CD90 Human

What is CD90 and what is its expression pattern across human primary cells?

CD90 (also known as Thy-1) is a cell surface glycoprotein initially identified on mouse thymocytes. In humans, CD90 has been identified on various stem cells and at different expression levels in non-lymphoid tissues including fibroblasts, brain cells, and activated endothelial cells . Research has established specific expression patterns across human primary cells:

• High expression: Normal Human Dermal Fibroblasts (NHDF), activated Human Umbilical Vein Endothelial Cells (HUVEC)

• Low/No expression: Human Corneal Epithelial Cells (CEC), Retinal Pigmented Epithelial (RPE) cells, Normal Human Epidermal Keratinocytes (NHEK), Normal Human Bronchial Epithelial (NHBE) cells, non-activated HUVEC

This differential expression pattern makes CD90 a valuable biomarker for identifying specific cell populations in research and clinical applications.

What methodologies are most effective for detecting and quantifying CD90 expression?

The gold standard methodology for CD90 detection and quantification is flow cytometry using fluorescent-labeled anti-human CD90 monoclonal antibodies (α-CD90 MAb). The standard protocol includes:

• Sample preparation: Trypsinization of adherent cells followed by washing with growth medium

• Cell suspension preparation: Centrifugation at 200 × g for 5 min and resuspension in PBS with 2% FBS

• Antibody staining: Incubation with fluorescein isothiocyanate-conjugated anti-human CD90 antibody (typically 1:200 dilution) for 30 min on ice in dark conditions

• Viability staining: Addition of 7-AAD (1:100) for 10 min before analysis to exclude dead cells

• Analysis: Final centrifugation, resuspension, filtration through a 70 μm filter, and analysis using flow cytometry

Additional methods include direct immunofluorescence for qualitative assessment and immunohistochemistry for tissue samples .

How do CD90+ cells from human adipose tissue demonstrate stem cell properties?

CD34+/CD90+ cells isolated from the stromal vascular fraction (SVF) of human adipose tissue exhibit multiple characteristic stem cell properties:

• Sphere cluster formation: When grown in non-adherent conditions, these cells form sphere clusters, a hallmark of stem-like behavior

• High proliferative capacity: They demonstrate superior proliferation compared to differentiated cells

• Elevated telomerase activity: Their telomerase activity is significantly higher than in differentiated cells, indicating self-renewal potential

• Side population phenotype: A fraction of these cells display the side population phenotype, associated with stemness

• Multipotent differentiation: These cells can rapidly differentiate into adipocytes when cultured in adipogenic medium and into endothelial cells (CD31+/VEGF+/Flk-1+) when appropriately stimulated

• Angiogenic potential: When placed in methylcellulose, they form capillary-like structures and produce high levels of VEGF, measurable by ELISA

These properties collectively establish CD34+/CD90+ adipose-derived stem cells as promising candidates for tissue reconstruction in regenerative medicine, particularly for vascular disease treatments .

How can CD90 expression be leveraged to eliminate contaminating fibroblasts from primary cell cultures?

Fibroblast contamination represents a significant challenge in maintaining specialized human primary cell cultures due to fibroblasts' rapid proliferation. CD90's differential expression can be utilized to develop an effective purification strategy:

• Identification: First confirm fibroblast contamination through CD90 expression analysis using flow cytometry

• Magnetic bead separation:

  • Incubate mixed cell population with α-CD90 antibodies conjugated to magnetic beads

  • Pass the cell suspension through a magnetic column

  • CD90+ cells (predominantly fibroblasts) are retained in the column

  • CD90- target cells flow through and are collected

This method preserves the viability and functionality of the target cells while efficiently removing contaminating fibroblasts. This approach is particularly valuable for developing pure cell populations for cell therapy applications where maintaining specifically differentiated cultures is essential .

What is the prognostic significance of CD90 expression in human cancers?

Meta-analysis data reveals significant correlations between CD90 overexpression and adverse clinical outcomes in cancer patients:

Sensitivity analysis confirmed that these associations remain statistically significant even after accounting for the influence of individual studies, establishing CD90 overexpression as a reliable prognostic biomarker for cancer progression and patient outcomes .

How do CD90+ and CD90- fibro-adipogenic progenitors differ in human skeletal muscle, particularly in metabolic disorders?

