Recombinant Human C-C motif chemokine 27 protein (CCL27) (Active)

What is CCL27 and what are its alternative nomenclatures in scientific literature?

CCL27, also known as Cutaneous T-cell-attracting chemokine (CTACK), is a member of the CC chemokine family that primarily functions as a chemotactic factor attracting skin-associated memory T-lymphocytes. This protein plays a crucial role in mediating the homing of lymphocytes to cutaneous sites through binding to its receptor CCR10. In the scientific literature, CCL27 may be referenced by several alternative names, including ILC, SCYA27, ESkine, IL-11 R-alpha-locus chemokine, Skinkine, and Small-inducible cytokine A27 . Understanding these nomenclature variations is essential when conducting comprehensive literature searches or when comparing research findings across different publications.

What is known about CCL27's physiological expression pattern and regulation?

CCL27 is constitutively expressed by keratinocytes in the skin and is upregulated in response to inflammatory stimuli and during wound healing processes. The expression of CCL27 is tightly regulated and can be increased under various pathological conditions, including inflammatory skin disorders and certain malignancies. In normal physiology, CCL27 operates as a homeostatic chemokine involved in immune surveillance of the skin. During inflammation, its expression increases to recruit T cells expressing the chemokine receptor CCR10 to the affected area. The protein cooperates with CCL17/TARC in inducing the migration of cutaneous lymphocyte antigen (CLA) positive memory T cells to the skin during inflammatory responses . This selective expression pattern makes CCL27 particularly relevant for research on skin immunology and dermatological conditions.

What are the optimal storage and reconstitution protocols for recombinant CCL27?

For optimal results with recombinant CCL27, researchers should adhere to the following storage and reconstitution protocols:

Reconstituted protein should be aliquoted to minimize freeze-thaw cycles, as repeated cycles can significantly reduce protein activity. For long-term storage, keep the lyophilized product at -20°C to -80°C. Once reconstituted, the protein can be stored at -80°C for up to 3 months, though specific activity may gradually decrease over time .

How should researchers determine the appropriate concentration of CCL27 for cell-based assays?

• Cell type (primary cells vs. cell lines)

• Receptor expression levels (endogenous vs. overexpressed CCR10)

• Specific readout (migration, calcium flux, signaling, etc.)

• Experimental duration

For migration assays, a range of 0.1-0.4 μg/mL is typically effective . For signaling studies such as beta-arrestin recruitment, researchers should begin with concentrations around the reported ED50 value and adjust accordingly. When establishing a new assay, it is advisable to perform a dose-response experiment using concentrations ranging from 10 ng/mL to 1 μg/mL to determine the optimal concentration for the specific experimental setup.

What cell models are most appropriate for studying CCL27 function?

The selection of appropriate cell models for studying CCL27 function depends on the specific research question being addressed. Based on published literature, the following models have proven effective:

When studying receptor-ligand interactions specifically, researchers have successfully used U2OS cells overexpressing human CCR10 with beta-arrestin and beta-galactosidase complementation systems . For migration studies, researchers should consider using primary human lymphocytes or established T cell lines expressing CCR10. The choice of model should align with the specific aspect of CCL27 biology being investigated.

How does the CCR10/CCL27-CCL28 axis contribute to tumor development?

The CCR10/CCL27-CCL28 axis plays complex and sometimes contradictory roles in tumor development, making it a fascinating area for cancer research. Current evidence suggests dual functions depending on the tumor microenvironment and immune context:

• Pro-tumorigenic effects:

  • Promotes tumor cell survival through activation of PI3K/AKT signaling

  • Enhances tumor cell migration and metastatic potential

  • May contribute to immune evasion mechanisms

• Anti-tumorigenic effects:

  • Recruits cytotoxic T cells and natural killer cells to the tumor site

  • Can enhance anti-tumor immune responses in some contexts

The specific outcome appears to be dependent on:

• The balance between CCL27 and CCL28 expression

• The tumor type and stage

• The composition of immune infiltrates

• Whether CCR10 is expressed by tumor cells, immune cells, or both

Recent research has also implicated this axis in lymphatic endothelial cell migration, which could influence tumor lymphangiogenesis and metastatic spread. A study published in Cancer Research demonstrated that CCL27/CCL28-CCR10 chemokine signaling mediates migration of lymphatic endothelial cells, potentially contributing to tumor progression .

