Recombinant Human Interleukin-11 (IL11), partial (Active)

What is the molecular structure of IL-11 and how does it form a functional signaling complex?

IL-11 is a member of the IL-6 family of cytokines that binds to the cognate IL-11 receptor alpha subunit (IL11RA) to form a hexameric IL11:IL11RA:gp130 signaling complex. The structural assembly occurs in three key steps:

• Initial formation of a 1:1 complex between IL-11 and IL-11Rα mediated through site-I of the cytokine

• Interaction between the binary IL-11/IL-11Rα complex and the first molecule of gp130, comprising two coupled interfaces: IL-11 and gp130 (site-IIA) and IL-11Rα and gp130 (site-IIB)

• Recruitment of a second gp130 molecule to complete the hexameric complex

The interaction of IL-11 with IL-11Rα has an affinity of approximately 23 nM and is strongly entropy-driven. The trimeric complex formation (IL-11/IL-11Rα/gp130) has a moderate affinity with KD of approximately 380 ± 190 nM .

How does the IL-11 signaling pathway function in cells?

IL-11 signaling operates through both canonical and non-canonical pathways:

• Canonical pathway: Binding to the membrane-bound IL11RA and IL6ST (gp130) activates JAK/STAT signaling, particularly STAT3 phosphorylation

• Non-canonical pathway: Activation of ERK and other MAP kinases

When IL-11 binds to its receptor, it induces phosphorylation of the receptor signaling subunit gp130, which then recruits and activates JAK/TYK kinases. These kinases phosphorylate STAT proteins (primarily STAT3), which then translocate to the nucleus to regulate gene expression .

In addition to the classic membrane-bound receptor signaling, IL-11 can also participate in "trans-signaling" when IL-11 and soluble IL11RA bind to IL6ST on target cells .

What cell types are most appropriate for studying IL-11 signaling and why?

Primary cells are strongly recommended over immortalized cell lines for studying IL-11 signaling due to their preservation of physiological receptor expression and signaling pathways. The following primary cells are particularly suitable:

• Primary hepatic stellate cells (HSCs): Express high levels of IL11RA

• Primary fibroblasts: Particularly human lung fibroblasts (HLFs)

• Primary hepatocytes: Ideal for studying IL-11 effects on liver function

• Primary megakaryocytes: For thrombopoietic studies

The specific IL11RA receptor is highly expressed on cells of the stromal and parenchymal niche but expressed at low levels on immune cells, highly passaged cells, or transformed cell lines. This differential expression makes primary stromal and parenchymal cells ideal for IL-11 research .

Using immortalized cell lines can lead to misleading results due to altered receptor expression patterns and dysregulated signaling pathways .

What are the critical experimental conditions for obtaining reliable IL-11 signaling data in vitro?

Several critical factors affect IL-11 signaling studies and can lead to experimental artifacts if not properly controlled:

• Cell type and passage number: IL11RA expression decreases with passage number; use low-passage primary cells (P1-P3)

• Cell culture conditions: Serum components can activate or inhibit IL-11 signaling; standardize serum conditions

• Recombinant IL-11 concentration: Use physiologically relevant concentrations (10-50 ng/ml)

• Timing of measurements: IL-11 induces rapid signaling events (minutes to hours)

• Choice of readouts: Include both transcriptional (STAT3) and post-transcriptional (ERK/mTOR) endpoints

Researchers should carefully plan experiments with primary cell material. Otherwise, physiologically relevant mechanisms may become dysfunctional and reproducible experimental artifacts can obscure the true cytokine biology .

How can researchers effectively study the contradictory effects of IL-11 reported in the literature?

The contradictory effects of IL-11 reported in the literature often stem from:

• Species-specific differences: Human rhIL-11 acts as a partial/incomplete agonist in mouse models

• Cell type-specific responses: Different cell types express varying levels of IL11RA

• Experimental conditions: Variations in dose, timing, and readouts

Researchers should:

• Use species-matched recombinant IL-11 (e.g., mouse IL-11 for mouse studies)

• Compare effects of endogenous IL-11 vs. recombinant IL-11

• Include both gain-of-function and loss-of-function approaches (antibody neutralization)

• Use multiple readouts to capture diverse signaling effects

• Control for the potential blocking effect of human rhIL-11 on endogenous mouse IL-11

These approaches will help reconcile the seemingly contradictory data on IL-11 as either protective or detrimental in disease models .