CD90 expression distinguishes functionally distinct fibro-adipogenic progenitor (FAP) subpopulations in human skeletal muscle, with significant implications for metabolic disorders:

Phenotypic differences:

• CD90+ FAPs are larger in size

• CD90+ FAPs proliferate faster

• CD90+ FAPs express higher levels of extracellular matrix genes compared to CD90- FAPs

Pathological implications:

• CD90+ FAPs are associated with muscle degeneration in Type 2 diabetic (T2D) patients

• The dynamic relationship between CD90+ FAPs and insulin resistance appears significant, though the precise mechanisms require further investigation

• CD90+ FAPs may contribute to the degenerative muscle microenvironment in metabolic disorders

Single-cell RNA sequencing analysis could further elucidate the heterogeneity of FAPs across healthy subjects, obese individuals, and T2D patients (both insulin-treated and non-treated). This approach would help clarify the molecular changes in FAP subpopulations during disease progression and potential therapeutic interventions .

What experimental approaches can determine the myogenic potential of CD90+ cells?

CD90 has been identified as a crucial molecular signature for cells with myogenic differentiation potential. The following experimental workflow can assess this potential:

• Isolation of CD90+ cells:

  • Flow cytometry sorting using fluorescein isothiocyanate-conjugated anti-human CD90 antibody

  • Protocol includes trypsinization, washing, antibody incubation (1:200 dilution, 30 min on ice), and 7-AAD staining for viability assessment

• Myogenic induction:

  • Transduction with retroviral vector containing doxycycline-inducible MYOD1 expression system (MOI of 200)

  • Culture in differentiation medium with doxycycline and 5 μM DZNep for initial 3 days

  • Continued culture in differentiation medium without DZNep, with medium changes every 3 days

• Assessment of myogenic differentiation:

  • Evaluation of fusion and myotube formation

  • Expression analysis of myogenic markers: MYMK, MYMX, MYOG, and MYHC

  • Trajectory analysis using scRNA-seq data to trace cell fate during differentiation

Research indicates that while CD90 itself doesn't promote differentiation, CD90+ cells respond more effectively to myogenic induction factors like MYOD1, suggesting the presence of additional factors in CD90+ cells that facilitate myogenic differentiation .

How can single-cell RNA sequencing enhance our understanding of CD90+ cell heterogeneity and function?

Single-cell RNA sequencing (scRNA-seq) offers powerful capabilities for investigating CD90+ cell populations:

• Heterogeneity characterization:

  • Identifies distinct subpopulations within CD90+ cells

  • Reveals unique transcriptomic signatures associated with specific functional states

  • Distinguishes CD90+ cells across different tissues and pathological conditions

• Trajectory analysis:

  • Maps differentiation pathways of CD90+ cells

  • Identifies intermediate states during lineage progression

  • Confirms differentiation potential into specific cell types, such as myogenic cells

• Comparative analysis:

  • Enables comparison of CD90+ cell populations between:

    • Healthy individuals and disease states (e.g., T2D patients)

    • Different treatment conditions (e.g., insulin-treated vs. non-treated)

    • Various tissue sources

• Regulatory network identification:

  • Uncovers transcription factors and signaling pathways governing CD90+ cell fate

  • Identifies potential molecular targets for therapeutic intervention

  • Elucidates mechanisms underlying CD90+ cell function in different contexts

This comprehensive approach provides insights that traditional bulk analysis methods cannot deliver, particularly in understanding the complex heterogeneity and dynamic nature of CD90+ cell populations in human tissues .

How can CD90-expressing cells contribute to regenerative medicine approaches?

CD90-expressing cells offer significant potential for regenerative medicine applications:

• Vascular regeneration:

  • CD34+/CD90+ adipose-derived stem cells (ASCs) demonstrate robust angiogenic potential

  • These cells form capillary-like structures in methylcellulose

  • They produce high levels of VEGF, promoting vascularization

  • Their endothelial differentiation capacity makes them suitable for vascular tissue engineering

• Skeletal muscle regeneration:

  • CD90+ subpopulations from urine-derived cells (UDCs) show high myogenic potential when induced with MYOD1

  • These cells could serve as a cell source for muscular disease modeling

  • Potential applications in testing therapeutic approaches for conditions like Duchenne muscular dystrophy (DMD)

• Translational considerations:

  • Accessibility of source tissues (adipose tissue, urine) offers practical advantages

  • Non-invasive collection methods reduce patient burden

  • Autologous applications minimize immunological concerns

  • Established protocols for isolation and expansion facilitate clinical translation

These characteristics position CD90-expressing cells as promising candidates for tissue reconstruction strategies, particularly for patients requiring treatments for vascular disease and muscle disorders .

