Research Article

Effect of nano‑hydroxyapatite on protein adsorption and cell adhesion

of poly(lactic acid)/nano‑hydroxyapatite composite microspheres
H. M. C. Suboda Wijerathne1,2 · Dong Yan1 · Bin Zeng1 · Yanping Xie1 · Hongchao Hu1 · M. Nirmali Wickramaratne2 ·
Yingchao Han1 

Received: 18 January 2020 / Accepted: 14 March 2020 / Published online: 20 March 2020
© Springer Nature Switzerland AG 2020

Abstract
Surface properties of biomaterials are significant as they are intended to interact with biological system interfaces. This
study was aimed to reveal the effect of nano-hydroxyapatite (Nano-HAP) on the properties of poly(lactic acid) (PLA)
microspheres. Nano-HAP particles of 10–30% content were successfully embedded in PLA microspheres by emulsion
solvent evaporation method. The incorporation of Nano-HAP particles resulted in the increases of surface hydrophilicity
and surface rough of microspheres that depended on the content of Nano-HAP. With the increase of Nano-HAP content,
the adsorption capacity for protein of PLA/Nano-HAP composite microspheres was increased significantly. Moreover,
the incorporation of Nano-HAP could promote the adhesion and proliferation of rat Mesenchymal Stem cells (rMSCs)
on microspheres, which could be attributed to the component of Nano-HAP as well as the increases of surface hydro-
philicity and rough. In addition, PLA/Nano-HAP composite microspheres showed significant promotion on osteogenic
differentiation of rMSCs due to the facilitation of Nano-HAP. Accordingly, the incorporation of Nano-HAP could improve
the physicochemical and biological properties of PLA microspheres, and the resulting PLA/Nano-HAP composite micro-
spheres could be expected to achieve good application in tissue repair.

Keywords  Poly(lactic acid) · Nano hydroxyapatite · Protein adsorption · Cell proliferation

1 Introduction the basic step of changing surface properties to biologi-

cal properties. The composition and conformation of the
Surgical involvements are required for severe bone defects adsorbed protein layer is considered to be one of the
associated with trauma, tumor and other infections. Tissue major factors in determining the nature of cell interaction
engineering is a promising strategy for bone regeneration with the materials. The enhancement of cell adhesion and
in repairing massive bone defects [1, 2]. There into, the proliferation on the bone scaffolds is usually responsible
biomaterials with higher potential of bone regeneration for eventual tissue integration and the amount of new
are crucial for repairing bone defects. Moreover, protein bone formation [3–5].
adsorption onto biomaterials is very important as it is

H. M. C. Suboda Wijerathne and Dong Yan: Co-first authors.

Electronic supplementary material  The online version of this article (https​://doi.org/10.1007/s4245​2-020-2531-8) contains
supplementary material, which is available to authorized users.

*  Yingchao Han, hanyingchao@whut.edu.cn | 1State Key Laboratory of Advanced Technology for Materials Synthesis and Processing,
Biomedical Materials and Engineering Research Center of Hubei, Wuhan University of Technology, Wuhan 430070, Hubei,
People’s Republic of China. 2Department of Physical Sciences and Technology, Faculty of Applied Sciences, Sabaragamuwa University of Sri
Lanka, Belihuloya 70140, Sri Lanka.

SN Applied Sciences (2020) 2:722 | https://doi.org/10.1007/s42452-020-2531-8

Vol.:(0123456789)
Research Article SN Applied Sciences (2020) 2:722 | https://doi.org/10.1007/s42452-020-2531-8

Poly(lactic acid) (PLA) is a promising candidate as 2.3 Preparation of PLA/Nano‑HAP composite

