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(A) The immunostaining of fi bronectin expression in PGAC cells on TCPS, which showed a random distribution pattern (bar: 25 m m). (B) The Western blot of adsorpted fi bronectin on PLGA and TCPS surface. (C) SEM images of PLGA and TCPS surface (bar: 30 m m).
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As a potential solution for patients to retrieve their lost salivary gland functions, tissue engineering of an auto-secretory device is profoundly needed. Under serum-free environment, primary human parotid gland acinar (PGAC) cells can be obtained. After reaching confluence, PGAC cells spontaneously form three-dimension (3D) cell aggregations, ter...
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... of cultured cells. In our previous study [5], human PGAC cells have been successfully isolated and expanded by using K-SFM- based medium (Fig. 1) and cells in over-crowded regions could form three-dimensional PCSs to exhibit higher expression levels and different location patterns of function-related proteins compared to the 2D cells (Fig. 2). This is consistent with previous studies that 3D culture systems can mimic the in vivo environment for acinar cells to recover their function. For example, it has also been shown that the ability of hair follicle induction of dermal papilla cells [17], albumin production of hepatocytes [23], dopamine secretion of PC12 cells [24] are enhanced when they are cultured into 3D aggregates. A number of methods can be employed to culture cells into 3D aggregates. The simplest way is to maintain cells in spinner fl asks where the culture system is continuously rotated, thereby pre- venting cells from adhering to the culture vessel. Another in vitro method of inducing 3D structures involves culturing cells within a gel system. When cells are seeded in gels, their movement may be con fi ned and grow into dense aggregates. Third, cell aggregates can also be obtained by direct scraping under high cell densities. However, these methods do not allow investigation of the cell self- assembly process. Therefore, in the present study, we disclose the feasibility of using the FDA-approved biomaterial PLGA to culture PGAC cells for promoting the formation of three-dimensional PCS. Compared to TCPS, PLGA can facilitate PCS formation, both in the sizes and the numbers (Fig. 3). It is reasonable to assume that cells form 3D structure on a 2D surface via migration after close intercellular contact can be reached. We may envision the rate-determining steps of PCS formation to be either the rate of cell migration to the PCS surface (migration control by cell-substrate interaction) or the rate of adhesion of cell to the PCS surface (adhesion control by cell e cell interaction). Straightforwardly, the higher cell migration rate the more possibility for cell e cell contact, so the formation of more PCSs can be expected. Based on this assumption, PLGA should be able to promote PGAC cell migration to form more and lager PCSs than TCPS. However, Fig. 5 shows PGAC cells displayed more active random movement on TCPS than on PLGA. Therefore, cell migration may not be essential for the formation of PCS on PLGA. This is consistent with the fact that PCS was observed after PGAC cells reached con fl uence. In addition to the dynamics of cell migration, another possibility accounting for the formation of PCS is that the static force of cell e cell interaction is able to provide strong intercellular adhesion. The factors affecting cell e cell interaction includes the trophic factors, serum factors, integrins, and adhesions molecules. It has been shown that E-cadherin is the major mediator for cell cohesion [25]. As shown in Fig. 6, obvious E-cadherin pattern alongside the cell boundary within 3D PCS was observed. Thus, we proposed that surface adhesion molecules, especially E-cadherin, may play an important role in mediating homophilic intercellular binding during the process of PCS formation. If this is the case, the enhancement of E-cadherin expression may be bene fi cial for the formation of PCS and the inhibition of E-cadherin expression is able to retard the formation of PCS. The observation that Src signaling induces a switch in adhesion type predominance from cadherin- to integrin-based adhesions [26] leads us to hypothesize that Src activity could have an opposite role in E-cadherin-mediated PCS formation. Certainly, we demonstrated that the Src inhibitor PP1 exactly caused up-regulation of E- cadherin protein in PGAC cells and enhanced cell e cell adhesiveness in a low-seeding-density cell aggregation assay (Fig. 7). Further- more, PGAC treated with antibody against E-cadherin abolished the formation of PCS. These results suggest that the decision of PGAC cells to initiate PCS formation requires the formation of E-cadherin cell e cell junctions. Hence, the static force of homophilic interaction