Why does human rhIL-11 produce different effects in mouse models than in human systems?

The species-specific differences in IL-11 effects stem from receptor binding and signaling complex activation properties:

• Human rhIL-11 binds strongly to mouse Il11ra1 but incompletely activates the resulting (rhIL-11:Il11ra1:gp130) signaling complex

• This partial activation creates a competitive inhibition effect, where rhIL-11 acts as a partial/incomplete agonist that blocks endogenous mouse Il11 signaling

• Paradoxically, rhIL-11 can partially activate gp130 signaling in mouse cells lacking Il11ra1

This explains why rhIL-11 often shows protective effects in mouse models of disease while endogenous mouse Il11 shows detrimental effects. In multiple studies, rhIL-11 dose-dependently inhibited endogenous mouse Il11-dependent effects, creating opposing results between human and mouse systems .

What binding kinetics explain the species-specific differences in IL-11 activity?

The binding kinetics of recombinant IL-11 show important species differences that explain the variable effects observed in experimental systems:

These differences in binding kinetics result in different signaling outcomes when human IL-11 is used in mouse systems compared to human systems . Researchers should consider these species-specific binding differences when designing cross-species experiments.

How does IL-11 regulate cell metabolism and what signaling pathways are involved?

IL-11 regulates cell metabolism through a complex signaling cascade that involves:

• Activation of ERK/P90RSK

• Inhibition of LKB1/AMPK

• Activation of mTOR signaling

The sequential events occur as follows:

• IL-11 stimulates ERK and P90RSK phosphorylation

• Activated ERK and P90RSK phosphorylate LKB1 at specific sites

• Dual phosphorylation inhibits LKB1 activity

• Decreased LKB1 activity leads to reduced AMPK phosphorylation

• Reduced AMPK activity results in increased mTOR activation

This pathway has been observed in multiple cell types including fibroblasts, epithelial cells, and hepatocytes. Inhibition of either ERK (with U0126) or P90RSK (with BI-D1870) prevents the downstream effects of IL-11 on LKB1/AMPK/mTOR signaling .

What are the contrasting effects of IL-11 on different cell types?

IL-11 exerts distinct and sometimes opposing effects on different cell types:

Megakaryocytes and hematopoietic cells:

• Promotes proliferation and maturation

• Increases platelet production

• Directly acts on megakaryocytes through IL-11 receptor expression on CD41+ cells

• Activates STAT3 and gp130 signaling in these cells

Fibroblasts and stromal cells:

• Activates fibroblasts and promotes fibrogenesis

• Increases collagen production

• Stimulates ERK/mTOR signaling leading to fibrotic phenotypes

Epithelial cells:

• Can cause epithelial cell dysfunction

• Promotes epithelial-mesenchymal transition (EMT)

• Impacts ion transport across epithelial barriers

• Modifies electrogenic ion transport in intestinal mucosa

Endothelial cells:

• Stimulates directed migration and tubule formation

• Promotes angiogenesis and collateral vessel growth

• Increases CD34+/VEGFR2+ cell mobilization

These cell-type specific effects highlight the importance of selecting appropriate experimental systems when studying IL-11 biology.

What pharmacokinetic principles should guide rhIL-11 dosing in research protocols?

Pharmacokinetic studies of rhIL-11 provide important guidance for dosing in research protocols:

• Linear pharmacokinetics: rhIL-11 shows linear pharmacokinetics after both intravenous infusion and subcutaneous administration

• Absorption-limited kinetics: Comparison of t1/2 and MRT values after intravenous vs. subcutaneous administration indicates that rhIL-11 pharmacokinetics after subcutaneous administration are absorption rate-limited

• Bioavailability: Subcutaneous administration provides approximately 65% bioavailability

• Elimination: rhIL-11 is primarily eliminated by metabolism (not detected in urine)

• Stable kinetics with repeated dosing: No significant change in pharmacokinetic profile occurs with repeated subcutaneous administration

Based on these principles, research protocols typically use subcutaneous dosing of 3-50 μg/kg for single-dose studies and 3-15 μg/kg for repeated dosing studies .

How can researchers explain the mixed results from rhIL-11 clinical trials in different disease settings?

