Overview

Also Known As:Flnb KO

These Flnb knockout mice exhibit abnormal bone development with vertebral fusion and reduced ossification in long bones. This strain may be useful in studies of skeletal development and skeletal dysplasias such as spondylocarpaltarsal synostosis syndrome.

Donating Investigator

Volney L. Sheen, Beth Israel Deaconess Medical Center

Genetic overview

Genetic Background	Generation

Flnb^tm1Vshn

Allele Type	Gene Symbol	Gene Name
Targeted (Null/Knockout)	Flnb	filamin, beta

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Research Applications

Internal/Organ Research
Developmental Biology Research

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Base Price

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$2,854.50 Domestic price Cryo Recovery

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Details

Detailed Description

Filamin beta stabilizes 3D actin filament networks, connecting the cell membrane to the actin cytoskeleton. It has roles in regulating intracellular signaling, cell motility, organ development, tumor growth and metastasis. Mutations of FLNB are the cause of a group of human skeletal dysplasia disorders including spondylocarpotarsal syndrome, boomerang dysplasia, Larsen syndrome and atelosteogenesis I and III. These mice carry a knock out mutation for the Flnb gene in which exons 3 through 5 are replaced by a NEO cassette. Mice that are homozygous for the targeted mutation have a high death rate around the time of birth. Adult male homozygotes are fertile but exhibit mating problems due to deformity. Some surviving homozygous females exhibit dystocia. Heterozygotes are viable and fertile.

No gene product (protein) is detected by Western blot analysis of liver from 7 day old homozygotes and growth plate of the radius from E16.5 homozygotes. Surviving male homozygotes exhibit a small body size, lower body weight, shortened distal limbs, rib and vertebrae fusion, abnormal spinal curvatures, and dysmorphic facial/calvarial bones. Female homozygotes display a small body size, vertebral fusions and joint laxity. Homozygotes have a delay in endochondral bone development, reduced bone ossification, and disrupted extracellular matrix organization. Chondrocyte differentiation in long bones is delayed while proliferation is disrupted.

Development

A targeting vector containing a NEO cassette was used to disrupt exons 3 through 5 of the targeted gene. The construct was electroporated into 129S6/SvEvTac derived iTL1 embryonic stem (ES) cells. Correctly targeted ES cells were injected into C57BL/6J blastocysts. The resulting chimeric animals were tested for germline transmission.
The mice were then crossed to C57BL/6 and/or FVB.
Upon arrival at The Jackson Laboratory, the mice were crossed to C57BL/6J (Stock No. 000664) at least once to establish the colony.

Control Suggestions

Wild-type from the colony

Additional Information

Considerations for Choosing Controls

Selected References

Lu J; Lian G; Lenkinski R; De Grand A; Vaid RR; Bryce T; Stasenko M; Boskey A; Walsh C; Sheen V. 2007. Filamin B mutations cause chondrocyte defects in skeletal development. Hum Mol Genet 16(14):1661-75PubMed: 17510210 MGI: J:125095

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Genetics

Flnb^tm1Vshn

Allele Symbol: Flnb^tm1Vshn

Allele Name	targeted mutation 1, Volney Sheen
Allele Type	Targeted (Null/Knockout)
Allele Synonym(s)	Flnb^-
Gene Symbol and Name	Flnb, filamin, beta
Gene Synonym(s)
Strain of Origin	129S6/SvEvTac
Chromosome	14
Molecular Note	Exons 3-5 were replaced by a neo selection cassette causing a frame shift mutation. Western blot analysis of P7 livers confirmed graded loss of protein expression in heterozygous and homozygous mutant mice.

