IGFBP6 (28-240) Human

What is IGFBP6 and what are its primary functions?

IGFBP6 is one of the six members of the insulin-like growth factor binding protein family. It has both IGF-dependent and IGF-independent cellular functions. IGFBP6 is involved in numerous cellular activities and represents an important factor in immune responses, particularly in human dendritic cells (DCs). Its functions include induction of chemotaxis, increasing oxidative burst and neutrophil degranulation, altering metabolic profiles of dendritic cells, and regulating the Sonic Hedgehog (SHH) signaling pathway during fibrosis development . Additionally, IGFBP6 acts as an acute-phase protein, being rapidly produced in response to damage, as demonstrated by experiments showing its production after DCs and fibroblast exposure to H₂O₂ .

How does IGFBP6 (28-240) differ from full-length IGFBP6?

The IGFBP6 (28-240) fragment represents the mature, processed form of human IGFBP6 after removal of the signal peptide. This region contains the core functional domains responsible for IGF binding and interactions with other signaling molecules. Research indicates that this processed form retains the immunomodulatory properties of IGFBP6, including its ability to induce chemotaxis and affect neutrophil functions . When designing experiments, researchers should consider that this specific fragment may show differential activity compared to the full-length protein in certain cellular contexts.

What expression patterns does IGFBP6 show in normal versus pathological conditions?

• In cystic fibrosis: IGFBP6 expression is higher in F508del-CFTR bronchial epithelial cells compared to wild-type cells. Furthermore, inflammatory stimuli like LPS or IL-1β+TNFα increase IGFBP6 expression in these cells .

• In ovarian cancer: IGFBP6 is significantly downregulated in primary and metastatic ovarian cancer tissues compared to normal ovarian tissues .

• In primary myelofibrosis (PMF): IGFBP6 shows increased expression in PMF patients wild type for JAK2V617F mutation compared to healthy subjects and PMF patients with the JAK2V617F mutation .

These differential expression patterns suggest context-specific regulation of IGFBP6 that may contribute to disease pathophysiology.

What are the recommended methods for measuring IGFBP6 expression in experimental systems?

Multiple complementary approaches are recommended for comprehensive IGFBP6 expression analysis:

• mRNA quantification: qRT-PCR using IGFBP6-specific primers normalized to housekeeping genes like GAPDH is effective for measuring transcript levels .

• Protein detection:

  • Western blotting using anti-IGFBP6 antibodies with appropriate loading controls (e.g., calnexin)

  • ELISA for quantification of secreted IGFBP6 in cell culture supernatants or biological fluids

• Cell-specific expression: For tissue samples, immunohistochemistry can determine cell-specific expression patterns and localization.

When publishing results, researchers should report both mRNA and protein data to account for potential post-transcriptional regulation.

How should researchers design in vitro experiments to study IGFBP6 functions?

Based on published methodologies, a comprehensive experimental approach should include:

• Dosage considerations: Use physiologically relevant concentrations. Studies have employed IGFBP6 at ranges from 0.2-200 ng/ml for dose-response experiments .

• Timing: Both acute (4-24 hours) and chronic (multiple days) exposure should be evaluated as different effects may manifest at different time points.

• Functional readouts:

  • Inflammatory responses: measure pro-inflammatory cytokine expression (TNFα, IL-6, IL-1β)

  • Migration/chemotaxis: transwell assays for immune cells

  • Oxidative burst: ROS production measurement

  • Cell-specific functional assays depending on the cell type under investigation

• Controls:

  • Include neutralizing antibodies against IGFBP6 to confirm specificity of observed effects

  • Compare recombinant IGFBP6 effects with conditioned media containing naturally secreted IGFBP6

What cell and tissue models are most appropriate for IGFBP6 functional studies?

Based on the current literature, several experimental models have proven valuable:

• Cell lines:

  • Bronchial epithelial cells (CFBE41o-) for respiratory studies

  • Matched cancer cell lines like PEA1/PEA2 (pre/post platinum resistance) for cancer research

  • Dendritic cells for immunological investigations

• Primary cells:

  • Human nasal epithelial (HNE) cultures from patients

  • Neutrophils for studying oxidative burst and degranulation

  • Bone marrow-derived mesenchymal stem cells

• Experimental manipulations:

  • Stress conditions (hyperthermia, oxidative stress with H₂O₂, hypoxia) to study IGFBP6 regulation

  • Inflammatory stimuli (LPS, IL-1β+TNFα) to mimic disease conditions

The choice of model should be guided by the specific aspect of IGFBP6 biology under investigation.

