Page 243 - IJB-10-5

International Journal of Bioprinting 3D printed hydrogels for tumor therapy

2-methylpropiophenone, and ammonium hydroxide (PBS; pH 7.4, Gibco) at 50°C under constant magnetic
solution (NH OH; 28%) were purchased from Sigma- stirring for 1 h. Subsequently, 0.8 mL (for GelMAL) or 8.0
4
Aldrich (St. Louis, MO, USA). Dopamine hydrochloride mL (for GelMAH) methacrylic anhydride (MA) was added
(98%) and doxorubicin hydrochloride (DOX) were bought dropwise to the gelatin solution. The reaction continued
from Aladdin Co., Ltd., China. Sodium chloride (NaCl), for 3 h and was then stopped by adding five times the
sodium hydroxide (NaOH), anhydrous chloride (CaCl ), volume of PBS to the system. Afterward, the solution was
2
magnesium chloride hexahydrate (MgCl ·6H O), disodium transferred into a dialysis tubing cellulose membrane and
2
2
hydrogen phosphate dodecahydrate (Na HPO ·12H O), dialyzed against DI water at 40°C for 1 week. The DI water
4
2
2
and ethanol were supplied by Sinopharm Chemical was refreshed daily. Finally, GelMA was freeze-dried and
Reagent Co., Ltd., China. Dialysis tubing cellulose stored at 4°C for further use. The degree of methacrylation
1
membrane (MWCO: 10 kDa) was bought from Thermo of GelMA was characterized using H NMR spectroscopy
Fisher Scientific. All reagents were used as-received (Bruker Avance III 400, Germany). The secondary
without further purification. structure of GelMA was studied using a circular dichroism
(CD) spectrometer (JASCO J-815, Japan) at 4 and 37°C,
2.2. Synthesis of MgHAp nanocomposites, GelMA, respectively. The characteristic groups of GelMA were
and PDA@DOX particles investigated using an FT-IR spectrometer under ATR
HAp and MgHAp nanocomposites were synthesized mode. Moreover, the phase transition temperatures of
via a biomimetic mineralization process following the GelMAH and GelMAL solutions were determined using a
procedure as described in the previous study. Briefly, 0.5 Rheometer (MCR 302, Anton Paar, Austria).
35
g COL1 was dissolved in 100 mL deionized (DI) water
under constant magnetic at a 37°C water bath. Then, 500 PDA and PDA@DOX particles were synthesized
mL 0.1 M CaCl , 300 mL 0.1 M Na HPO , 40 mL 1 M following the procedure described in our previous study
2
37
4
2
citric acid, and 3.6 g NaCl were added in a sequence order. with slight modifications. Briefly, 350 mL absolute
Subsequently, a 4 M NaOH solution was added dropwise ethanol, 12 mL NH OH, and 288 mL DI water were mixed
4
to adjust the pH to 9–10. The reaction system was then homogenously under constant magnetic stirring for 30
incubated in an incubator at 37°C for 7 days. Afterward, min. Dopamine hydrochloride (2.5 g) was dissolved in 50
the white precipitate was suspended in 75% ethanol. The mL DI water. The dopamine solution was then added to
precipitate was centrifuged at 12,000 rpm/min for 10 min the mixture for reaction at room temperature in an open-
and washed with DI water three times and then freeze- air environment for 24 h. PDA particles were obtained
dried. Finally, HAp nanocomposites were obtained and followed the centrifugation at 12,000 rpm/min for 10 min
stored at 4°C for further use. For the synthesis of MgHAp and subsequently dried in an oven at 80°C. PDA@DOX
nanocomposites, 500 mL 0.1 M CaCl was replaced by particles were synthesized using the same procedure but
2
450 mL 0.1 M CaCl and 50 mL 0.1 M MgCl , while the with the addition of 0.5 g DOX dissolved in the dopamine
solution. The morphology and microstructure of PDA
2
2
other procedures remained the same. The morphology and PDA@DOX particles were examined using SEM and
and microstructure of HAp and MgHAp nanocomposites TEM. Moreover, the diameter of the PDA and PDA@DOX
were characterized using a field emission scanning particles was calculated using the ImageJ software.
electron microscope (SEM; Hitachi S4800, Japan) and
transmission electron microscope (TEM; Themis, Thermo 2.3. Rheological properties of printing inks
Scientific, USA). The crystallographic phase of HAp and The compositions of the printing inks are listed in Table 1.
MgHAp nanocomposites was investigated using an X-ray Briefly, taking MgHAp/GelMA-PDA as a typical example,
diffractometer (XRD; 7000S, Shimadzu, Japan) over a a 20% w/v GelMA solution was prepared by dissolving 10%
range of 2θ from 10° to 70°. The characteristic groups in w/v GelMAL and 10% w/v GelMAH in PBS at 40°C in a
HAp and MgHAp nanocomposites were examined using water bath under constant magnetic stirring. Subsequently,
an FT-IR spectrometer (PerkinElmer, USA) under the 2% w/v MgHAp and 0.5% w/v PDA were added to the
attenuated total reflection (ATR) mode. GelMA solution. Photoinitiator was added to the printing
Two sets of GelMA with a high degree of methacrylation ink with a final concentration of 0.2% w/v.
(~80%) and a low degree of methacrylation (~20%) were The phase transition temperatures of the printing inks
synthesized according to the previous reported method. were characterized from 10 to 40°C using a rotational
26
Accordingly, GelMA with a high degree of methacrylation rheometer equipped with a parallel plate unit with a 20 mm
was designated as GelMAH, while GelMA with a low degree diameter and 1.0 mm measurement gap (1% strain and 1
of methacrylation was designated as GelMAL. Briefly, 10 g Hz). The shear-thinning behavior of the printing inks was
−1
gelatin was dissolved in 100 mL phosphate-buffered saline studied at shear rate ranging from 0.1 to 100 s at 25°C.

Volume 10 Issue 5 (2024) 235 doi: 10.36922/ijb.3526