What methodologies are recommended for measuring CCL27-induced cell migration?

For robust assessment of CCL27-induced cell migration, researchers should consider the following methodological approaches:

• Transwell migration assays:

  • Use 5-8 μm pore size depending on cell type

  • Optimize incubation time (typically 3-6 hours for lymphocytes)

  • Include positive controls (other established chemokines)

  • Test CCL27 concentrations in the range of 0.1-0.4 μg/mL

  • Quantify migrated cells by flow cytometry or colorimetric methods

• Real-time cell migration tracking:

  • Employ live-cell imaging systems with automated tracking

  • Calculate directionality and velocity parameters

  • Analyze chemotactic index (directed vs. random movement)

• 3D migration in extracellular matrix:

  • Incorporate ECM components (collagen, fibronectin)

  • Simulate tissue environments more accurately

  • Account for CCL27 binding to glycosaminoglycans in the matrix

• In vivo migration models:

  • Utilize fluorescently labeled cells and intravital microscopy

  • Employ adoptive transfer experiments with labeled CCR10+ cells

  • Use tissue-specific CCL27 expression systems

When analyzing results, researchers should differentiate between chemokinesis (increased random movement) and chemotaxis (directed movement) by including appropriate controls such as checkerboard assays where CCL27 is present in both upper and lower chambers. Additionally, blocking antibodies against CCR10 should be used to confirm specificity of the observed migration response.

What is known about CCL27's role in inflammatory skin conditions and its potential as a therapeutic target?

CCL27 has been implicated in several inflammatory skin conditions, with elevated serum levels reported in atopic dermatitis, psoriasis vulgaris, and mycosis fungoides . The protein plays a critical role in these conditions through several mechanisms:

• Atopic dermatitis:

  • Serum CCL27 levels correlate with disease severity

  • Contributes to recruitment of Th2 cells to lesional skin

  • May serve as a biomarker for disease activity and treatment response

• Psoriasis vulgaris:

  • Upregulated in lesional keratinocytes

  • Promotes infiltration of CLA+ memory T cells

  • Forms part of the inflammatory cascade with other chemokines

• Cutaneous T cell lymphoma (including mycosis fungoides):

  • Elevated expression correlates with disease progression

  • May contribute to malignant T cell localization in the skin

These findings suggest CCL27 as a potential therapeutic target, with several approaches under investigation:

Therapeutic targeting must consider the homeostatic functions of CCL27 in normal skin, and strategies may need to focus on reducing excessive levels rather than complete blockade. Combination approaches targeting multiple chemokines may prove more effective than single-agent strategies in complex inflammatory conditions.

What are the key differences between E. coli-derived and HEK293-derived recombinant CCL27 that researchers should consider?

When selecting between E. coli-derived and HEK293-derived recombinant CCL27 for research applications, several important differences should be considered:

How can researchers validate the biological activity of recombinant CCL27 before use in critical experiments?

Validation of recombinant CCL27 biological activity is essential to ensure experimental reliability. The following approaches are recommended:

• Receptor binding assays:

  • Radio- or fluorescently-labeled CCL27 binding to CCR10+ cells

  • Competition binding with unlabeled protein

  • Surface plasmon resonance for binding kinetics

• Functional assays:

  • Beta-arrestin recruitment assay using U2OS cells overexpressing human CCR10/beta-arrestin/beta-galactosidase complementation system (as used by commercial suppliers)

  • Calcium flux assays in CCR10-expressing cells

  • ERK1/2 phosphorylation or other downstream signaling events

• Cell migration assays:

  • Transwell migration of CCR10+ lymphocytes

  • Scratch wound assays for keratinocyte migration

• Positive controls:

  • Include commercially validated CCL27 as reference

  • Compare to reported ED50 values (≤ 295 ng/ml)

A properly validated batch of CCL27 should demonstrate dose-dependent activity in at least one functional assay, with potency comparable to reference standards. For critical experiments, researchers should consider performing validation using the same cell type and readout that will be used in the final experiments to account for cell-specific differences in response.