on surfaces of individual cells, but not the dynamics of cell migration, may play a more important role in PCS formation. We next asked whether biomaterial could play a role in the E- cadherin-dependent cell adhesion to mediate PCS formation. It is known the cell e biomaterial interaction is of extreme importance in regulating the numerous functions necessary for cell adhesion, growth and differentiation. The direct contact between cell and substrate is the fi rst step for cell adhesion onto the substrate, but it is generally considered to be a multistep process involving adsorption of ECM proteins onto the substrate surface and recog- nition of ECM components by cell surface receptors, followed by cytoskeletal rearrangements that lead to cell spreading [27,28]. Indeed, the fi bronectin adsorption test in Fig. 4 reveals that PLGA and TCPS had different ECM adsorption ability in this study. Among cell surface receptors, integrin receptor is mainly responsible for cell-ECM interactions to induce the formation of focal adhesions and to generate a cascade of phosphorylation of signal transduction molecules to modulate signaling pathways. Focal adhesion kinase (FAK) is located at focal contact and plays a key role in regulating cell spreading and migration. On the other hand, the role of integrin-linked kinase (ILK) has been found to interact with b 1 and b 3 integrins and to transduce signals from ECM components to downstream signaling components [19]. The activation of ILK in epithelial cells has been shown to result in loss of cell e cell adhesion, due to the down-regulation of E-cadherin expression [19]. The down-regulation of E-cadherin expression also involves ILK-mediated activation of the E-cadherin suppressor, snail [29]. Thus, interaction between integrin and ECM proteins also modulates snail phosphorylation [30]. Based on these previous studies, we hypothesize that biomaterials can modulate the PCS formation in PGAC cells via the ligand-binding of integrins, ILK signaling pathway, and regulation of the E-cadherin expression [22]. In this study, PGAC cells on TCPS showed more developed focal adhesion contacts and more effective cell attachment than those on PLGA (Fig. 8). By Western blot analysis, PLGA mediated lower p-FAK and ILK expression of PGAC cells, which in turn regulates the expression of snail and E-cadherin with a reciprocal relationship (Fig. 9). Since PGAC cells were found to have a greater tendency to form PCS on PLGA than on TCPS (Fig. 3), our data demonstrate that biomaterial can support PGAC cells to form more PCS through the effects on enhancing E-cadherin expression, which is associated with the suppression of FAK/ILK/snail in PGAC cells. Thus, selective appropriate biomaterials may be potentially useful in generating PCS and recover epithelial characteristics. On the other hand, since inhibition of Src also resulted in stimulating E-cadherin expression, we examined whether the expression of E-cadherin could be regulated by Src, on different biomaterials. However, as shown in Fig. 7D, this does not seem to be the case, since phosphorylation of P38 (data not shown) and Src had no signi fi cant difference for PGAC cells cultured on PLGA or TCPS. Thus, Src signaling pathway is likely not involved in the biomaterial-stimulated E-cadherin expression. By the way, it is known that the conversion of an epithelial cell to a mesenchymal cell is a crucial step of tumorigeneses. The hallmarks for the epithelial to mesenchymal transformation (EMT) have been associated with the loss of epithelial phenotype, increased fi bronectin matrix deposition, production of transcription factor snail able to inhibit E-cadherin expression, and activation signaling molecules such as FAK and ILK [30 e 32]. Interestingly, this study suggests that E-cadherin can be considered to have an opposite role in PCS formation and EMT. Thus, application of biomaterial may also lead to another way of restoring the frequently observed down-regulation of E-cadherin expression in many types of carcinomas. When PGAC cells were cultured on PLGA, higher level of PCS formation was observed than on TCPS. This different formation tendency was regulated by the differential expression of E-cadherin molecules on both substrates. PLGA provided a more weaker cell- substrate adhesion for PGAC cells and serially down-regulated the downstream FAK, ILK, and Snail expression; however, the descending of Snail reversibly enhanced the endogenous E-cadherin expression, fi nally lead into a stronger cell e cell adhesion and the higher level of PCS formation. This study revealed the possibility of controlling cell aggregation behaviors by choosing different biomaterials. The authors thank National Science Council of the Republic of China for their fi nancial support of this research. This work is supported in part by the National Science Council of the Republic of China (NSC99-2628-B-002-050 MY3) to P-J ...
