CDH11 Human

What is CDH11 and how does it differ from other cadherins in the human genome?

CDH11, also known as cadherin-11 or OB-cadherin, is a type-II classical cadherin that mediates calcium-dependent cell-cell adhesion. Unlike type-I classical cadherins (such as E-cadherin/CDH1) that primarily maintain cohesive tissue integrity, CDH11 plays a more intricate role in tissue orchestration and is involved in functions such as migration and differentiation .

Methodologically, CDH11 can be distinguished from other cadherins by:

• Its unique amino acid sequence in the adhesive interface

• Formation of weaker adhesive bonds with a higher turnover rate compared to type-I cadherins

• Distinct expression patterns in mesenchymal tissues and developing neural structures

• Specific antibody recognition patterns using validated antibodies targeting the C-terminus

What are the recommended experimental methods for detecting CDH11 expression in human tissues?

For reliable detection of human CDH11 in research settings:

Western Blotting:

• Use validated antibodies such as mouse IgG1 clone 5B2H5 (Thermo Fisher #32-1700) at 2 μg/ml concentration

• Include appropriate loading controls and positive control cell lines

• Compare against CDH11 knockdown/knockout controls when possible

Immunocytochemistry/Immunofluorescence:

• Multiple antibodies have been validated including goat anti-CDH11 (Santa Cruz sc-6461) and mouse anti-CDH11

• Optimal working dilutions typically range from 3-4 μg/ml

• Co-staining with membrane or junction markers helps confirm localization

Immunoprecipitation:

• Typically requires 7 μg of antibody per reaction

• Verify specificity through reverse co-immunoprecipitation approaches

How does CDH11 regulate mesenchymal stem cell differentiation pathways?

CDH11 exerts lineage-specific effects on mesenchymal stem cell (MSC) differentiation:

Adipogenic Differentiation:

• CDH11 is indispensable for adipogenic differentiation of human MSCs

• Knockdown of CDH11 significantly inhibits adipogenic differentiation in both monolayer and aggregate cultures

• This effect appears consistent across different culture dimensionalities

Osteogenic Differentiation:

• The role of CDH11 appears to be context-dependent

• In aggregate cultures, CDH11 may influence mineralized matrix deposition

• CDH11 knockdown has been shown to decrease expression of type VI collagen, potentially affecting osteogenic capacity

Methodologically, researchers should assess differentiation through multiple complementary approaches:

• Histological staining (Oil Red O for adipogenic, Alizarin Red S for osteogenic)

• Gene expression analysis of lineage-specific markers

• Protein analysis of differentiation-associated factors

• Matrix component quantification

What mechanisms explain CDH11's influence on the extracellular matrix composition?

CDH11 regulates extracellular matrix (ECM) composition through several mechanisms:

TGFβ1 Pathway Regulation:

• CDH11 modulates the TGFβ1 pathway through SMAD2/3 signaling

• This regulation occurs in a temporal manner, affecting early and late ECM production differently

• The pathway represents a key mechanism by which CDH11 controls ECM synthesis despite lacking intrinsic signaling activity

ECM Component Regulation:

• Cadherin-11 regulates both collagen and fibronectin expression

• These effects can be observed using Western blotting and immunofluorescence analysis

• Analysis should be performed at both early and late time points to capture temporal dynamics

RTK-Mediated Signaling:

• CDH11 influences receptor tyrosine kinase (RTK) signaling, particularly through PDGFRβ

• These changes in RTK profiles lead to downstream alterations in MAPK pathway activation

• The RTK-MAPK axis represents another mechanism by which CDH11 influences cell behavior and potentially matrix production

For experimental approaches, researchers should:

• Combine genetic manipulation (CDH11 knockdown/knockout) with pathway inhibition studies

• Assess both intracellular signaling (phospho-SMAD2/3, MAPK components) and extracellular matrix outcomes

• Consider temporal dynamics by analyzing multiple time points

What are the most effective genetic manipulation approaches for studying CDH11 function?

Knockdown Strategies:

• shRNA-mediated stable knockdown has been effectively used to create sh-CDH11 cell lines

• Typical knockdown efficiency should be validated at both mRNA and protein levels

• Multiple target sequences should be tested to identify optimal knockdown

Knockout Models:

• Cdh11 knockout mice (Cdh11^tm1Mta^/HensJ available from Jackson Laboratory) provide a complete loss-of-function model

• CRISPR-Cas9 gene editing can be employed for human cell lines

• Inducible knockout systems allow for temporal control of CDH11 expression

Rescue Experiments:

• Re-expression of CDH11 in knockout/knockdown models should be used to confirm specificity of observed phenotypes

• Domain-specific mutants can identify functional regions responsible for specific activities

• Species-matched CDH11 should be used for optimal rescue

How should researchers design experiments to distinguish between dimensional effects and cadherin-specific effects in 3D cell cultures?