Disease/Phenotype

Disease Terms

Research Areas By Phenotype

This mouse can be used to support research in many areas including:

Internal/Organ Research
- Skeleton
  - Bone
Developmental Biology Research
- Skeletal Defects
- Growth Defects
  - Growth Defects (homozygous)

Mammalian Phenotype Terms by Genotype

The following phenotype information is associated with a similar, but not exact match to this JAX^® Mice strain

Genotype: Flnb^tm1Vshn/Flnb^tm1Vshn
involves: 129S6/SvEvTac * C57BL/6

cellular phenotype

abnormal cell adhesion
- inhibition of beta1-integrin in these cells leads to further impairment in cell spreading
- mutant chondrocytes exhibit decreased adhesion to ECM substrates

abnormal extracellular matrix morphology
- at E16.5, electron micrographs of the matrix compartments of mutant growth plates revealed increased collagen fibril density relative to wild-type controls, suggesting a disruption of the ECM

limbs/digits/tail phenotype

abnormal limb bone morphology
- limb bones are poorly calcified

abnormal patella morphology
- patellar bones are poorly calcified

abnormal radius morphology
- an increase in apoptosis is noted in the hypertrophic (H) zone of E16.5 radius, as shown by TUNEL analysis; increased rates of cell death occur along the periphery of the radius bones between E14.5 and E17.5, with dying cells restricted primarily to the perichondral regions, adjacent to the H zone
- erichondral regions, adjacent to the H zone
- the radius appears less calcified and more fragile, as shown by von Kossa staining

abnormal ulna morphology
- the ulna appears less calcified and more fragile, as shown by von Kossa staining

brachyphalangia
- the phalanx (fore claw) is significantly shorter at E16.5 and at 2 months

decreased diameter of radius
- the radius is thinned

decreased diameter of ulna
- the ulna is thinned

short limbs
- shortened distal limbs

short radius
- radius length is reduced by ~1.4 mm at P1 and up to 4 mm by 8 weeks
- the radius is significantly shorter at E16.5 and at 2 months

mortality/aging

preweaning lethality, incomplete penetrance
- only 7.7% of homozygotes are obtained within the first week of life, suggesting embryonic or early postnatal lethality

skeleton phenotype

abnormal chondrocyte morphology
- a delay in chondrocyte progression and differentiation is seen in the H zone at 12 hr after the BrdU pulse, coinciding with a decrease in the length of the H zone of the E16.5 radius
- early defect affecting hypertrophic chondrocyte differentiation within the more distal rather than proximal appendages
- homozygotes show a progressive decline in the number of rapidly proliferating chondrocytes and premature differentiation characterized by an enlarged prehypertrophic zone, a widened Col2a1+/Col10a1+ (proliferative/hypertrophic) overlapping region, but a relatively reduced hypertrophic zone length (seen at P7 and at 2 weeks)
- the greatest differences in the length of the H zone occur at E14.5 and E15.5, suggesting a delay in chondrocyte maturation and hypertrophy
- the size of the prehypertrophic zone is variable and often blurred, as shown by Indian hedgehog staining

abnormal chondrocyte physiology
- at baseline, cultured mutant chondrocytes exhibit reduced cell spreading relative to wild-type chondrocytes
- at E16.5, fewer phospho-beta1-integrin (Ser785) positive chondrocytes are observed in the mutant radius, indicating reduced chondrocyte cell adhesion
- decreased adhesion is further reduced by dominant negative beta1-integrin transfection relative to either mutant chondrocytes or wild-type chondrocytes transfected with the dominant negative beta1-integrin alone, indicating disruption of the ECM-integrin pathway through loss of cell adhesion
- in vitro, the % of proliferating chondrocytes remaining in G1/G0 (BrdU+, Ki67-/low) is increased by ~36%, relative to wild-type controls
- in vivo, co-staining with Ki67 and BrdU revealed that the % of proliferating chondrocytes remaining in the G1/G0 phase (BrdU+, Ki67-/low) in the radius is increased by ~10% and ~13% at E16.5 and P7, respectively, relative to wild-type controls, indicating an increased number of actively proliferating chondrocytes adopting a more differentiated state
- mutant chondrocytes display decreased adhesion to several extracellular substrates including collagen and fibronectin, and show reduced binding to other ECM components (i.e. type IV collagen, vitronectin and laminin)
- pathway through loss of cell adhesion