What are common pitfalls in CCL27 research and how can they be avoided?

Researchers working with CCL27 should be aware of several common pitfalls that can compromise experimental outcomes:

Additionally, researchers should be aware that CCL27 binds to glycosaminoglycans in the extracellular matrix, which can affect its distribution and availability in complex experimental systems. When designing in vitro assays, consider the impact of matrix components on chemokine presentation and function. For in vivo studies, remember that CCL27 interacts with determinants on the surface of fibroblasts and endothelial cells, which may influence its biodistribution beyond simple diffusion .

What are emerging areas of research regarding CCL27 function beyond its established role in skin immunity?

While CCL27's role in skin immunity is well-established, several emerging areas of research are expanding our understanding of this chemokine's functions:

• Wound healing and tissue regeneration:

  • CCL27 induces migration of keratinocyte precursors from bone marrow to injured skin

  • Contributes to re-epithelialization and tissue repair processes

  • Potential applications in regenerative medicine and wound healing therapies

• Cancer immunotherapy:

  • Modulation of CCL27 as a strategy to enhance T cell infiltration into tumors

  • Development of CCL27-based chimeric proteins to direct immune cells to tumors

  • Role in resistance to anticancer kinase inhibitors

• Lymphatic vessel development and function:

  • CCL27/CCL28-CCR10 signaling mediates lymphatic endothelial cell migration

  • Implications for lymphangiogenesis in development and disease

  • Potential target for controlling lymphatic metastasis

• Non-skin epithelial immunity:

  • Emerging evidence for CCL27/CCR10 function in mucosal immunity

  • Overlap with CCL28 functions at mucosal surfaces

  • Contributions to epithelial barrier function beyond the skin

These expanding areas of research suggest CCL27 has broader physiological and pathological roles than initially recognized, presenting new opportunities for therapeutic development and diagnostic applications. Researchers entering these fields should consider interdisciplinary approaches combining immunology, oncology, and regenerative medicine perspectives.

What novel methodologies are being developed to study the CCL27-CCR10 axis in complex tissue environments?

The study of CCL27-CCR10 interactions in complex tissue environments is advancing through several innovative methodologies:

• Organoid and 3D tissue models:

  • Skin equivalents incorporating keratinocytes, fibroblasts, and immune cells

  • Microfluidic "organ-on-chip" systems modeling CCL27 gradients

  • Co-culture systems to study cell-cell communication mediated by CCL27

• Advanced imaging techniques:

  • Multiphoton intravital microscopy to track CCR10+ cell migration in living tissues

  • Mass cytometry imaging to map CCL27 distribution and cell responses

  • CODEX (CO-Detection by indEXing) for highly multiplexed tissue imaging

• Single-cell technologies:

  • scRNA-seq to identify CCL27-responsive cell populations

  • CyTOF analysis of signaling events at single-cell resolution

  • Spatial transcriptomics to map chemokine-receptor interactions in tissue context

• Chemokine reporter systems:

  • CCR10 biosensors for real-time monitoring of receptor activation

  • Conditional CCL27 expression systems for temporal control

  • CRISPR-based genetic screens for CCL27-CCR10 pathway components

These methodologies enable researchers to move beyond reductionist approaches and study CCL27 function in contexts that more closely resemble in vivo conditions. The integration of these approaches with computational modeling is particularly promising for understanding how CCL27 gradients establish and function within complex tissue architectures.