MLO-Y4 Cell Line

Brief Introduction
Product List

General Information
Organism	Mus musculus, mouse
Cell Line Description	MLO-Y4 is a transformed murine long-bone osteocyte-like cell line established from cells isolated by sequential collagenase digestion of long bones from transgenic mice expressing SV40 large T antigen under control of the osteocalcin promoter. The cells extend complex dendritic processes, form functional gap junctions, produce abundant osteocalcin and connexin 43, and respond to fluid flow and mechanical strain. MLO-Y4 supports studies of osteocyte mechanotransduction, dendrite biology, intercellular communication, survival, prostaglandin signaling, osteoclast regulation, and extracellular-matrix-dependent gene expression. The model represents a proliferative osteocyte-like state rather than a fully mature primary osteocyte, particularly because standard two-dimensional culture produces little or no physiologically relevant sclerostin.
Cell Type	Murine osteocyte-like cell
Tissue of Origin	Long bone
Disease	No associated disease; derived from transgenic mouse bone
Mouse Background	(C57BL/6J × BALB/cJ) F2 transgenic background
Sex	Not specified
Age at Sampling	14 days
Immortalization	SV40 large T antigen expressed from the osteocalcin promoter in the source transgenic mice
Morphology	Stellate osteocyte-like cells with extensive, complex dendritic processes
Growth Mode	Adherent; collagen-coated culture surfaces support attachment, maintenance, and growth
Applications	1. Osteocyte biology and gene-expression studies 2. Fluid-flow, substrate-stretching, compression, and mechanical-loading experiments 3. Osteocyte mechanotransduction and prostaglandin signaling 4. Gap-junction communication and connexin 43 biology 5. Dendrite formation, elongation, and E11/gp38 function 6. Osteocyte survival, apoptosis, and stress-response studies 7. Osteocyte–osteoblast and osteocyte–osteoclast communication 8. Osteoclast formation and activation assays 9. RANKL/OPG and bone-remodeling research 10. Two-dimensional and three-dimensional extracellular-matrix studies 11. Sost/sclerostin regulation under mineralized or hydroxyapatite-containing conditions 12. Bone metastasis and multicellular co-culture models
Characteristics
Transformation Status	Transformed cell line with integrated SV40 large T antigen originating from an osteocalcin-promoter-driven transgenic mouse
Dendritic Phenotype	Produces extensive, complex dendritic processes characteristic of osteocyte-like morphology
Osteocalcin Expression	High osteocalcin production relative to primary osteoblasts
Alkaline Phosphatase	Low alkaline-phosphatase production relative to primary osteoblasts
Type I Collagen	Very low type I collagen production and mRNA abundance relative to primary osteoblasts
Connexin 43 and Gap Junctions	High connexin 43 production with functional gap junctions that communicate through dendritic processes
Osteopontin and CD44	Positive for osteopontin and CD44; abundance is similar to the osteoblast comparison used in the original characterization
Mechanical Response	Fluid flow increases prostaglandin production and functional gap-junction activity. Mechanical strain regulates E11/gp38 and dendrite elongation, while three-dimensional loading activates mechanosensitive transcriptional programs.
Osteoclast-Regulatory Activity	Supports osteoclast formation and activation in co-culture systems
Sost/Sclerostin Limitation	Standard two-dimensional culture on collagen-coated plastic produces little or no physiologically relevant Sost/sclerostin. Hydroxyapatite-containing two-dimensional matrices induce Sost, and three-dimensional collagen–hydroxyapatite scaffolds further increase expression toward physiological levels.
Mycoplasma Test	Negative
Culture Conditions and Handling
Culture Conditions	37°C in a humidified atmosphere containing 5% CO₂
Dissociation Reagent	Trypsin–EDTA
Subculturing	1. Prepare fresh type I collagen-coated culture vessels before passaging. 2. Remove the spent medium and rinse the adherent cells with a suitable balanced salt solution. 3. Add enough trypsin–EDTA to cover the monolayer. 4. Monitor detachment microscopically and stop the reaction promptly with serum-containing complete medium. 5. Gently disperse the cells without excessive mechanical stress. 6. Seed the suspension onto collagen-coated vessels in complete α-MEM. 7. Return the cultures to 37°C and 5% CO₂ and monitor attachment, dendritic morphology, and growth.
Extracellular-Matrix Control	Use the same collagen source, coating concentration, coating time, and surface format within an experiment. Matrix composition and dimensionality directly influence metabolic activity, dendritic phenotype, and Sost expression.
Mechanical-Stimulation Studies	Standardize cell density, confluence, substrate stiffness, coating, loading waveform, frequency, magnitude, duration, and post-loading collection time. Include static controls cultured on an equivalent substrate.
Three-Dimensional Culture Note	Three-dimensional collagen-based systems can provide a more physiologically relevant environment than collagen-coated plastic. Inclusion of hydroxyapatite can restore Sost expression and modify the response to fluid flow.

Cat.No.	Product Name	Price
CSC-RO01206	Cas9 Stable Cell Line - MLO-Y4	Inquiry
CSC-RR00698	Luciferase Reporter Cell Line - MLO-Y4	Inquiry

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* For research use only. Not intended for any clinical use.

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