abnormal limb bone morphology
- limb bones are poorly calcified

abnormal long bone epiphyseal plate morphology
- at P1 and 2 weeks old age, but not at P7, the length of Col2a1+/Col10a1+ overlapping expression relative to the radius growth plate length is increased relative to that in wild-type controls (17.7% vs 8.6% at P1 and 24.2% vs 19.1% at 2 weeks, respectively)
- at P1, P7 and 2 weeks, the ratio of Pthr1+ length to growth plate length is increased relative to that in wild-type controls (51.7% vs 39.2% at P1, 38.7% vs 23.0% at P7, and 38.6% vs 28.1% at 2 weeks, respectively)
- at P7 and 2 weeks, the ratio of Ihh+ length to growth plate length is also increased (34.2% vs 19.6% at P7, and 37.0% vs 27.7% at 2 weeks)
- at P7 to 2 weeks age, but not at P1, the ratio of Col10a1+ length relative to Col2a1+ length (hypertrophic to proliferative zone ratio), is decreased in the radius growth plate relative to that in wild-type controls (33.8% vs 45.3% at P7 and 71.9% vs 89.6% at 2 weeks, respectively), suggesting a delay in skeletogenesis
- immunostaining of the growth plates with Pthr1 and Ihh (intermediate differentiation/prehypertrophic zone markers) revealed an increase in the prehypertrophic zone

abnormal long bone epiphyseal plate proliferative zone
- a slight decrease in the length and width of the proliferative (P) zone is observed by collagen II staining
- at E14.5, E16.5 and P7, the % of cells positive for proliferation markers Sox9, BrdU (cells in S-phase), Ki67, and PH3 (cells in M-phase) is progressively decreased within the proliferative zone of the radius growth plate
- at P7, fewer Sox9+ labeled chondrocytes (i.e. less differentiated chondrocytes) are noted in the rapid proliferative zone and prehypertrophic zone relative to wild-type controls (47.8% vs 81.4%, respectively)
- at P7, signal intensity for Runx2 (a chondrocyte marker that is up-regulated during both endochondral and intramembranous ossification) is increased within chondrocyte progenitors in the proliferative zone relative to wild-type controls (32.6 vs 28.9 luminosity, respectively)
- Normal - however, no increase in TUNEL staining is noted within the proliferative zone from E14.5 to P7
- while proliferation in the P zone of E16.5 radius is largely unaffected (1 h post BrdU injection), the distribution of proliferating chondrocyte progenitors incorporating BrdU is shifted toward the resting zone (more distal to the H zone)

abnormal neurocranium morphology
- dysmorphic calvaria

abnormal occipital bone morphology
- occipital bones are poorly calcified

abnormal parietal bone morphology
- parietal bones are poorly calcified

abnormal patella morphology
- patellar bones are poorly calcified

abnormal perichondrium morphology
- increased apoptosis along the periphery of the hypetrophic (H) zone of the radius at E16.5; apoptotic cells reside within the perichondrium, adjacent to the H zone
- increased cell death is noted along the bone collar and is consistently seen through all skeletal developmental ages

abnormal radius morphology
- an increase in apoptosis is noted in the hypertrophic (H) zone of E16.5 radius, as shown by TUNEL analysis; increased rates of cell death occur along the periphery of the radius bones between E14.5 and E17.5, with dying cells restricted primarily to the perichondral regions, adjacent to the H zone
- erichondral regions, adjacent to the H zone
- the radius appears less calcified and more fragile, as shown by von Kossa staining

abnormal skeleton development
- at E18.5, homozygotes display hypoplastic cartilaginous skeletons
- overall findings are consistent with a delay in skeletal development

abnormal thoracic cage morphology
- thoracic cages show increased radiolucency suggestive of osteopenia

abnormal ulna morphology
- the ulna appears less calcified and more fragile, as shown by von Kossa staining

abnormal viscerocranium morphology
- dysmorphic facial bones

brachyphalangia
- the phalanx (fore claw) is significantly shorter at E16.5 and at 2 months

