Recombinant Human Interleukin-11 protein (IL11) (Active)

What is the molecular structure of recombinant human IL-11?

Recombinant human IL-11 is a full-length protein spanning amino acids 22-199, with a molecular structure that facilitates binding to its cognate receptor IL-11RA before engaging with the shared gp130 (IL6ST) coreceptor to form a hexameric signaling complex . The protein sequence begins with PGPPPGPPR and contains multiple structural domains that contribute to its functional specificity and receptor binding properties . Proper folding of this 178-amino acid sequence is critical for biological activity, and most commercial preparations express the protein in HEK293 cells to ensure proper post-translational modifications and maintaining ≥95% purity with minimal endotoxin contamination .

How does IL-11 signaling differ from other IL-6 family cytokines?

IL-11 signaling shares the common gp130-mediated pathway with other IL-6 family cytokines but has distinct receptor expression patterns and signaling dynamics . Unlike IL-6, which primarily targets immune cells through IL-6R expression, IL-11 predominantly acts on stromal cells (fibroblasts, adipocytes, vascular smooth muscle cells) where IL-11RA is highly expressed . IL-11 activates JAK/STAT3 signaling that is more transient and less potent compared to IL-6 or Oncostatin M, while showing more prolonged and biphasic ERK activation . Additionally, IL-11 uniquely signals through the ERK/P90RSK pathway to inhibit LKB1, activate mTOR, and inhibit GSK3β, triggering a mesenchymal transition program not observed with all IL-6 family members .

What distinguishes "classic" from "trans" IL-11 signaling pathways?

Classic IL-11 signaling occurs when IL-11 binds to membrane-bound IL-11RA which then complexes with membrane-bound gp130, primarily affecting cells expressing the IL-11RA receptor . In contrast, trans-signaling involves the binding of IL-11 to soluble IL-11RA, creating a complex that can activate gp130 on cells lacking IL-11RA . This distinction is functionally significant as trans-signaling expands the range of potential target cells beyond those expressing IL-11RA, potentially mediating different biological effects and contributing to pathological processes . The dominant form of IL-11 activity under physiological versus pathological conditions requires further research, though current evidence suggests classic signaling predominates under normal conditions .

What are the critical factors for maintaining IL-11 activity in experimental settings?

Maintaining IL-11 activity requires careful attention to storage conditions, reconstitution protocols, and exposure to freeze-thaw cycles . For optimal results, recombinant IL-11 should be reconstituted in sterile, serum-free media containing carrier protein (0.1-1% BSA or HSA) and stored at -20°C to -80°C in single-use aliquots to avoid repeated freeze-thaw cycles . Working concentrations should be established through titration experiments, as effective doses range from 0.02-0.12 ng/mL for certain bioassays to 10-10,000 ng/mL in tissue-based experiments . Additionally, researchers should verify protein activity through functional assays specific to their experimental system, as activity can diminish over time even under optimal storage conditions.

How should researchers address species-specificity issues when working with human IL-11 in animal models?

Species-specificity presents a significant challenge when interpreting IL-11 studies, as recombinant human IL-11 (rhIL-11) has been shown to act as a partial antagonist of mouse Il11 signaling rather than a true agonist . Researchers should use species-matched recombinant proteins (e.g., mouse Il11 for mouse studies) to accurately assess gain-of-function effects . If using rhIL-11 in mouse models, scientists must recognize that observed effects likely represent inhibition of endogenous mouse Il11 function rather than human IL-11 gain-of-function . Alternative approaches include using transgenic animals expressing human IL-11RA or validating findings with neutralizing antibodies against endogenous Il11 to confirm mechanistic interpretations . This species-specificity issue explains many contradictory findings in the literature, particularly regarding anti-inflammatory versus pro-inflammatory effects.

What controls are essential for IL-11 signaling experiments?

Essential controls for IL-11 signaling experiments include vehicle controls, dose-response analyses, time-course studies, and pathway-specific inhibitors . Researchers should include positive controls such as IL-6 or Oncostatin M to benchmark signaling intensity and kinetics . JAK inhibitors (e.g., ruxolitinib), STAT3 inhibitors, and ERK pathway inhibitors should be employed to confirm pathway specificity . When studying inflammatory responses, parallel stimulation with known pro-inflammatory factors (TNF-α, IL-1β) provides contextual comparison . For studies involving receptor interactions, controls with receptor-blocking antibodies or soluble receptors are critical for distinguishing classic from trans-signaling effects . In animal models, genetic controls (receptor knockouts) and species-matched cytokines are vital to avoid misinterpretation of species-specific effects described in the literature .