tissue engineering scaffold due to its good biocompat- microspheres
ibility and biodegradability. However, pure PLA shows a
hydrophobic surface and the acidic degradation prod- PLA/Nano-HAP composite microspheres were prepared
uct easily causes aseptic inflammation, which are not by emulsion solvent evaporation method. 5 mg/mL PVA
conducive to the adhesion and proliferation of cells. aqueous solution (water phase) was prepared by dis-
Hydroxyapatite (HAP) is the major inorganic component solving PVA in UP water and 50 mg/mL PLA solution (oil
of human bones, and has good biocompatibility, bioac- phase) was prepared by dissolving PLA in DCM. 0.25 g,
tivity and osteogenic activity. It exists as Nano crystals 0.5 g and 0.75 g of Nano-HAP was introduced to PLA
in the organic/inorganic composite structure of human solution and ultrasonically dispersed for one minute
bones. The incorporation of HAP can endow the bioac- in order to prepare 10%, 20% and 30% Nano-HAP con-
tivity and osteogenic activity, and improve cell affinity tained composite microspheres, respectively. Then PLA/
[6–8]. Therefore, the organic/inorganic composites by Nano-HAP mixture was introduced to PVA solution with
mimicking natural bone attract attentions, aiming to 1:10 of oil to water ratio and the mixture was mechani-
improve the natural process of bone regeneration. cally agitated for 240  min. Then the obtained micro-
Microspheres as injectable scaffolds are among highly spheres were washed three times with UP water and
effective candidates for the application of bone defect separated by centrifugation. Finally the samples were
repair [9–13]. In this study, PLA/Nano-HAP composite freeze dried and used for further analysis.
microspheres were prepared, and the effect of Nano-
HAP on the surface property of microspheres including
hydrophilicity, protein adsorption and cell adhesion was 2.4 Characterization of samples
investigated. This study is strictly aimed to the improve-
ment of protein adsorption, cell adhesion and prolif- X-ray diffraction (XRD, D/Max-IIIA, Rigaku Co., Tokyo,
eration of PLA microspheres, which is significant and Japan) and fourier transform infrared spectroscopy (FT-
highly demanded for applications of PLA in bone tissue IR, Thermo Nicolet 6700, USA) were used to identified
engineering. Nano-HAP. Microstructure of sample was characterized
by field emission scanning electron microscope (FESEM,
Zeiss Ultra Plus) and transmission electron microscope
(TEM, JEM-1400Plus). The wetting property of sample
2 Experimental procedure was measured using contact angle meter (JC2000C,
POWEREACH).
2.1 Materials

Analytical pure ­CaCl 2·2H 2O, ­( NH 4) 2HPO 4, ­N H 4OH and 2.5 Analysis of protein adsorption of samples
dichloromethane (DCM) were perchased from Sinopharm
Chemical Reagent Co., Ltd. Analytical pure polyvinyl alco- BSA standard solution series (0.2–5.0 mg/mL) were pre-
hol (PVA) was purchased from Aladdin. Biomedical grade pared by serial dilution of 5.0 mg/mL BSA stock solution
PLA with a molecular weight of 100,000 was purchased with PBS. BCA A and BCA B from the protein assay kit
from JINAN Daigang BIO Engineer Limited Co. (Beyotime, China) was mixed with 1:50 ratio to obtain
the protein assay mixture. 20 µL BSA standard solution
series were added in 96-wells plate and 200 µL BCA
2.2 Preparation of Nano‑HAP solution was added subsequently. After incubation
of 30  min at 37 ºC, the absorbance was measured at
0.0334 mol/L ­CaCl2·2H2O aqueous solution and 0.02 mol/L 562 nm using microplate reader (Multiskan Go, Thermo
­( NH 4 ) 2 HPO 4 was prepared by ultrapure (UP) water. Scientific). The standard curve was obtained while
Then ­(NH4)2HPO4 aqueous solution was introduced to BSA concentration was from 0.2 mg/mL to1.8 mg/mL
­CaCl2·2H2O solution with 1:1 of volume to volume ratio (Y = 0.02649 + 0.45892X, ­R2 = 0.9996).
and the mixture was stirred with magnetic stirrer. Con- The adsorption of BSA on microspheres vs time was
centrated ­NH4OH was added to the mixture with 3:200 carried out. 40 mg of microsphere was put into 10 mL
of volume to volume ratio (­ NH4OH: solution) in order to Eppendorf tube. Then, 10 mL 37 ºC-incubated BSA solu-
maintain the pH of the solution between 9 and 10. This tion (1.0 mg/mL) was added and Eppendorf tube was
mixture was maintained at 37 ºC for three hours to prepare incubated at 37 ºC. At setting incubation times of 0.5,
Nano-HAP.