When studying CDH11 in different culture dimensions:

Experimental Design Considerations:

• Compare 2D monolayers vs. 3D aggregates using the same cell numbers

• Include CDH11 knockdown/knockout in both 2D and 3D systems

• Analyze cadherin expression patterns in both culture systems, as they may differ significantly

Control Approaches:

• Test scaffold-free aggregates vs. cells embedded in biomaterials (e.g., alginate hydrogels)

• Compare systems with and without cell-ECM interaction motifs (e.g., RGD peptides)

• Vary aggregate size while maintaining consistent cell numbers

Analysis Methods:

• Assess both morphological outcomes and molecular signaling changes

• Quantify differentiation markers across culture systems

• Measure CDH11 and partner protein levels in different dimensional contexts

Research has demonstrated that "cell culture dimensionality influences cell fate through cadherin-2 and cadherin-11," indicating that the dimensional environment itself affects cadherin function and subsequent cellular behaviors .

How does CDH11 influence neural circuit development and what are the implications for autism spectrum disorder?

CDH11 plays significant roles in neural development with implications for autism spectrum disorder (ASD):

Neuronal Morphology Effects:

• Cdh11^-/-^ knockout mice exhibit increased dendritic complexity in hippocampal neurons

• This altered morphology correlates with changes in neuronal and synaptic activity

• The effects suggest CDH11 normally acts as a constraint on excessive dendritic branching

Synaptic Protein Regulation:

• Loss of CDH11 leads to increased levels of excitatory synaptic markers, including:

  • Neuroligin-1 (an excitatory synaptic marker)

  • Postsynaptic density protein-95 (PSD-95)

• These changes suggest CDH11 influences excitatory synapse formation or stability

Relationship with Other Cadherins:

• CDH11 knockout leads to compensatory upregulation of cadherin-8 (CDH8)

• This relationship mirrors expression patterns observed in autism, where CDH8 is upregulated and CDH11 is downregulated

• The reciprocal regulation suggests interconnected roles in neurodevelopment

These findings suggest that alterations in CDH11 expression may contribute to the neural circuit abnormalities observed in autism spectrum disorder .

What human model systems are most informative for studying CDH11's role in neurodevelopmental disorders?

iPSC-Derived Neural Models:

• Induced pluripotent stem cell (iPSC)-derived cortical neural precursor cells (NPCs) from individuals with autism show altered CDH11 expression

• These cells exhibit downregulated CDH11 and upregulated CDH8, mirroring patterns in mouse models

• iPSC-derived models allow for patient-specific investigation of cadherin dysregulation

Cortical Organoids:

• 3D cortical organoids generated from individuals with autism also display CDH11/CDH8 expression changes

• These complex models recapitulate aspects of human cortical development

• Organoids allow assessment of cadherin function in a physiologically relevant 3D context

Complementary Approaches:

• Compare findings between rodent models and human cellular systems

• Validate mechanisms identified in animal models using human-derived cells

• Correlate expression patterns with specific clinical phenotypes in ASD patients

When utilizing these models, researchers should:

• Establish isogenic control lines when possible

• Compare multiple patient-derived lines to account for genetic heterogeneity

• Assess both cellular phenotypes and molecular signatures

• Validate findings across different model systems

What are the critical controls for CDH11 antibody-based detection methods?

Antibody Validation Controls:

• CDH11 knockout/knockdown cells or tissues serve as negative controls

• Overexpression systems provide positive controls

• Competing peptides can confirm specificity of antibody binding

• Multiple antibodies targeting different epitopes should yield consistent results

Recommended Antibodies and Dilutions:
Based on published research, validated antibodies include:

• Mouse monoclonal (IgG1) antibody clone 5B2H5 targeting C-terminus (Thermo Fisher #32-1700)

  • Western blot: 2 μg/ml

  • Immunoprecipitation: 7 μg per reaction

• Goat polyclonal antibody targeting C-terminus (Santa Cruz sc-6461)

  • Western blot: 1:500 dilution

• Mouse antibody targeting N-terminus (DSHB CAD8-1)

  • Western blot: 0.5 μg/ml

  • Immunocytochemistry: 3-4 μg/ml

  • Immunoprecipitation: 2 μg per reaction

How can researchers address variable expression of CDH11 across different experimental conditions?

Standardization Approaches:

• Maintain consistent culture conditions (passage number, confluence, serum batch)

• Establish baseline expression profiles before experimental manipulation

• Include multiple reference genes for qPCR normalization

• Account for cell density effects on cadherin expression

Contextual Factors to Control:

• Culture dimensionality significantly affects cadherin expression and function

• Cell-cell contact density influences cadherin clustering and activity

• Media composition, particularly calcium concentration, affects cadherin function

• Consider temporal dynamics by analyzing multiple time points

Quantification Recommendations:

• Use absolute quantification methods where possible

• Normalize protein expression to multiple loading controls

• Employ digital PCR for more precise mRNA quantification

• Combine mRNA and protein level measurements to confirm changes

By implementing these comprehensive controls and standardization approaches, researchers can obtain more reliable and reproducible results in CDH11 studies across various experimental systems.