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... proteins have been extensively studied with respect to substrate adsorption and cellular response [18]. As shown in Fig. 4A, PGAC cells cultured in serum-free medium could secrete fibronectin, which was involved in the ILK pathway and E-cadherin expression regulation [19]. To confirm whether PLGA and TCPS have different ECM protein adsorption ability to influence PCS formation, PLGA and TCPS were incubated with 5 mg/cm 2 fibronectin for 48 h and then the ...
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... secrete fibronectin, which was involved in the ILK pathway and E-cadherin expression regulation [19]. To confirm whether PLGA and TCPS have different ECM protein adsorption ability to influence PCS formation, PLGA and TCPS were incubated with 5 mg/cm 2 fibronectin for 48 h and then the adsorption amount was quantified by Western blot analysis. Fig. 4B shows both PLGA and TCPS have a homogeneous smooth surface without considering surface roughness bias. Fig. 4C shows TCPS exhibited higher adsorption quantity of fibronectin than PLGA. These results indicate that PLGA and TCPS could adsorb different ECM proteins to talk with cultured ...
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... whether PLGA and TCPS have different ECM protein adsorption ability to influence PCS formation, PLGA and TCPS were incubated with 5 mg/cm 2 fibronectin for 48 h and then the adsorption amount was quantified by Western blot analysis. Fig. 4B shows both PLGA and TCPS have a homogeneous smooth surface without considering surface roughness bias. Fig. 4C shows TCPS exhibited higher adsorption quantity of fibronectin than PLGA. These results indicate that PLGA and TCPS could adsorb different ECM proteins to talk with cultured ...
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... step for cell adhesion onto the substrate, but it is generally considered to be a multistep process involving adsorption of ECM proteins onto the substrate surface and recognition of ECM components by cell surface receptors, followed by cytoskeletal rearrangements that lead to cell spreading [27,28]. Indeed, the fibronectin adsorption test in Fig. 4 reveals that PLGA and TCPS had different ECM adsorption ability in this ...
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Biodegradation is a key property for biodegradable polymer-based tissue scaffolds because it can provide suitable space for cell growth as well as tailored sustainability depending on their role. Ultrashort pulsed lasers have been widely used for the precise processing of optically transparent materials, including biodegradable polymers. Here, we d...
Citations
... Various culture techniques, including those for the generation of synthetic scaffolds that mimic the topographical and mechanical properties of the ECM, have been utilized to regulate salivary gland cell function and/or salivary tissue organization. Several research groups have reported that salivary epithelial cells and/or progenitors incorporated into synthetic or natural scaffolds using polylactic-co-glycolic acid (PLGA), poly(vinylidene fluoride) (PVDF), [29][30][31] and PEG 32 or chitosan-, 33 fibrin-, 34,35 and hyaluronic acid-based hydrogels 36,37 exhibit salivary tissue organization. Biomaterial-based culture processes that facilitate cell-to-cell or cell-to-ECM contact through the embedding or seeding of cells into ECM-based scaffolds 38 or micropatterned nanofibrous scaffolds 39,40 have also been reported to be of potential use for the modulation of salivary cell function. ...
Three-dimensional spheroid culture enhances cell-to-cell interactions among stem cells and promotes the expression of stem cell properties; however, subsequent retrieval and delivery of these cells remain a challenge. We fabricated a thermoresponsive fiber-based microwell scaffold by combining electrospinning and hydrogel micropatterning. The resultant scaffold appeared to facilitate the formation of cellular spheroids of uniform size and enabled the expression of more stem cell-secreting growth factor genes ( EGF, IGF-1, FGF1, FGF2, and HGF), pluripotent stem cell-related genes ( SOX2 and NANOG), and adult epithelial stem cell-related genes ( LGR4, LGR5, and LGR6) than salivary gland stem cells in a monolayer culture (SGSC monolayer ). The spheroids could be retrieved efficiently by decreasing temperature. SGSC-derived spheroid (SGSC spheroid ) cells were then implanted into the submandibular glands of mice at 2 weeks after fractionated X-ray irradiation at a dose of 7.5 Gy/day. At 16 weeks post-irradiation, restoration of salivary function was detected only in SGSC spheroid -implanted mice. The production of submandibular acini specific mucin increased in SGSC spheroid -implanted mice, compared with PBS control. More MIST1 ⁺ mature acinar cells were preserved in the SGSC spheroid -implanted group than in the PBS control group. Intriguingly, SGSC spheroid -implanted mice exhibited greater amelioration of tissue damage and preservation of KRT7 ⁺ terminally differentiated luminal ductal cells than SGSC monolayer -implanted mice. The SGSC spheroid -implanted mice also showed less DNA damage and apoptotic cell death than the SGSC monolayer -implanted mice at 2 weeks post-implantation. Additionally, a significant increase in Ki67 ⁺ AQP5 ⁺ proliferative acinar cells was noted only in SGSC spheroid -implanted mice. Our results suggest that a thermoresponsive fiber-based scaffold could be of use to facilitate the production of function-enhanced SGSC spheroid cells and their subsequent retrieval and delivery to damaged salivary glands to alleviate radiation-induced apoptotic cell death and promote salivary gland regeneration.