cervical vertebral fusion
- fusion of cervical spinal vertebrae in some mice

decreased bone mineral density
- bone densitometry measurements of E20, P7 and adult vertebral segments revealed a significant reduction in bone density
- micro-CT scans of mutant skeletons revealed a generalized mottled osteopenia at various developmental ages

decreased bone mineralization
- micro-CT scans revealed demineralization of the mutant skeleton at various developmental ages (E20, P3, P13, P20 and adult)
- several of the trabecular and cortical bones (calvaria, patella and appendicular limbs) appear to be poorly calcified, as evidenced by the reduced radiolucency on CT scans
- whole skeleton preparations of E18.5 mutant embryos, stained with Alizarin red (bone) and Alcian blue (cartilage), show a decrease in bone mineralization

decreased bone ossification
- bone densitometry measurements of the vertebral segments revealed a significant reduction in bone ossification

decreased diameter of radius
- the radius is thinned

decreased diameter of ulna
- the ulna is thinned

decreased length of long bones
- at P1 to 8 weeks, appendicular long bones are shortened

decreased long bone epiphyseal plate size
- at P1 and P7, the length of the growth plate in the radius is significantly shorter than that in wild-type controls
- H&E staining of E16.5 distal bone appendages revealed that mutant growth plates are significantly smaller than wild-type, as shown in the ulna, radius and phalanx
- Normal - however, no differences in chondrocyte size and shape, chondrocyte rotation and column integrity are seen
- the greatest reduction is observed in the more distal radius, ulna and digital bones, as opposed to the more proximal humerus

decreased width of hypertrophic chondrocyte zone
- an increase in apoptosis is noted in the H zone of E16.5 radius, as shown by TUNEL analysis; increased rates of cell death occur along the periphery of the radius bones between E14.5 and E17.5, with dying cells restricted primarily to the perichondral regions, adjacent to the H zone
- collagen X staining confirmed a clear reduction in the length and width of the H zone
- ions, adjacent to the H zone
- the greatest differences in the length of the H zone occur at E14.5 and E15.5, suggesting a delay in chondrocyte maturation and hypertrophy
- the hypertrophic (H) zone is significantly shortened in length in both the radius and phalanx

delayed bone ossification

delayed endochondral bone ossification
- delayed endochondral ossification at E18.5

delayed intramembranous bone ossification
- delayed intramembranous ossification at E18.5

increased width of hypertrophic chondrocyte zone
- at P1, the relative thickness of the Col10a1+ hypertrophic zone (Col10a1+/whole growth plate ratio) is increased by 10% relative to that in wild-type controls (38.7% vs 28.5%, respectively), suggesting premature hypertrophic differentiation
- Normal - however, no significant change in the Col10a1+/whole growth plate ratio is noted at P7 and at 2 weeks, suggesting a slowing of the early maturation process over time

joint laxity
- female homozygotes display joint laxity

kyphosis
- exaggerated thoracolumbar kyphosis in some mice

lordosis
- exaggerated lower thoracic lordosis in some mice

rib fusion
- abnormal fusion of the proximal ribs in some mice

scoliosis

short radius
- radius length is reduced by ~1.4 mm at P1 and up to 4 mm by 8 weeks
- the radius is significantly shorter at E16.5 and at 2 months

small thoracic cage

thin ribs
- ribs appear thin and gracile

vertebral fusion
- female homozygotes display vertebral fusions

craniofacial phenotype

abnormal neurocranium morphology
- dysmorphic calvaria

abnormal occipital bone morphology
- occipital bones are poorly calcified

abnormal parietal bone morphology
- parietal bones are poorly calcified

abnormal viscerocranium morphology
- dysmorphic facial bones

growth/size/body region phenotype

postnatal growth retardation
- Normal - in contrast, female homozygotes show absence of clear growth and size differences
- male homozygotes show a significant decrease in postnatal growth rate relative to wild-type controls

slow postnatal weight gain
- male, but not female, homozygotes never attain the size of adult wild-type controls by 8 weeks of age

References

Lu J; Lian G; Lenkinski R; De Grand A; Vaid RR; Bryce T; Stasenko M; Boskey A; Walsh C; Sheen V. 2007. Filamin B mutations cause chondrocyte defects in skeletal development. Hum Mol Genet 16(14):1661-75PubMed: 17510210 MGI: J:125095