How does IGFBP6 influence immune cell function?

IGFBP6 exerts multiple effects on immune cells:

• Chemotactic properties: IGFBP6 induces chemotaxis of monocytes and T-lymphocytes, promoting their migration through epithelial monolayers .

• Neutrophil activation: IGFBP6 increases oxidative burst with reactive oxygen species (ROS) production and triggers degranulation of primary granules in neutrophils .

• Dendritic cell modulation: IGFBP6 induces metabolic changes in dendritic cells and is upregulated in response to hyperthermia, which impacts the immunostimulatory capacity of DCs .

• Role in inflammatory conditions: IGFBP6 is overexpressed in the serum and joints of rheumatoid arthritis patients , suggesting a role in chronic inflammation.

These diverse effects indicate IGFBP6 functions as an immunomodulatory molecule with both pro-inflammatory and potentially anti-inflammatory actions depending on the cellular context.

What is the relationship between IGFBP6 and inflammatory signaling pathways?

IGFBP6 interacts with several key inflammatory signaling pathways:

• TNFα signaling via NF-κB: Gene expression analysis reveals that IGFBP6 can modulate this pathway, though the direction of regulation appears context-dependent. In ovarian cancer cells, this pathway is differentially regulated in platinum-sensitive versus platinum-resistant cells exposed to IGFBP6 .

• IL-6 signaling: IGFBP6 impacts IL-6 signaling, which may contribute to its effects in inflammatory conditions .

• TLR4 signaling: Research suggests a relationship between IGFBP6, sonic hedgehog, and TLR4 signaling, particularly in conditions like primary myelofibrosis .

• Anti-inflammatory properties: Interestingly, IGFBP6 can reduce pro-inflammatory cytokine expression in a dose-dependent manner in certain contexts, such as cystic fibrosis bronchial epithelial cells challenged with LPS .

The seemingly contradictory effects of IGFBP6 on inflammation highlight its complex role as a modulator rather than a simple activator or inhibitor of inflammatory responses.

Does IGFBP6 have dual pro-inflammatory and anti-inflammatory functions?

Evidence supports a dual role for IGFBP6 in inflammation:

Pro-inflammatory functions:

• Induces chemotaxis of immune cells

• Increases neutrophil oxidative burst and degranulation

• Is elevated in inflammatory conditions like rheumatoid arthritis

Anti-inflammatory functions:

• Reduces pro-inflammatory cytokine expression in bronchial epithelial cells challenged with LPS

• Addition of neutralizing antibodies against IGFBP6 increases pro-inflammatory cytokine expression under LPS challenge

• May improve mitochondrial fitness and reduce ROS production in certain cell types

This dual nature suggests that IGFBP6 functions as a context-dependent immunomodulator. The specific outcome likely depends on the cellular environment, disease state, and presence of other inflammatory mediators. Researchers should design experiments to capture this complexity rather than categorizing IGFBP6 as strictly pro- or anti-inflammatory.

How is IGFBP6 involved in cancer biology?

IGFBP6 has a complex and sometimes contradictory role in cancer:

These pathways are activated in platinum-sensitive cells but inhibited in platinum-resistant cells following IGFBP6 exposure .

• Drug resistance associations: The IGFs/IGFBPs axis appears involved in modulating drug sensitivity in cancer cells, with IGFBP6 specifically implicated in the proliferation of chemoresistant tumor cells in human glioblastoma .

These findings suggest that IGFBP6 may serve as a potential biomarker or therapeutic target in cancer, particularly in the context of treatment resistance.

What role does IGFBP6 play in fibrosis development?

IGFBP6 is increasingly recognized as a significant factor in fibrosis development:

• Regulation of the Sonic Hedgehog (SHH) signaling pathway: IGFBP6 has been shown to regulate the SHH pathway during fibrosis development . This pathway is critical for tissue repair and can contribute to pathological fibrosis when dysregulated.

• Connection to immunity and stroma: There are well-established relationships between immunity, stromal activity, and fibrosis that impact cancer immunotherapy outcomes. IGFBP6 appears to be at the nexus of these interactions .

• Response to tissue damage: As an acute-phase protein rapidly produced in response to damage (e.g., after exposure to H₂O₂), IGFBP6 likely plays a role in the initial phases of the wound healing response that can lead to fibrosis under pathological conditions .

• TLR4 signaling axis: The IGFBP6/sonic hedgehog/TLR4 signaling axis has been implicated in bone marrow microenvironment alterations, which can contribute to fibrotic conditions .