How has understanding of IL-11's role in hematopoiesis evolved, and what implications does this have for research?

The understanding of IL-11's role in hematopoiesis has undergone significant revision . Initially characterized as a key hematopoietic factor, studies of IL-11 receptor knockouts (IL11RA-/- mice) and long-term neutralizing antibody administration revealed no significant effects on blood counts, contradicting the initial classification . The acute thrombocytosis observed after high-dose rhIL-11 administration likely represents pharmacological activation of gp130-related signaling in the bone marrow, similar to Oncostatin M, rather than reflecting IL-11's physiological function . Researchers should therefore approach IL-11 not primarily as a hematopoietic factor but instead focus on its roles in tissue injury responses, inflammation, and fibrosis . This paradigm shift necessitates reinterpretation of earlier literature and suggests that therapeutic applications in thrombocytopenia may represent off-target effects rather than physiological replacement.

What methodological approaches best capture IL-11's effects on epithelial-mesenchymal transition (EMT) and fibrosis?

Investigating IL-11's effects on epithelial-mesenchymal transition (EMT) and fibrosis requires multi-parametric approaches combining molecular, cellular, and tissue-level analyses . At the molecular level, researchers should measure changes in EMT markers (decreased E-cadherin, increased vimentin, fibronectin) alongside activation of downstream signaling pathways (JAK/STAT3, ERK/P90RSK, GSK3β/SNAI1) . Time-course experiments are essential, as IL-11 induces a program of mesenchymal transition that evolves over time rather than operating as an immediate switch . Three-dimensional cell culture models better recapitulate the complex cellular interactions versus traditional monolayer cultures, while organoid systems can reveal tissue-specific responses . In animal models, conditional cell-specific knockouts of IL-11 or IL-11RA provide more precise insights than global knockouts, which should be complemented with histological assessments of tissue architecture, collagen deposition, and inflammatory infiltration to fully characterize the fibrotic response .

How can researchers reconcile contradictory findings regarding IL-11's pro- versus anti-inflammatory effects?

Reconciling contradictory findings regarding IL-11's inflammatory effects requires careful attention to experimental contexts, species differences, and dosing regimens . The apparent anti-inflammatory effects observed in earlier mouse studies using rhIL-11 likely resulted from rhIL-11 acting as a partial antagonist of endogenous mouse Il11 signaling . In contrast, studies using species-matched Il11 or human cells with rhIL-11 reveal pro-inflammatory effects, including upregulation of factors such as SERPINB2, TNFRSF18, IL33, CCL20, IL1RL1, CXCL3/5/8, and ICAM1 . To resolve these contradictions, researchers should: (1) use species-matched recombinant proteins, (2) validate findings with receptor knockouts or neutralizing antibodies, (3) distinguish between acute versus chronic effects, and (4) consider the tissue-specific microenvironment . Additionally, examining the effects of IL-11 on specific inflammatory cell populations rather than just measuring global inflammatory markers provides mechanistic clarity regarding seemingly contradictory outcomes.

What considerations are important when using IL-11 for inflammatory bowel disease (IBD) research models?

When investigating IL-11 in inflammatory bowel disease (IBD) models, researchers must consider several critical factors that influence experimental outcomes and interpretation . First, distinguish between preventive versus therapeutic intervention timing, as rhIL-11 shows different efficacy depending on whether it's administered before inflammation onset or during active disease . Second, assess both clinical parameters (diarrhea, weight loss) and molecular/histological markers (mucosal healing, ion transport function, myeloperoxidase activity) to comprehensively evaluate treatment effects . Third, recognize that rhIL-11 modulates epithelial ion transport differently in inflamed versus healthy tissue, affecting outcomes in a context-dependent manner . Fourth, investigate the differential effects on jejunum versus colon, as these tissues respond distinctly to both inflammation and IL-11 intervention . Finally, consider potential discrepancies between acute pharmacological effects versus chronic physiological roles of IL-11 in intestinal homeostasis, which may explain conflicting results in the literature.

How should researchers approach IL-11 inhibition studies for fibrotic and inflammatory diseases?

Approaching IL-11 inhibition studies requires strategic consideration of inhibition mechanisms, dosing regimens, and appropriate disease models . Researchers can target IL-11 signaling through several approaches: neutralizing antibodies against IL-11 itself, receptor-blocking antibodies against IL-11RA, or small molecule inhibitors of downstream signaling components like JAK2 . Timing of intervention is critical—prophylactic administration addresses preventive potential while intervention during established disease tests therapeutic efficacy . Dose-finding studies should establish optimal inhibition while monitoring for potential compensatory upregulation of other IL-6 family cytokines . Appropriate models include those with documented IL-11 pathway activation, such as TGFβ-driven fibrosis models or certain inflammatory conditions . Readouts should comprehensively assess tissue architecture, function, inflammatory markers, and fibrosis-associated gene expression to evaluate efficacy . Finally, genetic validation using conditional knockout approaches helps confirm that observed effects result specifically from IL-11 inhibition rather than off-target effects.

