FGF 1 Mouse, His

What is the expression pattern of FGF1 in mouse tissues?

FGF1 shows a distinctive tissue-specific expression pattern in mice. The Fgf1 gene contains at least three upstream promoters that are alternatively spliced to the first protein coding exon, giving rise to different mRNA variants (1A, 1B, and 1G) . Among these variants, the Fgf1A transcript is predominantly expressed in the heart and kidney . Unlike FGF2, FGF1 is expressed relatively little outside the nervous system, where it is predominantly expressed by neurons . During development, FGF1 is expressed at low concentrations until embryonic day 16 (E16), after which it rises to adult levels .

To study FGF1 expression patterns experimentally:

• Use RT-PCR to differentiate between Fgf1 transcript variants in various tissues

• Employ immunohistochemistry with FGF1-specific antibodies for protein localization

• Consider using the F1A-CreERT2 mouse line, which allows for lineage tracing of Fgf1A-expressing cells in vivo

What phenotype do FGF1 knockout mice exhibit?

Interestingly, despite FGF1's involvement in multiple biological processes, FGF1 knockout mice show no severe deficits or abnormalities . This is documented in comprehensive knockout phenotype tables where FGF1 is described as having a "Normal" phenotype . This lack of an overt phenotype suggests potential functional redundancy within the FGF family or compensatory mechanisms that activate during development.

When working with FGF1 knockout mice:

• Perform careful physiological assessments beyond gross morphology

• Consider challenging the mice with stress conditions to reveal conditional phenotypes

• Investigate molecular and cellular changes that might not manifest as obvious physical traits

How can I effectively use the F1A-CreERT2 mouse model for FGF1 research?

The F1A-CreERT2 transgenic mouse line enables time-dependent and lineage tracing of Fgf1A-expressing cells in vivo . This model uses the Fgf1A promoter to drive expression of tamoxifen-inducible Cre recombinase (CreERT2).

To effectively use this model:

• Cross F1A-CreERT2 mice with reporter mice (e.g., ROSA26 reporter mice)

• Administer tamoxifen to activate Cre-mediated recombination at your desired timepoint

• Analyze reporter expression (e.g., LacZ or RFP) to identify cells where the Fgf1A promoter was active

The model has been validated through several approaches:

• mRNA expression of CreERT2 in F1A-CreERT2 mice matches the tissue-specific pattern of endogenous Fgf1A

• After tamoxifen administration, LacZ-positive signals are detected exclusively in the heart

• RFP-positive cells co-localize with cardiac troponin T (cTnT)-positive cardiomyocytes and FGF1-positive cells

How can FGF1 signaling be manipulated to study cardiomyocyte regeneration?

FGF1 has demonstrated potential in stimulating cardiomyocyte regeneration and improving cardiac function after myocardial infarction . To study this process:

Experimental Approaches:

• In vitro manipulation:

  • Treat neonatal cardiomyocytes with recombinant FGF1 to study cell cycle reentry through the FGFR1/Fn14 pathway

  • Use Cx43-knockdown models, which upregulate FGF1 expression in cardiomyocytes

• In vivo models:

  • Deliver recombinant FGF1 protein to the myocardium after induced infarction

  • Use the F1A-CreERT2 mouse model crossed with fluorescent reporters to trace endogenous Fgf1A-expressing cells during cardiac regeneration

  • Combine FGF1 with neurogenin1 to stimulate cardiomyocyte proliferation and facilitate cardiac remodeling after myocardial infarction

• Signaling pathway analysis:

  • Study the FGF1/FGFR/PKC signaling axis involved in cardiogenesis

  • Investigate autocrine vs. paracrine signaling, as FGF1 appears to function in an autocrine manner on cardiomyocytes but in a paracrine manner on fibroblasts and endothelial cells

What are the methodological considerations for differentiating between autocrine and paracrine effects of FGF1?