Vol:.(1234567890)
SN Applied Sciences (2020) 2:722 | https://doi.org/10.1007/s42452-020-2531-8 Research Article

1.0, 1.5, 2.0, 3.0, 5.0 and 7.0 h, 20 µL aliquots were col- 3 Results and discussion
lected for BCA assay, respectively. P values were calcu-
lated from SPSS statistical software. 3.1 Characterization of Nano‑HAP
Similarily, the adsorption of BSA on microspheres vs and microspheres
BSA concentration was also carried out. All samples were
incubated at 37 ºC for 4 h. The Nano-HAP was first synthesized for the preparation
of PLA/Nano-HAP composite microspheres. The XRD pat-
tern (Fig. S1 (a)) showed the broadened diffraction peaks
2.6 In vitro cellular evaluation assigned to HAP (JCPDS 84-1998), indicating a low crystal-
linity of resulting sample. The FTIR spectrum (Fig. S1 (b))
Rat mesenchymal stem cells (rMSCs) were seeded and further displayed the characteristic vibrations assigned to
cultured on microspheres to assess the biological prop- HAP (3563 cm−1: OH– group in HAP; 1104 cm−1, 1035 cm−1,
erties of microspheres. In cell adhesion test, after 7 and 963 cm−1, 603 cm−1, 565 cm−1 and 472 cm−1: ­PO43− group
14 days of cell culture, anti-osteopontin (OPN) antibody in HAP) [14, 15]. TEM observation (Fig. S2) revealed that
was added to recognize OPN. Then FITC-conjugated sample consisted of short rod-like Nanoparticles with size
secondary antibody was used to complete the immu- of about 35.78 nm × 8.06 nm.
nostaining. Cell nuclei were stained with DAPI. After Figure S3 illustrates the SEM images of pure PLA
immunostaining, cells were observed by fluorescence microspheres and PLA/Nano-HAP composite micro-
microscope (Nikon Eclipse CI, Japan). To further observe spheres, respectively. Results showed that the regular
the morphology of cells on microspheres, cells were microspheres of PLA and PLA/Nano-HAP composite were
fixed and then examined by SEM (TESCAN, VEGA3LMU) obtained. The mean diameter of pure PLA microspheres
on day 14. ALP assay was performed by the modified was 16.55 ± 9.47 µm. With the incorporation of Nano-HAP,
Gomori’s calcium-cobalt method and examined with the mean diameter of PLA/Nano-HAP composite micro-
biological microscope (Nikon Eclipse CI) on day 7 and spheres was increased to about 30  µm; however, the
14. Mineralized nodules stained with alizarin red were composite microspheres with 10%, 20% and 30% Nano-
observed by biological microscope (Nikon Eclipse CI) on HAP showed no significant difference in mean diameters
day 21. (29.86 ± 6.47 µm, 28.12 ± 5.48 µm and 29.81 ± 5.33 µm).

Fig. 1  Surface morphol-
ogy of microspheres. a Pure
PLA; b PLA/10% Nano-HAP;
c PLA/20% Nano-HAP; d
PLA/30% Nano-HAP

Vol.:(0123456789)
Research Article SN Applied Sciences (2020) 2:722 | https://doi.org/10.1007/s42452-020-2531-8

The surface morphology of microspheres was changed

due to the incorporation of Nano-HAP (Fig. 1). The pure
PLA microspheres had very smooth surface. However, the
composite microspheres showed rough surface due to the
incorporation of Nano-HAP. Moreover, the surface became
much rougher with the increase of Nano-HAP content
from 10 to 30%. The Nano-HAP particles were embed-
ded in PLA matrix. While the content of Nano-HAP was
10%, Nano-HAP particles exhibited almost single-particle
dispersion in PLA. At higher Nano-HAP content, Nano-
HAP particles were embedded in aggregations of about
200–400 nm; however, the distribution of Nano-HAP par-
ticles was almost uniform on micrometer scale. In addition,
the elemental mapping test was used to reveal the distri-
bution of Nano-HAP in composite microspheres (Fig. 2). Fig. 3  Contact angle of microspheres and nHAP. A: pure PLA; B:
PLA/10% Nano-HAP; C: PLA/20% Nano-HAP; D: PLA/30% Nano-
The observed signals of Ca demonstrated that Nano-HAP HAP; E: 37ºC-synthesized Nano-HAP
particles were distributed uniformly in the surface layer of
composite microspheres.
The surface hydrophilicity of microspheres was also composite microspheres with 10%, 20% and 30% Nano-
varied with the incorporation of Nano-HAP as shown in HAP were 97°, 71° and 66°, respectively. Compared to PLA
Fig. 3. PLA microspheres, a hydrophobic polymer material, microspheres, PLA/Nano-HAP composite microspheres
showed a contact angle of about 114° to water. However, showed an improvement of hydrophilicity due to the
Nano-HAP, a hydrophilic inorganic material, displayed a embedding of Nano-HAP, depending on the content of
contact angle of about 44° to water. The contact angles of Nano-HAP.

