Skip to content
Brilliant Blue Biosciences logoBrilliant BlueBiosciences
Acta Biomaterialia2019ResearchNon-viral Gene Delivery

Injectable PLGA microspheres with tunable magnesium ions release for promoting bone regeneration

Zuoying Yuan, Pengfei Wei, Yiqian Huang, Wenxin Zhang, Fuyu Chen, Xu Zhang, Jianping Mao, Dafu Chen, Qing Cai, Xiaoping YangDOI 10.1016/j.actbio.2018.12.017

Summary

Magnesium ions (Mg²⁺) promote bone regeneration, but their effects are strongly concentration-dependent: too little has minimal effect, while excess Mg²⁺ can impair osteoblast activity and bone metabolism. There is no well-established bone tissue engineering scaffold that precisely controls local Mg²⁺ release. Injectable, biodegradable microspheres that provide tunable Mg²⁺ release could fill irregular bone defects and improve bone regeneration. Tunable Mg²⁺ release: Higher MgO fractions gave faster release and higher initial Mg²⁺ concentrations; higher MgCO₃ fractions gave slower, longer-term sustained release. PMg-III released approximately 30–50 ppm Mg²⁺ in.

Keywords

PLGAPolymericNanoparticlesNanocarriersGene deliveryDrug deliveryImmune cells
Purpose: Magnesium ions (Mg²⁺) promote bone regeneration, but their effects are strongly concentration-dependent: too little has minimal effect, while excess Mg²⁺ can impair osteoblast activity and bone metabolism. There is no well-established bone tissue engineering scaffold that precisely controls local Mg²⁺ release. Injectable, biodegradable microspheres that provide tunable Mg²⁺ release could fill irregular bone defects and improve bone regeneration.
Hypothesis: If fast-degrading MgO and slow-dissolving MgCO₃ are co-embedded into PLGA microspheres at different weight ratios, then the Mg²⁺ release rate and duration can be precisely tuned; microspheres with a proper sustained Mg²⁺ release profile will promote bone marrow mesenchymal stromal cell (BMSC) migration, proliferation, and osteogenic differentiation, and enhance in vivo bone regeneration in a critical-sized calvarial defect model.
Aims: Primary aim: Prepare injectable PLGA microspheres co-embedded with MgO and MgCO₃ at a fixed total loading of 20 wt.% and different MgO:MgCO₃ ratios (1:0, 3:1, 1:1, 1:3, 0:1).
  • Secondary aim 1: Characterize microsphere morphology, size, Mg loading, hydrophilicity, degradation, pH changes, and Mg²⁺ release kinetics.
  • Secondary aim 2: Evaluate in vitro bioactivity — biomineralization, antibacterial activity against S. aureus, BMSC viability, attachment, migration, and osteogenic differentiation.
  • Secondary aim 3: Assess in vivo bone regeneration in a critical-sized rat calvarial defect (8 mm) using the optimized PMg-III microspheres compared with PLGA microspheres and an unfilled control.
Delivery system:

Component: Polymer; Description: PLGA (lactide:glycolide = 75:25, MW ≈ 50,000)

Component: Microsphere Type; Description: Injectable bioresorbable microspheres prepared by solid-in-oil-in-water (S/O/W) emulsion

Component: Payload; Description: MgO nanoparticles (~20 nm) and MgCO₃ microparticles (~15 µm) co-embedded at total 20 wt.%

Component: Ratios Tested; Description: MgO:MgCO₃ = 1:0 (PMg-I), 3:1 (PMg-II), 1:1 (PMg-III), 1:3 (PMg-IV), 0:1 (PMg-V)

Component: Targeting Ligand; Description: None

Component: Size; Description: PMg-I: 190.9 ± 40.4 µm; PMg-V: 98.2 ± 22.5 µm; PLGA: 84.3 ± 27.3 µm

Component: Surface; Description: Rougher than PLGA; pits from embedded inorganic powders

Component: Key Design Feature; Description: Tuning MgO/MgCO₃ ratio controls Mg²⁺ release: more MgO → faster, higher initial release; more MgCO₃ → slower, longer sustained release

Component: Optimized Formulation; Description: PMg-III (MgO:MgCO₃ = 1:1) selected for in vivo bone regeneration

Approach: In Vitro: - Cells: Sprague-Dawley rat BMSCs. - Culture modes: Transwell non-contact co-culture and direct contact seeding on microspheres. - Osteogenic induction: 21 days with ascorbic acid, β-sodium glycerophosphate, and dexamethasone. - Antibacterial: S. aureus live/dead staining after 24 h incubation with microspheres. - Biomineralization: Microspheres soaked in 1.5× simulated body fluid (1.5SBF) for 7 days. - Migration: Boyden chamber assay with microspheres in lower chamber.