The search results indicate that FGF1 functions through distinct autocrine and paracrine mechanisms in different cell types . To differentiate between these signaling modes:

Methodological Approaches:

• Lineage tracing with F1A-CreERT2 mice:

  • Use F1A-CreERT2 mice crossed with reporter lines to identify cells that express FGF1

  • Compare reporter expression with FGF1 receptor (FGFR) expression patterns

  • Data suggests LacZ expression is detected in cardiomyocytes but not in fibroblasts or endothelial cells, indicating autocrine signaling in cardiomyocytes and paracrine signaling to other cell types

• Conditional knockout approaches:

  • Generate cell-type specific FGF1 knockouts using Cre-lox technology

  • Compare phenotypes when FGF1 is deleted from cardiomyocytes versus endothelial cells or fibroblasts

• Co-culture systems:

  • Establish co-cultures of cardiomyocytes with fibroblasts or endothelial cells

  • Use conditioned media experiments to isolate paracrine effects

  • Implement FGFR blockade in specific cell populations to determine signaling directionality

How can researchers investigate FGF1's role in metabolic regulation?

FGF1 has emerged as an important regulator of metabolic processes, particularly in the context of diabetes and obesity .

Research Approaches:

• Mouse models of metabolic disease:

  • Study Fgf1A expression in high-fat diet-induced obesity mouse models

  • Investigate FGF1 knockout mice, which show aggressive diabetic phenotypes when fed high-fat diets

  • Assess Fgf1A promoter activation in metabolic tissues using F1A-CreERT2 reporter mice

• Therapeutic administration studies:

  • Test subcutaneous FGF1 injection in mouse models of diabetes (ob/ob, db/db, diet-induced obesity)

  • Measure improvements in insulin sensitivity and reduction in serum glucose levels

  • Note that effects can be seen within 24 hours and last more than 48 hours

  • Consider intracerebroventricular FGF1 injection, which has been shown to reduce disease progression in rodent models of type 2 diabetes

• Mechanistic studies:

  • Investigate FGF1's role in ameliorating systemic inflammation in ob/ob mice through the FGFR1 pathway

  • Study how FGF1 functions as an insulin sensitizer similar to thiazolidinediones

  • Explore correlations between serum FGF1 levels and metabolic parameters in newly diagnosed type 2 diabetes patients

What are the neurobiological applications of FGF1 in mouse models?

FGF1 has significant neurobiological functions that can be studied in mouse models:

Research Applications:

• Neurodevelopmental studies:

  • Investigate FGF1's role in neuronal maturation and maintenance using culture experiments

  • Study the interaction between FGF1 and its receptors, particularly FGFR1, which is found mostly on neurons and neural stem cells

• Neuroplasticity research:

  • Examine FGF1's contribution to adult neuroplasticity

  • Investigate the relationship between FGF1 and FGFR1, which modulates proliferation of neural progenitor cells, neurogenesis, memory consolidation, and long-term potentiation

• Affective disorder models:

  • Explore FGF1 as a potential genetic predisposing factor for anxiety, depression, or substance abuse

  • Study how FGF1 may exert both developmental organizational effects and rapid "on-line" influences on behavior

  • Consider FGF1 as a candidate biomarker and treatment target for affective and addictive disorders

How should recombinant His-tagged FGF1 be used in experimental protocols?

When working with recombinant His-tagged FGF1 protein:

Best Practices:

• Quality control:

  • Verify protein purity (≥95% is standard for research applications)

  • Check endotoxin levels (≤0.005 EU/μg is appropriate for in vivo and cell culture experiments)

  • Validate protein identity via SDS-PAGE and/or mass spectrometry

• Experimental considerations:

  • For in vitro studies, use serum-free media to avoid interference from serum growth factors

  • Include heparin as a co-factor to stabilize FGF1 and enhance receptor binding

  • When studying signaling pathways, use phospho-specific antibodies to detect activation of FGFR and downstream targets

• Biological activity assessment:

  • Validate biological activity using established assays such as:

    • Proliferation of FGF-responsive cell lines

    • Phosphorylation of FGF receptors and downstream signaling molecules

    • Cardiomyocyte proliferation or cell cycle reentry assays

How can researchers reconcile the normal phenotype of FGF1 knockout mice with its numerous reported functions?