Fig. 2  Elemental mapping (C,

O, Ca) of PLA/10% Nano-HAP
composite microspheres

Vol:.(1234567890)
SN Applied Sciences (2020) 2:722 | https://doi.org/10.1007/s42452-020-2531-8 Research Article

Fig. 4  The variation of BSA adsorption on microspheres with time. Fig. 5  The variation of BSA adsorption on microspheres with con-
(a): pure PLA; (b): PLA/10% Nano-HAP; (c): PLA/20% Nano-HAP; (d): centration. (a): pure PLA; (b): PLA/10% Nano-HAP; (c): PLA/20%
PLA/30% Nano-HAP Nano-HAP; (d): PLA/30% Nano-HAP

3.2 Analysis of protein adsorption of microspheres enhanced. With BSA concentration over 2.0 mg/mL, the
change of adsorption quantity became flat. Differently,
As shown in Fig. 4, PLA and PLA/Nano-HAP microspheres PLA/Nano-HAP composite microspheres exhibited higher
showed similar adsorption profiles for BSA with the adsorption quantity for BSA than pure PLA microspheres.
increase of adsorption time. The BSA adsorption quickly Moreover, the larger content of Nano-HAP led to higher
increased in the first 3 h and then reached to an equilib- adsorption quantity. Accordingly, the incorporation of
rium. However, there were some differences between PLA Nano-HAP in PLA was beneficial for the adsorption of BSA
microsphere and PLA/Nano-HAP microspheres. HAP has on microspheres. The adsorption isotherm studies were
good adsorption property for protein and can be used done for pure PLA microspheres and PLA/20% Nano-HAP
as protein delivery system [16–18]. The incorporation of composite microspheres using the data shown in Fig. 5.
Nano-HAP resulted in a little increase of adsorption rate for Langmuir model, Freundlich model and Templin model
BSA. In addition, at same time point, the adsorption quan- were considered to describe the equilibrium behavior of
tity was significantly enhanced with the increase of Nano- microspheres (Fig. S6). Results showed that the adsorp-
HAP content from 0 to 30% (p < 0.05). These indicated tions of PLA microspheres and composite microspheres
that Nano-HAP could promote the adsorption of BSA on for BSA were all most fitted to Langmuir type adsorption
microspheres including adsorption rate and adsorption isotherm model.
quantity. As an example of PLA/20% Nano-HAP composite
microspheres, the kinetic behavior of BSA adsorption by 3.3 In vitro cellular evaluation
microspheres was studied using dynamic data pretended
in Fig. S4. The pseudo first order kinetic model, pseudo sec- As shown in Fig. 6, PLA microspheres and PLA/Nano-HAP
ond order kinetic model and intraparticle diffusion model composite microspheres could both support cells adhe-
were proposed to clarify the adsorption kinetics. As shown sion. However, PLA/Nano-HAP composite microspheres
in Fig. S5, the higher regression coefficient ­(R2) of 0.92072 showed better adhesion and proliferation of rMSCs than
demonstrated that the pseudo first order kinetic model PLA microspheres. This could be attributed to the com-
was more appropriate to describe the dynamic behavior ponent of Nano-HAP in PLA microspheres [19]. Moreover,
of protein adsorption on composite microspheres. the rough surface due to the incorporation of Nano-HAP
The adsorption of BSA on microspheres was further might promote the adhesion of rMSCs [20–22].
determined with varying BSA concentration. As shown In addition, results (Fig. 7) demonstrated that Nano-
in Fig. 5, PLA and PLA/Nano-HAP microspheres displayed HAP showed a facilitation on osteogenic differentia-
similar adsorption profiles for BSA with the increase of BSA tion of rMSCs. After 7 days and 14 days of cell culture
concentration. While BSA concentration was increased (Fig. 7a–d), PLA/Nano-HAP sample displayed more posi-
from 0.2 to 2.0 mg/mL, the adsorption quantity was quickly tive staining area for ALP activity compared with pure