Additional - Flnb^tm1Vshn related

Technical Support

CONTACT TECHNICAL SUPPORT

Genotyping Protocols
- Standard PCR:Flnb-Alternate 4
- Standard PCR:Generic Neo
- Probe:Generic Neo
- Genotyping resources and troubleshooting
Breeding Considerations

When maintaining a live colony, heterozygous mice may be bred together, to wildtype siblings, or to C57BL/6J inbred mice (Stock No. 000664). Most homozygotes die perinatally. Surviving homozygous females exhibit dystocia and surviving homozygous males exhibit mating problems due to deformity.
- Additional Breeding and Husbandry Support
Citation
When using the Flnb KO mouse strain in a publication, please cite the originating article(s) and include JAX stock #027233 in your Materials and Methods section.

Animal Health Reports

Facility Barrier Level Descriptions

Production of mice from cryopreserved embryos or sperm occurs in a maximum barrier room, G200

Pricing & Availability

Cryo Recovery

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Live Mouse

Age

Genotype

Price

weeks

Cryorecovery - Pricing

Service/Product	Description	Price
	Heterozygous or wildtype for Flnb<tm1Vshn>

Related Products and Services

Frozen Mouse Embryo	STOCK Flnb<tm1Vshn>/J Frozen Embryo	$2595.00
Frozen Mouse Embryo	STOCK Flnb<tm1Vshn>/J Frozen Embryo	$2595.00
Frozen Mouse Embryo	STOCK Flnb<tm1Vshn>/J Frozen Embryo	$3373.50
Frozen Mouse Embryo	STOCK Flnb<tm1Vshn>/J Frozen Embryo	$3373.50

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The Jackson Laboratory has rigorous genetic quality control and mutant gene genotyping programs to ensure the genetic background of JAX® Mice strains as well as the genotypes of strains with identified molecular mutations. JAX® Mice strains are only made available to researchers after meeting our standards. However, the phenotype of each strain may not be fully characterized and/or captured in the strain data sheets. Therefore, we cannot guarantee a strain's phenotype will meet all expectations. To ensure that JAX® Mice will meet the needs of individual research projects or when requesting a strain that is new to your research, we suggest ordering and performing tests on a small number of mice to determine suitability for your particular project. We do not guarantee breeding performance and therefore suggest that investigators order more than one breeding pair to avoid delays in their research.

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Also Known As:Flnb KO

Donating Investigator

Genetic overview

Research Applications

Base Price

Detailed Description

Development

Control Suggestions

Additional Information

Selected References

Flnbtm1Vshn

Allele Symbol: Flnbtm1Vshn

Disease Terms

Research Areas By Phenotype

This mouse can be used to support research in many areas including:

Mammalian Phenotype Terms by Genotype

Genotype: Flnbtm1Vshn/Flnbtm1Vshninvolves: 129S6/SvEvTac * C57BL/6

cellular phenotype

limbs/digits/tail phenotype

mortality/aging

skeleton phenotype

craniofacial phenotype

growth/size/body region phenotype

References

Additional - Flnbtm1Vshn related

Genotyping Protocols

Breeding Considerations

Citation

Animal Health Reports

Production of mice from cryopreserved embryos or sperm occurs in a maximum barrier room, G200

Live Mouse

Cryorecovery - Pricing

Related Products and Services

Payment Terms and Conditions

The Jackson Laboratory's Genotype Promise

Terms Of Use

Additional Use Restrictions Apply

Licensing Information

JAX® Mice, Products & Services Conditions of Use

No Warranty

Credit for PRODUCTS or SERVICES

Animal Care and Use for SERVICES

No Liability

Flnb^tm1Vshn

Allele Symbol: Flnb^tm1Vshn

Genotype: Flnb^tm1Vshn/Flnb^tm1Vshn
involves: 129S6/SvEvTac * C57BL/6

Additional - Flnb^tm1Vshn related