In Vivo: - Model: Female SD rats (5–6 weeks old), critical-sized calvarial defect (φ = 8 mm). - Groups (n = 10/group): (1) unfilled control, (2) PLGA microspheres, (3) PMg-III microspheres. - Time points: 4, 8, and 16 weeks post-operation (3, 3, and 4 rats per group sacrificed at each time point, respectively). - No rat died or was excluded.

Key methods:

Technique: SEM and elemental mapping; Purpose: Microsphere morphology, size, Mg distribution

Technique: Thermogravimetric analysis (TGA); Purpose: Quantify inorganic loading and residual MgO

Technique: Water contact angle; Purpose: Assess surface hydrophilicity

Technique: ICP-OES; Purpose: Quantify Mg²⁺ release in PBS over 28 days

Technique: pH measurements; Purpose: Monitor medium pH changes during degradation

Technique: CCK-8, live/dead, phalloidin/Hoechst, SEM; Purpose: BMSC viability, attachment, spreading, and morphology

Technique: Boyden chamber; Purpose: BMSC migration

Technique: ALP activity, Col-I ELISA, alizarin red, qPCR; Purpose: Osteogenic differentiation markers (ALP, OPN, OCN, Col-I)

Technique: S. aureus live/dead staining; Purpose: Antibacterial activity

Technique: Micro-CT; Purpose: Bone volume fraction (BV/TV) and bone mineral density (BMD)

Technique: H&E, Masson’s trichrome, immunohistochemistry; Purpose: Histological bone regeneration, OPN and OCN expression

Key results: Tunable Mg²⁺ release: Higher MgO fractions gave faster release and higher initial Mg²⁺ concentrations; higher MgCO₃ fractions gave slower, longer-term sustained release. PMg-III released approximately 30–50 ppm Mg²⁺ in vitro and maintained medium pH near 7. - PMg-III best promoted osteogenic differentiation: Among PMg-I, PMg-III, and PMg-V, PMg-III showed the strongest upregulation of ALP, OPN, OCN, and Col-I genes/proteins and the richest alizarin red calcium deposition. - Migration: BMSC migration increased in the order PMg-I > PMg-III > PMg-V, correlating with Mg²⁺ release. - Antibacterial activity: PMg-I (highest MgO) reduced viable S. aureus to 35.8 ± 3.8%; PMg-III had inferior antibacterial activity. - In vivo bone regeneration at 16 weeks: PMg-III achieved BV/TV = 32.9 ± 5.6% and BMD = 325.7 ± 20.2 mg/cm³, significantly higher than PLGA (BV/TV 8.1 ± 2.5%; BMD 124 ± 35.8 mg/cm³) and control (BV/TV 4.6 ± 0.7%; BMD 45.7 ± 23.2 mg/cm³). New bone, collagen-rich matrix, vascular-like structures, and strong OPN/OCN expression were observed in the PMg-III group.
Interpretation: The authors conclude that co-embedding fast-degrading MgO and slow-dissolving MgCO₃ in PLGA microspheres enables precise tuning of Mg²⁺ release. The optimized PMg-III formulation, with sustained moderate Mg²⁺ release, significantly promotes BMSC migration, osteogenic differentiation, and in vivo bone regeneration. This injectable microsphere platform is proposed as a promising strategy for bone tissue engineering and a valuable guideline for controlling bioactive metal ion release in scaffold design.
Limitations: Only PMg-III was evaluated in vivo; the full range of MgO/MgCO₃ ratios was not tested in the animal model. - The study used a rat calvarial defect model only; no large-animal or load-bearing bone defect validation. - The optimal total MgO/MgCO₃ loading and optimal polymer degradation rate were not determined. - Local in vivo Mg²⁺ concentration could not be measured and may differ from in vitro estimates. - Long-term safety, immune response, and degradation by-product effects were not fully assessed. - Antibacterial testing was limited to S. aureus. - No direct comparison with clinically used bone graft materials or growth-factor-based therapies.

Related articles

Biomaterials Science

Approaches towards Biomaterial-Mediated Gene Editing for Cancer Immunotherapy

Theranostics

Bioengineering of Artificial Antigen Presenting Cells and Lymphoid Organs

Pharmaceutical Research

Cancer Immunotherapy and Nanomedicine

← Back to all articles

Let's engineer the next delivery breakthrough together

We co-develop nanocarrier and biosensing programs with pharma, biotech and academic groups — from target selection through GMP supply.

Start a conversationExplore the pipeline
Injectable PLGA microspheres with tunable magnesium ions release for promoting bone regeneration | Brilliant Blue Biosciences