SPP1 Human, Active

What are the primary biological functions of SPP1?

SPP1 is involved in numerous biological processes, making it a versatile protein in research contexts:

• Bone remodeling through strong binding to hydroxyapatite

• Formation of mineralized matrix in bone tissue

• Mediation of cell-matrix interactions across various tissue types

• Immune functions in fibroblasts, macrophages, and lymphocytes during inflammation and wound healing

• Protection against cardiac ischemia-reperfusion injury through late preconditioning mechanisms

• Involvement in neuroinflammatory processes, particularly through microglial activation

• Regulation of cancer stem cell maintenance and tumor progression

How can I validate the functional activity of recombinant SPP1 protein in my experimental system?

The functional activity of SPP1 Human, Active protein can be validated through cell adhesion assays. A standard approach involves:

• Immobilize SPP1 on culture plates at a concentration of 10μg/ml

• Add HEK293 human embryonic kidney cells to the coated plates

• Measure the adhesion effect, which should reach approximately 40% or greater compared to controls

This assay leverages SPP1's native function in mediating cell-matrix interactions and serves as a reliable indicator that the recombinant protein retains its biological activity.

What experimental models are most appropriate for studying SPP1 function?

Based on recent research, several experimental models have proven effective for studying SPP1:

When designing experiments, incorporating both in vitro and ex vivo approaches provides complementary insights into SPP1 function in different contexts .

What are effective approaches for modulating SPP1 expression in experimental systems?

Several approaches have been validated for manipulating SPP1 expression in research contexts:

• siRNA-mediated knockdown: Effective for transient reduction of SPP1 expression. Studies have achieved approximately 80% reduction in SPP1 mRNA levels using this approach .

• shRNA-mediated interference: Provides more stable knockdown for longer-term experiments, particularly useful in sphere formation studies that run for 7+ days .

• Targeting upstream regulators: Knockdown of transcription factors like GLI1 that regulate SPP1 expression can indirectly modulate SPP1 levels .

• Forced differentiation: In cancer stem cell models, inducing differentiation has been shown to reduce SPP1 expression, allowing for study of its role in stemness maintenance .

When implementing these approaches, it is critical to validate knockdown efficiency through qPCR and/or protein level measurements before interpreting downstream functional effects.

How is SPP1 associated with cognitive decline and Alzheimer's disease?

Recent research from the Religious Orders Study and Rush Memory and Aging Project (ROSMAP) has revealed significant associations between SPP1 and neurodegenerative processes:

• Expression of SPP1 and its encoded protein osteopontin is associated with faster cognitive decline in older adults .

• Higher SPP1 levels correlate with greater odds of common neuropathologies, including those characteristic of Alzheimer's disease .

• At the single-cell level, SPP1 appears to be linked to specific microglial populations that are associated with Alzheimer's disease and cerebral amyloid angiopathy .

This evidence suggests SPP1 may serve as both a biomarker for cognitive decline and a potential therapeutic target in neurodegenerative conditions. The economic impact of these findings is considerable, given that global dementia costs are expected to surpass US$2.8 trillion by 2030 .

How does SPP1 interact with microglia in the context of neuroinflammation?

SPP1's role in neuroinflammation appears to be mediated through specific interactions with microglial cells:

• SPP1 is expressed by CD11c+ cells and is associated with microglial activation in neuroinflammatory conditions .

• ITGAX (which encodes CD11c) is highly expressed in specific microglial subpopulations that are linked to Alzheimer's disease pathology .

• These SPP1-expressing microglial subsets may represent a disease-associated microglial phenotype that contributes to neuroinflammatory processes .

This relationship between SPP1 and microglial activation provides a mechanistic link between SPP1 expression and neurodegenerative outcomes, suggesting targeted approaches to modulating neuroinflammation might benefit from focusing on the SPP1 pathway.

What mechanisms regulate SPP1 expression in cancer cells?

SPP1 expression in cancer cells is regulated through several mechanisms that distinguish it from normal tissue:

• Transcription factor regulation: SPP1 is regulated by stemness transcription factors like GLI1 and OCT4 that are expressed in glioblastoma cells but not in normal astrocytes. Chromatin immunoprecipitation (ChIP) assays have demonstrated direct binding of GLI1 to the proximal promoter of the SPP1 gene in human glioma cell lines but not in normal human astrocytes .

• Promoter binding sites: Two putative GLI1 binding sites have been identified in the human SPP1 gene promoter, and the stemness factor Oct4 has been shown to bind within the first intron of the murine SPP1 gene .

• Epigenetic regulation: The Nencki Genomics Database has been used to identify conserved sequence motifs and epigenetic modifications in the non-coding regions of the SPP1 gene, suggesting epigenetic control mechanisms .

• Feedback loops: Knockdown of GLI1 in glioma cells significantly reduces SPP1 mRNA and protein production, indicating regulatory feedback mechanisms .

These findings suggest that cancer cells reactivate embryonic types of SPP1 transcriptional regulation, potentially contributing to their abnormal growth patterns and stem-like properties.