Understanding the specific mechanisms by which IGFBP6 influences fibrosis could lead to novel therapeutic approaches for fibrotic diseases.

How does IGFBP6 contribute to cystic fibrosis pathophysiology?

Recent research has elucidated several aspects of IGFBP6's role in cystic fibrosis (CF):

• Altered expression: IGFBP6 mRNA and protein levels are upregulated in F508del-CFTR bronchial epithelial cells compared to wild-type CFTR cells at basal conditions .

• Response to inflammatory stimuli: LPS and IL-1β+TNFα treatments further increase IGFBP6 mRNA levels in CF bronchial epithelial cells .

• Anti-inflammatory potential: Exogenous IGFBP6 reduces the level of pro-inflammatory cytokines in both CF bronchial epithelial cells and primary nasal epithelial cells, without affecting rescued CFTR expression and function .

• Neutralization effects: Adding a neutralizing antibody against IGFBP6 increases pro-inflammatory cytokine expression under LPS challenge, suggesting an endogenous anti-inflammatory role for IGFBP6 in CF airways .

• Response to anti-inflammatory treatment: Dimethyl fumarate (DMF), an anti-inflammatory agent, reduces IGFBP6 expression in CF cells challenged with inflammatory stimuli .

These findings suggest IGFBP6 may act as part of a compensatory mechanism attempting to counterbalance excessive inflammation in CF, making it a potential therapeutic target for addressing the inflammatory component of CF pathology.

What are the IGF-independent functions of IGFBP6?

Research has revealed multiple IGF-independent functions of IGFBP6:

• Immunomodulation:

  • Induction of chemotaxis in monocytes and T-lymphocytes

  • Enhancement of neutrophil oxidative burst and degranulation

  • Alteration of dendritic cell metabolic profiles

• Signaling pathway regulation:

  • Regulation of the Sonic Hedgehog (SHH) signaling pathway

  • Modulation of TLR4-dependent signaling

• Metabolic effects:

  • Improvement of mitochondrial fitness and redox status

  • Reduction of mitochondrial ROS production

  • Modulation of lactate metabolism in specific cell types

• Cell migration effects:

  • IGFBP6 may have opposing effects on the migration of different ovarian cancer cell lines, indicating context-dependent functions

These IGF-independent functions highlight the multifaceted nature of IGFBP6 and suggest that therapeutic approaches targeting this protein would need to consider these diverse biological activities.

How does the IGFBP6/Sonic Hedgehog/TLR4 signaling axis function?

The IGFBP6/Sonic Hedgehog/TLR4 signaling axis represents an emerging area of research:

• Pathway interconnection: IGFBP6 has been found to regulate the Sonic Hedgehog (SHH) signaling pathway during fibrosis, with this pathway further connecting to TLR4 signaling .

• Microenvironment alterations: This signaling axis drives bone marrow microenvironment alterations in conditions like primary myelofibrosis .

• Role in JAK2 mutation contexts: Interestingly, IGFBP6 expression shows significant increases in PMF patients wild type for JAK2V617F mutation compared to healthy subjects and PMF patients carrying the JAK2V617F mutation .

• Potential therapeutic relevance: Understanding this axis could provide novel therapeutic targets for conditions characterized by dysregulated tissue repair and fibrosis.

Further research is needed to fully elucidate the molecular mechanisms and functional consequences of this signaling axis in different pathological contexts.

What are the current gaps in IGFBP6 research that should be addressed?

Several important knowledge gaps remain in IGFBP6 research:

• Structure-function relationships: More detailed understanding of how specific domains within IGFBP6 (28-240) contribute to its various functions would help design more targeted experimental and therapeutic approaches.

• Receptor identification: While IGFBP6 has IGF-independent functions, the specific receptors mediating these effects remain incompletely characterized.

• Context-dependent effects: The seemingly contradictory roles of IGFBP6 in different cellular contexts (pro- vs. anti-inflammatory, pro- vs. anti-tumorigenic) require further investigation to delineate the molecular switches determining these outcomes.

• Therapeutic potential: Exploration of IGFBP6 as a therapeutic target or biomarker in conditions like cancer, fibrosis, and inflammatory diseases merits systematic investigation.

• Post-translational modifications: How glycosylation and other post-translational modifications affect IGFBP6 function remains underexplored.

Addressing these gaps would significantly advance our understanding of IGFBP6 biology and potentially lead to novel therapeutic strategies for diseases involving dysregulated IGFBP6 signaling.