Vol.:(0123456789)
Research Article SN Applied Sciences (2020) 2:722 | https://doi.org/10.1007/s42452-020-2531-8

Fig. 6  Cell adhesion on PLA

and PLA/Nano-HAP samples

PLA sample, indicating Nano-HAP markedly promoted and mineralized nodules. Spindle cell and its outspread
osteogenic differentiation of rMSCs. The mineralized pseudopods were observed on PLA/Nano-HAP sample
nodules stained positive for alizarin red on PLA/Nano- (Fig. 7f ). Moreover, some small particles secreted by cells
HAP sample further proved the expression of osteo- were found on the surface of cells, and the acceleration
genic phenotype of cells under the effect of Nano-HAP and agglomeration of these particles formed the miner-
(Fig. 7e). SEM images exhibited details of cell adhesion alized nodules (Fig. 7g).

Vol:.(1234567890)
SN Applied Sciences (2020) 2:722 | https://doi.org/10.1007/s42452-020-2531-8 Research Article

Fig. 7  a ALP staining of PLA

sample on 7 days , b ALP
staining of PLA/HAP sample
on 7 days, c ALP staining of
PLA sample on 14 days, d ALP
staining of PLA/HAP sample on
14 days, e mineralized nodules
stained with alizarin red on
PLA/HAP sample, f and g SEM
images of cells on PLA/HAP
sample

4 Conclusions Moreover, the incorporation of Nano-HAP promoted the

adsorption capacity of PLA microspheres for BSA. The
PLA/Nano-HAP composite microspheres were successfully protein adsorption on PLA/Nano-HAP composite micro-
prepared, achieving the embedding of Nano-HAP particles spheres followed pseudo first order kinetic model and
of 10–30% content in PLA microspheres. Results showed Langmuir adsorption isotherm model. Cell culture experi-
the increases of surface hydrophilicity and surface rough mental results showed that PLA/Nano-HAP composite
of microspheres depending on the content of Nano-HAP. microspheres had higher cell adhesion and proliferation

Vol.:(0123456789)
Research Article SN Applied Sciences (2020) 2:722 | https://doi.org/10.1007/s42452-020-2531-8