How does SPP1 contribute to cancer stem cell maintenance?

SPP1 plays a crucial role in maintaining cancer stem cell properties, particularly in glioma:

• SPP1 is overexpressed in glioma initiating cells defined by high rhodamine 123 efflux, sphere-forming capacity, and stemness marker expression .

• When glioma sphere cultures undergo forced differentiation, SPP1 expression is reduced, suggesting its specific role in the undifferentiated state .

• Experimental knockdown of SPP1 in LN18 glioma cells reduces their ability to form spheres by approximately 20%, even with a transient reduction in SPP1 levels .

• SPP1-CD44 signaling appears to be critical for maintenance of glioma initiating cells through both autocrine and paracrine mechanisms .

These findings indicate that targeting SPP1 or its downstream signaling could potentially disrupt cancer stem cell maintenance, offering a therapeutic strategy for reducing tumor recurrence and therapy resistance.

How does SPP1 expression correlate with cancer prognosis and how can this be reconciled with contradictory findings?

The relationship between SPP1 expression and cancer prognosis appears to be context-dependent:

In certain cancers:

• Expression of Osteopontin (SPP1) and CD44 in hepatocellular carcinoma is linked to advanced tumor stage and poor prognosis

• SPP1 is the most over-expressed gene in intrahepatic cholangiocarcinoma, suggesting a negative prognostic role

• SPP1 overexpression is associated with interstitial lung diseases and their progression

• In penile cancer, survival analysis based on immunohistochemistry data shows that the high-SPP1 group had a better prognosis than the low-SPP1 group

These contradictions might be explained by:

• Tissue-specific functions of SPP1

• Different SPP1 isoforms expressed across cancer types

• Varying interactions with the immune microenvironment (the high-SPP1 group in penile cancer showed distinct immune cell infiltration patterns)

• Cancer-specific signaling pathway interactions

How does SPP1 modulate immune cell populations in disease contexts?

SPP1 exhibits significant associations with various immune cell populations, particularly in cancer microenvironments:

• Bioinformatic analyses using algorithms like CIBERSORT and ssGSEA have revealed that SPP1 expression is significantly associated with multiple immune cell types including B cells, CD8+ T cells, CD4+ T cells, macrophages, helper T cells, neutrophils, and dendritic cells .

• Immunohistochemical studies have shown that high-SPP1 expression correlates with relatively high expression of CD16 (a marker for NK cells and certain macrophages) and relatively low expression of CD4 (a marker for helper T cells) .

• GSVA analysis indicates that high-SPP1 expression is significantly associated with immune-related pathways such as PD-L1 expression, the PD-1 checkpoint pathway in cancer, and TNF signaling .

These findings suggest SPP1 may serve as an immunomodulatory molecule that shapes the immune landscape in disease contexts, with potential implications for immunotherapy approaches.

Could SPP1 serve as a biomarker for immunotherapy response?

Given SPP1's associations with immune-related pathways, it shows promise as a potential biomarker for immunotherapy response:

• The association between SPP1 and immune checkpoint pathways (PD-1/PD-L1) suggests it might predict responsiveness to checkpoint inhibitor therapies .

• SPP1's role in modulating immune cell populations could affect the tumor immune microenvironment and subsequently impact immunotherapy efficacy .

• Differential immune cell profiles observed in high versus low SPP1-expressing tumors might help stratify patients for appropriate immunotherapy approaches .

To validate SPP1 as an immunotherapy biomarker, researchers should:

• Analyze SPP1 expression in relation to clinical responses to various immunotherapies

• Investigate how SPP1 mechanistically influences the tumor immune microenvironment

• Develop standardized assays for measuring SPP1 in clinical specimens with high reproducibility

What methodological approaches are recommended for comprehensive analysis of SPP1 in research contexts?

For rigorous investigation of SPP1 in research settings, a multi-modal approach is recommended:

Integration of these approaches, as demonstrated in the ROSMAP study with over 1200 samples, provides the most comprehensive understanding of SPP1 biology in both normal and pathological contexts .

How can discrepancies in experimental findings about SPP1 be addressed methodologically?

When encountering contradictory findings regarding SPP1, researchers should consider several methodological approaches:

• Isoform-specific analysis: Different SPP1 isoforms may have distinct functions. Researchers should use techniques that distinguish between isoforms rather than measuring total SPP1 .

• Context-dependent investigation: SPP1 function may vary by tissue type and disease state. Experimental designs should account for these variables through appropriate controls and comparative analyses .

• Integrated multi-omics approach: Combining transcriptomic, proteomic, and functional readouts can resolve apparent contradictions by revealing regulatory mechanisms not captured by single-modality approaches .

• Statistical considerations: When analyzing SPP1 expression data, researchers should apply appropriate statistical methods, such as dividing datasets into high and low SPP1 groups based on median expression values and using chi-square tests for dichotomous variables and log-rank tests for survival analyses .

By applying these methodological approaches, researchers can develop more nuanced understandings of SPP1's complex biology across different experimental and disease contexts.