In Vitro Biological Performance of Alginate Hydrogel Capsules for Stem Cell Delivery

Abstract

Encapsulation of biological components in hydrogels is a well described method for controlled drug delivery of proteins, tissue engineering and intestinal colonization with beneficial bacteria. Given the potential of tissue engineering in clinical practice, this study aimed to evaluate the feasibility of encapsulation of adipose tissue-derived mesenchymal stem cells (MSCs) of mules in sodium alginate. We evaluated capsule morphology and cell viability, immunophenotype and release after encapsulation. Circular and irregular pores were observed on the hydrogel surface, in which MSCs were present and alive. Capsules demonstrated good capacity of absorption of liquid and cell viability was consistently high through the time points, indicating proper nutrient diffusion. Flow cytometry showed stability of stem cell surface markers, whereas immunohistochemistry revealed the expression of CD44 and absence of MHC-II through 7 days of culture. Stem cell encapsulation in sodium alginate hydrogel is a feasible technique that does not compromise cell viability and preserves their undifferentiated status, becoming a relevant option to further studies of tridimensional culture systems and in vivo bioactive agents delivery.

Keywords: alginic acid; biocompatibility; biomaterial; cell encapsulation; tridimensional culture.

FIGURE 1

Observation of a capsule by a magnifier, showing a central concentration of cells compared to the edges. Scale bar: 1000 µm.

FIGURE 2

SEM analysis of capsules with MSCs showing the capsule surface, pores and superficial MSCs. Black arrows indicate pores (A) and MSCs entrapped in the alginate network (B). Magnification of 500x.

FIGURE 3

Morphological structure by SEM showing the capsule surface and presence of cells (arrows) at magnification of 1,200x (A) and 5,000x (B).

FIGURE 4

Swelling behavior of pristine alginate capsules (n = 10) and capsules with MSCs (n = 10). Although capsules with MSCs showed less stability in mass through time, there was a significant increase in mass of the group of capsules alone at 21 h compared to the initial time point (* indicates p < 0.05).

FIGURE 5

Moisture content of the capsules (n = 5). There was a pronounced reduction in moisture content after drying the material at 50°C for 3 h (**** means p < 0.0001).

FIGURE 6

(A) DAPI positive labelling showing cell nuclei stained in blue. (B) Qtracker positive labeling showing cell cytoplasm stained in red. (C) Labellings merged. White arrow indicates the border of the capsule.

FIGURE 7

(A) Viability of encapsulated MSCs at 0, 24, 48, 72 h and 7 days, showing a maintenance of high viability through time. (B) Cell migration from the scaffold to the plastic bottom of the wells (* means p = 0.0434).

FIGURE 8

MTT assay showing a decrease in metabolic activity at 24 and 72 h time points and a recovery at 7 days. Adherent MSC culture was used as control.

FIGURE 9

Immunophenotypic characterization by flow cytometry of MSCs 24 h (A) and 7 days (B) after encapsulation. M1 indicates the area of positivity for each marker. There was maintenance of positivity for CD90 and negativity for CD11b, CD45 and MHCII.

FIGURE 10

Immunohistochemistry of encapsulated MSCs. (A) Negative control of the sample; (B) Positive labelling of CD44 and C) Absence of labelling by MHC II. Harris’ hematoxylin counterstaining, 20x.