What are the key considerations for IL-11 pharmacokinetic/pharmacodynamic (PK/PD) studies in research applications?

Pharmacokinetic/pharmacodynamic (PK/PD) studies of IL-11 require careful attention to several parameters to ensure reliable and translatable results . For PK assessment, researchers should employ sensitive and specific immunoassays capable of distinguishing endogenous from exogenous IL-11, with appropriate sampling timepoints based on the expected half-life (~7-8 hours in humans) . Standard measurements should include maximum concentration (Cmax), area under the curve (AUC), elimination half-life (t1/2), and volume of distribution . For PD assessment, both proximal biomarkers (STAT3 phosphorylation, ERK activation) and distal functional outcomes (platelet counts, inflammatory markers) should be monitored to establish exposure-response relationships . Route of administration significantly impacts PK/PD profiles, with subcutaneous delivery providing more sustained exposure compared to intravenous administration . Species differences in IL-11 receptor binding and signaling necessitate caution when extrapolating PK/PD data across species, particularly between rodents and humans . Finally, researchers should consider potential differences in PK/PD parameters between healthy and disease states, as inflammation, fibrosis, or organ dysfunction may significantly alter IL-11 disposition and efficacy.

What are the critical quality control parameters for ensuring consistent IL-11 activity in research applications?

Ensuring consistent IL-11 activity requires rigorous quality control encompassing physical, chemical, and biological parameters . Purity ≥95% confirmed by SDS-PAGE and HPLC is essential, with contaminants below detection limits . Endotoxin levels must remain below 0.005 EU/μg to prevent experimental artifacts . Biological activity should be verified through standardized assays measuring proliferation of IL-11-dependent cell lines or STAT3 phosphorylation, with batch-to-batch consistency in EC50 values (typically 0.02-0.12 ng/mL for proliferation assays) . Physical stability should be confirmed through appropriate protein concentration validation, absence of aggregation, and consistent secondary structure . Identity confirmation through mass spectrometry and N-terminal sequencing ensures the correct protein sequence with proper post-translational modifications . Finally, functional testing in the specific experimental system of interest provides the ultimate verification of activity relevant to the research question.

How can researchers distinguish between classic and trans-signaling effects in IL-11 experiments?

Distinguishing classic from trans-signaling in IL-11 experiments requires specialized experimental designs and reagents . To isolate classic signaling, researchers can use cells expressing membrane-bound IL-11RA while blocking soluble receptor with neutralizing antibodies specific to soluble IL-11RA . Conversely, to study trans-signaling, IL-11RA-negative cells can be treated with pre-formed complexes of IL-11 with soluble IL-11RA . Receptor expression profiling through flow cytometry, immunofluorescence, or Western blotting establishes the baseline IL-11RA status of experimental systems . Genetic approaches using IL-11RA knockout systems with controlled reintroduction of membrane-bound or soluble forms provide definitive mechanistic insights . Differential pathway activation patterns may help distinguish signaling modes, as trans-signaling often produces more sustained STAT3 activation . Finally, pathway-specific inhibitors targeting different components of the signaling cascade can reveal mechanistic differences between classic and trans-signaling responses .

What strategies can address the reproducibility challenges in IL-11 research given historical misunderstandings?

Addressing reproducibility challenges in IL-11 research requires acknowledging historical misunderstandings while implementing rigorous experimental controls and transparent reporting . Researchers should explicitly state the species origin of both the IL-11 protein and the experimental system, recognizing that rhIL-11 in mouse systems represents a partial antagonist rather than agonist . Comprehensive signaling pathway analysis beyond simplistic "activated/not activated" readouts provides mechanistic clarity, while time-course experiments capture the dynamic nature of IL-11 responses . Genetic validation through receptor knockouts or knockdowns confirms specificity of observed effects . When comparing to published literature, researchers should critically evaluate whether apparent contradictions stem from species mismatch issues rather than genuine biological differences . Multi-laboratory validation for key findings enhances confidence, while detailed methodology reporting (protein source, purity, activity verification, dosing, timing) enables proper replication . Finally, publishing negative or contradictory results alongside positive findings provides a more complete picture of IL-11 biology and prevents perpetuation of misunderstandings.