a , IL-8 and INF-a. we used 10 ng /ml of IL-6 as final dose. but none of these cytokines has been upregulated. what do you suggest?

As you can see the impact of IL-6 on TNF-alpha depends on the experimental setting. Maybe, in your setting there is no impact at the concentration of 10 ng/mL or lower. Another reason may be the quality of the stimulus. HAve you checke activity of IL-6 used in your experiments as well as specificity of your analytical tools ans setting (qPCR, assays etc.)?

Moreover, did you checked successful transition of EpC to liver cells as well as expression of corresponding receptors, needed for IL-6-sependent signaling, and downstream effects (e.g. transcitional modification of potentail targets)?

Thank you so much Andreas Eisenreich for sharing your valuable knowledge with me. I have seen in many signalling pathways for cytokines that they have thier own pathways, and very few share partially small common path e.g. IL-10 and IL6....stat3 .... I am puzzeled if with seperate paths for TNF´-a and IL6 how and where in signalling pathway they effect each other. I know in cytokines network they may do that. Any idea?

This strongly depends on your experimental setting (cell types used, receptors and signaling pathways available etc.). One potential signaling pathway linking TNF-alpha and IL-6 expression is NFkappaB activation (e.g. see Clin Immunol. 2012 May; 143(2): 188–199 ). However, for further insights in this issue you have to check literature and advace your experimental setting.

Best wishes

Andreas

Dear Andreas, thank you so much. One final question as in my experiment I see upregulation of IL10 in cells treated with our samples. How can I mechanistically study this upregulation ?

I gratefully thank your reply in advance, Andreas Eisenreich.

In some disease settings (includig inflammation) IL-6 and IL-10 expression are linked (in the same direction). I hope this helps you finding adequate information from existing literature (e.g. see: Cell Mol Immunol. 2019 Mar;16(3):275-287. doi: 10.1038/cmi.2018.5. Epub 2018 Mar 19. ).

Best wishes Shokoufeh

Thank you so much Andreas.

I am really thankful :)

You`re welcome Shokoufeh.

Hi Shokoufeh,

IL-6 can trigger a signaling cascade involving different kinases and transcription factors, some of which are regulators of TNF-Alpha expression as well. In particular, IL-6 binding on its receptor transduce a JAK-dependent signaling leading to STAT3 activation (this is one of the more studied pathway but really there can be more downstream effectors). STAT3 is involved in several cellular processes, including inflammation, oncogenesis and cell proliferation. Other cytokines are able to induce a JAK-STAT signaling, similarly to IL-6. In fact cytokines commonly present redundant properties.

Thank you so much dear Valentino Bezzerri.

I believe that the body is so complicated and we are not able to understand exact mechanism. There are a number of active pathways and it is depending on different factors such as species, tissue, model (in vitro or in vivo), time of exposure and dosage of treatment. Don’t forget that gene expression is like taking a picture from tissue in a particular time, and cell is so intelligent, therefore it can be changed based on conditions. On the other hand, most of genes especially transcription factors are multifunction and have a small effect on phenotype, so it is difficult to say how and where they affect each other. Additionally, we should consider other factors such as non-coding RNAs. May be we have some up-regulated genes but we are not able to detect or measure their mRNA level because of microRNAs effect.

Dear Mojtaba Najafi thank you so much, I totally agree with you.

Some effects caused by IL-6 are similar to those observed under the action of IL-1 and TNF. However, the main effect of IL-6 is associated with its participation as a cofactor in the differentiation of B-lymphocytes, their maturation and transformation into plasma cells secreting immunoglobulins. In addition, IL-6 promotes the expression of the IL-2 receptor on activated immunocytes, and also induces the production of IL-2 by T-cells.