What methodological challenges exist in isolating and characterizing CD90+ cells for clinical applications?

Several methodological challenges must be addressed when isolating and characterizing CD90+ cells for clinical applications:

• Heterogeneity management:

  • CD90+ populations exhibit significant heterogeneity within and across tissues

  • Standardized protocols for isolating specific CD90+ subpopulations are needed

  • Functional characterization is essential to distinguish therapeutically relevant subsets

• Purification efficiency:

  • Achieving high purity CD90+ isolates requires optimization of antibody concentrations and flow cytometry parameters

  • Contamination with CD90- cells may affect downstream applications

  • Magnetic separation technologies must balance yield with purity

• Functional assessment:

  • Standardized assays for assessing CD90+ cell functionality across different contexts

  • Correlation of in vitro findings with in vivo performance

  • Long-term stability and safety evaluation of isolated populations

• Clinical-grade processing:

  • Translation of research protocols to GMP-compliant procedures

  • Development of xeno-free isolation and culture conditions

  • Scalability considerations for therapeutic applications

• Regulatory considerations:

  • Establishing quality control metrics for CD90+ cell products

  • Defining release criteria for clinical applications

  • Long-term follow-up strategies for safety assessment

Addressing these challenges through rigorous methodological development and standardization will facilitate the translation of CD90+ cell-based therapies from bench to bedside .

What are the key unresolved questions regarding CD90 function in human tissues?

Despite significant advances in CD90 research, several critical questions remain unresolved:

• Regulatory mechanisms:

  • How is CD90 expression regulated following injury and during tissue regeneration?

  • What molecular pathways control CD90 expression in different cell types?

  • What is the functional significance of dynamic CD90 regulation?

• Developmental origins:

  • What is the hierarchical relationship between CD90+ and CD90- cells within the same tissue?

  • Do CD90+ cells represent a distinct lineage or a functional state?

  • What lineage tracing experiments could clarify the developmental origins of CD90+ cells?

• Functional mechanisms:

  • What is the precise role of CD90 in controlling cell fate and behavior?

  • Does CD90 function primarily as a signaling molecule or adhesion receptor?

  • What are the interaction partners for CD90 in different cellular contexts?

• Disease associations:

  • How do metabolic changes associated with Type 2 diabetes affect CD90+ muscle FAPs?

  • What is the cross-talk between CD90+ FAPs and muscle stem cells in health versus disease?

  • Do CD90+ cells become resistant to insulin, and what are the implications?

• Therapeutic potential:

  • What factors cooperate with CD90 to determine cell fate?

  • How can CD90+ cells be optimally manipulated for therapeutic applications?

  • What are the safety considerations for clinical applications of CD90+ cells?

Addressing these questions will require integration of advanced technologies including single-cell transcriptomics, lineage tracing, and functional genomics approaches .

How might technological advances enhance CD90 research and applications?

Emerging technologies offer transformative potential for advancing CD90 research:

• Single-cell multi-omics:

  • Integration of transcriptomics, proteomics, and epigenomics at single-cell resolution

  • Comprehensive characterization of CD90+ cell states and functional heterogeneity

  • Identification of regulatory networks governing CD90+ cell behavior

• Spatial transcriptomics:

  • Mapping CD90+ cell distribution and interactions within tissue microenvironments

  • Understanding niche-dependent regulation of CD90+ cell function

  • Elucidating cell-cell communication networks involving CD90+ cells

• CRISPR-based functional genomics:

  • Systematic interrogation of genes regulating CD90 expression and function

  • Identification of factors that cooperate with CD90 in determining cell fate

  • Development of strategies to enhance CD90+ cell therapeutic potential

• Organoid and microphysiological systems:

  • Modeling CD90+ cell behavior in tissue-specific contexts

  • Studying dynamic interactions between CD90+ cells and other cell types

  • High-throughput screening of factors influencing CD90+ cell function

• Advanced computational modeling:

  • Prediction of CD90+ cell trajectory and fate decisions

  • Integration of multi-modal data for comprehensive understanding of CD90 biology

  • AI-assisted discovery of novel therapeutic applications for CD90+ cells

These technological advances will facilitate deeper understanding of CD90 biology and accelerate translation of findings into clinical applications for regenerative medicine and disease treatment .