The mixed results from rhIL-11 clinical trials can be explained by several factors:

• Species differences in signaling: As noted earlier, human rhIL-11 acts as a partial agonist in murine models, potentially creating misleading preclinical data

• Dose-dependent effects: Different doses can activate distinct signaling pathways with opposing effects

• Disease context specificity: IL-11 effects vary dramatically depending on the underlying pathophysiology

For example, while rhIL-11 showed promise in mouse models of various diseases (myocardial infarction, colitis, liver disease, etc.), the human clinical trials showed mixed results with none progressing to pivotal phase 3 studies. This suggests that the positive results in mouse models may have been due to rhIL-11's ability to block endogenous mouse Il11, rather than a true therapeutic effect .

In Crohn's disease trials, rhIL-11 at 15 μg/kg once weekly induced remission in a significantly greater proportion of patients compared to placebo (36.7% vs. 16.3%), while a 7.5 μg/kg twice weekly regimen was not significantly better than placebo. This highlights the importance of optimal dosing strategies in clinical applications .

What methodological approaches can reliably assess IL-11 activity in experimental and translational research?

To reliably assess IL-11 activity in research, multiple complementary approaches should be employed:

• Receptor signaling assessment:

  • Phosphorylation of gp130, JAK/STAT3 (Western blot)

  • ERK/P90RSK phosphorylation (Western blot)

  • LKB1/AMPK/mTOR pathway components (Western blot)

• Functional readouts:

  • For megakaryocytes: ploidy analysis, platelet count

  • For fibroblasts: collagen production, myofibroblast marker expression

  • For epithelial cells: ion transport measurements, barrier function

  • For endothelial cells: tubule formation, migration assays

• In vivo assessments:

  • Mobilization of CD34+/VEGFR2+ cells

  • Tissue-specific functional outcomes (e.g., perfusion in ischemia models)

  • Histological assessment of target tissues

• Species considerations:

  • Use species-matched recombinant IL-11

  • Include both gain-of-function (adding recombinant IL-11) and loss-of-function (IL-11 or IL-11RA neutralizing antibodies) approaches

These methodological approaches provide a comprehensive assessment of IL-11 activity across multiple biological systems .

How can researchers better understand the dual role of IL-11 in inflammatory processes?

To investigate the seemingly contradictory pro-inflammatory and anti-inflammatory effects of IL-11, researchers should:

• Use species-appropriate reagents: Species-matched recombinant IL-11 proteins to avoid the partial agonist effect

• Compare gain- and loss-of-function: Both addition of recombinant IL-11 and neutralization of endogenous IL-11

• Assess temporal aspects: Examine both acute and chronic effects of IL-11 intervention

• Analyze cell-specific responses: Isolate and study responses in specific cell populations

• Examine pathway specificity: Determine which inflammatory pathways are activated or suppressed

Research has shown that while rhIL-11 appears anti-inflammatory in mouse models of inflammatory bowel disease, this may be partly due to blocking effects on endogenous mouse IL-11. In human tissue, the effect of IL-11 on intestinal epithelial electrogenic ion transport appears to be concentration-dependent .

What approaches can differentiate between the canonical and non-canonical signaling effects of IL-11?

To differentiate between canonical (JAK/STAT) and non-canonical (ERK/mTOR) signaling effects of IL-11, researchers should:

• Use pathway-specific inhibitors:

  • JAK inhibitors (e.g., ruxolitinib) for canonical pathway

  • MEK/ERK inhibitors (e.g., U0126) for non-canonical pathway

  • mTOR inhibitors (e.g., rapamycin) for downstream effects

• Employ genetic approaches:

  • STAT3 conditional knockout or knockdown

  • ERK1/2 conditional knockout or knockdown

  • Expression of dominant-negative pathway components

• Temporal analysis:

  • Canonical STAT3 activation occurs rapidly (minutes)

  • Non-canonical ERK/mTOR effects may have different kinetics

• Cell-type specific analysis:

  • Different cell types may preferentially activate different pathways

• Readout selection:

  • Measure both transcriptional (STAT-dependent) and post-transcriptional outcomes

Experiments with pharmacologic inhibition of either ERK (U0126) or P90RSK (BI-D1870) in cells stimulated with IL-11 have demonstrated the importance of the non-canonical pathway in mediating effects on metabolism through LKB1/AMPK/mTOR signaling .