compared to pure PLA microspheres. The addition of 8. Cunniffe GM, Curtin CM, Thompson EM, Dickson GR, O’Brien
Nano-HAP can markedly promote osteogenic differen- FJ (2016) Content-dependent osteogenic response of nanohy-
droxyapatite: an in vitro and in vivo assessment within collagen-
tiation of rMSCs. Accordingly, PLA/Nano-HAP composite based scaffolds. ACS Appl Mater Interfaces 8:23477–23488
microsphere with an improved ability for bone regenera- 9. Nagata F, Miyajima T, Teraoka K, Yokogawa Y (2005) Prepara-
tion can be used as a promising material for bone tissue tion of porous poly (lactic acid)/hydroxyapatite microspheres
engineering. intended for injectable bone substitutes. Key Eng Mater
284–286:819–822
10. Lee TJ, Kang SW, Bhang SH, Kang JM, Kim BS (2010) Apatite-
coated porous poly(lactic-co-glycolic acid) microspheres as an
5 Supplementary information injectable bone substitute. J Biomater Sci-Polym E 21:635–645
11. Lee YK (2010) Nanocomposite microspheres of PLGA/HA with
antibiotics for injectable bone-graft materials. Tissue Eng Regen
Supplementary information is provided including XRD, Med 7:561–565
FTIR and TEM of Nano- HAP and the SEM of the PLA/Nano- 12. Ahmed I (2015) Development of microspheres for biomedical
HAP composite microspheres, particle size distribution of applications : a review. Prog Biomater 4:1–19
composite microspheres and adsorption data. 13. Liao HB, Lanao RPF, van den Beucken JJJP, Zhou N, Both SK,
Wolke JGC, Jansen JA (2016) Size matters: effects of PLGA-
microsphere size in injectable CPC/PLGA on bone formation. J
Acknowledgements  This work was supported by the National Tissue Eng Regen M 10:669–678
Key Research and Development Program of China (Grant 14. Poorraeisi M, Afshar A (2019) Synthesizing and comparing HA–
#2016YFB1101300) and by the Science and Technology Partner- TiO2 and HA–ZrO2 nanocomposite coatings on 316 stainless
ship Program of Ministry of Science and Technology of China (Grant steel. SN Appl Sci 1:155
#KY201602002). 15. Sundarabharathi L, Ponnamma D, Parangusan H et al (2020)
Effect of anions on the structural, morphological and dielectric
Compliance with ethical standards  properties of hydrothermally synthesized hydroxyapatite nano-
particles. SN Appl Sci 2:94
Conflict of interest  The authors declare no competing financial inter- 16. Kandori K, Mukai M, Yasukawa A, Ishikawa T (2000) Competi-
est. tive and cooperative adsorptions of bovine serum albumin
and lysozyme to synthetic calcium hydroxyapatites. Langmuir
16:2301–2305
17. Boonsongrit Y, Abe H, Sato K, Naito M, Yoshimura M, Ichikawa H,
References Fukumori Y (2008) Controlled release of bovine serum albumin
from hydroxyapatite microspheres for protein delivery system.
1. Wei G, Ma PX (2004) Structure and properties of nano- Mater Sci Eng B-Adv 148:162–165
hydroxyapatite/polymer composite scaffolds for bone tissue 18. Tanaka T, Takai Y, Nagase A, Teraguchi K, Minbu H, Ochiai A,
engineering. Biomaterials 25(19):4749–4757 Kimura I, Taniguchi M (2019) Protein adsorption characteristics
2. Khan F, Ahmad SR (2013) Chapter 5 Bioactive polymers and of nanoparticle-assembled hollow microspheres of hydroxyapa-
nanobiomaterials composites for bone tissue engineering, bio- tite and their composites with PLLA microporous membranes.
mimetics: advancing nanobiomaterials and tissue engineering. Heliyon 5:e01490
Wiley, Hoboken 19. Lee W, Loo C, Zavgorodniy AV, Ghadiri M (2013) A novel
3. Fu C, Yang X, Tan S, Song L (2017) Enhancing cell proliferation approach to enhance protein adsorption and cell proliferation
and osteogenic differentiation of MC3T3-E1 pre-osteoblasts by on hydroxyapatite : citric acid treatment. RSC Adv 3:4040–4051
BMP-2 delivery in graphene oxide-incorporated PLGA/HA bio- 20. Fan YW, Cui FZ, Chen LN, Zhai Y, Xu QY, Lee IS (2002) Adhesion
degradable microcarriers. Sci Rep 7:12549 of neural cells on silicon wafer with nano-topographic surface.
4. Szcześ A, Ho L, Chibowski E (2017) Synthesis of hydroxyapatite Appl Surf Sci 187:313–318
for biomedical applications. Adv Colloid Interface 249:321–330 21. Alves CM, Yang Y, Marton D, Carnes DL, Ong JL, Sylvia VL, Dean
5. Kucharska M, Walenko K, Lewandowska-szumiel M, Brynk T DD, Reis RL, Agrawal CM (2008) Plasma surface modification of
(2015) Chitosan and composite microsphere-based scaffold poly (D, L-Lactic Acid) as a tool to enhance protein adsorption
for bone tissue engineering: evaluation of tricalcium phosphate and the attachment of different cell types. J Biomed Mater Res
content influence on physical and biological properties. J Mater B Appl Biomater 87:59–66
Sci Mater Med 26:143 22. Baek SM, Shin MH, Moon J, Jung HS, Lee SA, Hwang W, Yeom JT,
6. Ni P, Bi H, Zhao G, Han Y, Wickramaratne MN, Dai H, Wang X Hahn SK, Kim HS (2017) Superior pre-osteoblast cell response
(2019) Electrospun preparation and biological properties in vitro of etched ultrafine-grained titanium with a controlled crystal-
of polyvinyl alcohol/sodium alginate/nano-hydroxyapatite com- lographic orientation. Sci Rep 7:44213
posite fiber membrane. Colloid Surf B 173:171–177
7. Santana-Melo GF, Rodrigues BVM, Silva E, Ricci R, Marciano Publisher’s Note Springer Nature remains neutral with regard to
FR, Webster TJ, Vasconcellos LMR, Lobo AO (2017) Electrospun jurisdictional claims in published maps and institutional affiliations.
ultrathin PBAT/nHAp fibers influenced the in vitro and in vivo
osteogenesis and improved the mechanical properties of neo-
formed bone. Colloid Surf B 155:544–552

Vol:.(1234567890)