The paradox between FGF1 knockout mice showing no severe deficits and the numerous reported functions of FGF1 requires careful consideration:

Interpretative Framework:

• Functional redundancy:

  • Other FGF family members (23 in total) may compensate for FGF1 absence during development

  • FGF2, which shares many properties with FGF1, might provide functional backup

• Context-dependent functions:

  • FGF1's role may become apparent only under specific physiological challenges

  • Knockout mice should be examined under stress conditions (e.g., after myocardial infarction, with high-fat diet challenges, or during aging)

• Methodological considerations:

  • Use more sensitive assays to detect subtle phenotypes at molecular and cellular levels

  • Employ conditional and/or inducible knockout models to bypass developmental compensation

• Translational implications:

  • The lack of severe phenotype in knockout mice suggests that FGF1-targeted therapeutics might have favorable safety profiles

  • Focus on gain-of-function rather than loss-of-function approaches in translational research

What explains the contradictory findings regarding FGF1's effects across different tissues?

The search results reveal some contradictory findings about FGF1's roles, particularly in different tissues:

Analytical Approach:

• Tissue-specific promoter usage:

  • The Fgf1 gene contains three promoters (1A, 1B, 1G) with tissue-specific expression patterns

  • Studies should specify which FGF1 transcript variant they are investigating

• Receptor distribution:

  • Different tissues express different complements of FGF receptors

  • FGFR1 is predominantly neuronal while FGFR2 shows primarily glial expression

  • The receptor profile determines the cellular response to FGF1

• Experimental variables:

  • Different concentrations of FGF1 may activate different signaling pathways

  • The presence or absence of co-factors (especially heparin) significantly affects FGF1 activity

  • The developmental stage at which FGF1 is administered or manipulated matters significantly

What are the optimal conditions for using the F1A-CreERT2 mouse model in lineage tracing experiments?

The F1A-CreERT2 mouse model enables precise temporal control over the labeling of Fgf1A-expressing cells . For optimal results:

Protocol Considerations:

• Tamoxifen administration:

  • Dosage: Typically 75-100 mg/kg body weight

  • Route: Intraperitoneal injection is common, but oral gavage is an alternative

  • Schedule: Single high dose or multiple lower doses over 3-5 days

  • Timing: Consider the developmental stage or disease progression point

• Reporter selection:

  • ROSA26 reporters are commonly used for their ubiquitous expression

  • Consider fluorescent reporters (e.g., tdTomato) for live imaging

  • LacZ reporters offer sensitivity for fixed tissue analysis

• Control experiments:

  • Include oil-injected F1A-CreERT2;Reporter mice as negative controls

  • Use ubiquitous CreERT2 lines as positive controls

  • Test for tamoxifen-independent recombination ("leakiness")

• Analysis methods:

  • For cardiac studies, co-staining with cTnT confirms cardiomyocyte identity of labeled cells

  • RFP-positive cells can be directly observed under fluorescence microscopy without immunostaining

  • Precise tissue processing is critical - detailed protocols are available in the literature

How should researchers prepare and validate His-tagged FGF1 for functional studies?

Working with His-tagged FGF1 requires careful preparation and validation:

Laboratory Protocol:

• Expression and purification:

  • Expression systems: HEK293 cells provide mammalian post-translational modifications

  • Purification: Use nickel or cobalt affinity chromatography followed by size exclusion

  • Quality control: Verify purity by SDS-PAGE (≥95% purity is standard)

• Functional validation:

  • Bioactivity assays: Proliferation of FGF-responsive cell lines

  • Receptor activation: Phosphorylation of FGFRs and downstream effectors

  • Heparin binding: Affinity chromatography using heparin columns

• Storage and handling:

  • Aliquot to avoid freeze-thaw cycles

  • Add carrier protein (e.g., BSA) for dilute solutions

  • Include heparin as a stabilizing co-factor

  • Store at -80°C for long-term or -20°C for short-term use