... Further, MSCs appear to be a reasonable therapeutic approach based on their availability, pluripotency, and immune-evasive properties. Successful regeneration will require identification of biomarkers for salivary stem/progenitor cells [15,40], elucidation of pathways/factors associated with salivary gland development/differentiation [75], and development of microenvironments (e.g., ECM/scaffold and specific cytokines/growth factors) to support proliferation and guide differentiation of salivary gland progenitors [41,76]. The results of these studies will facilitate stem cellebased therapy via regeneration of salivary gland tissue in vivo and engineering of artificial salivary glands in vitro. ...
... Nanofibers have been widely used in tissue engineering: they provide scaffolds with increased surface area, allowing for the organization of cells into a monolayer and resemble the native extracellular matrix (ECM) and basement membrane that are critical for gland development and regeneration [2][3][4]. Nanofiber scaffolds promote apico-basal polarity of salivary epithelial cells, a critical cell property for functional secretion. A common polymer used in engineering of both soft and hard tissues is poly-lactic-co-glycolic acid (PLGA) (stiffness~20 MPa-2.0 ...
Engineering salivary glands is of interest due to the damaging effects of radiation therapy and the autoimmune disease Sjögren's syndrome on salivary gland function. One of the current problems in tissue engineering is that in vitro studies often fail to predict in vivo regeneration due to failure of cells to interact with scaffolds and of the single cell types that are typically used for these studies. Although poly (lactic co glycolic acid) (PLGA) nanofiber scaffolds have been used for in vitro growth of epithelial cells, PLGA has low compliance and cells do not penetrate the scaffolds. Using a core-shell electrospinning technique, we incorporated poly (glycerol sebacate) (PGS) into PLGA scaffolds to increase the compliance and decrease hydrophobicity. PGS/PLGA scaffolds promoted epithelial cell penetration into the scaffold and apical localization of tight junction proteins, which is necessary for epithelial cell function. Additionally, co-culture of the salivary epithelial cells with NIH3T3 mesenchymal cells on PGS/PLGA scaffolds facilitated epithelial tissue reorganization and apical localization of tight junction proteins significantly more than in the absence of the mesenchyme. These data demonstrate the applicability of PGS/PLGA nanofibers for epithelial cell self-organization and facilitation of co-culture cell interactions that promote tissue self-organization in vitro.
... Recently, several authors reported that synthetic or natural polymers such as polyethylene glycol (PEG) [23], polylactic-coglycolic acid (PLGA) [24], fibrin [25], hyaluronic acid [26], chitosan [27], alginate [28], and silk fibroin [29] scaffolds could be used for the construction of an artificial gland to restore SG hypofunction because cultured primary SG cells could express acinar markers and TJ proteins. The nanofibrous bottom confers a number of advantages over conventional culture platforms since nanofibrous environments have consistently been shown to promote more in vivo-like cell behavior than standard culture surfaces [30]. ...
Statement of significance:
Three dimensional (3D) cultures of salivary glandular epithelial cells using nanofibrous bottom facilitate the formation of acinar-like organoids. In this study, we adapted a PEG hydrogel-micropatterned PCL nanofibrous microwell for the efficient bioengineering of human salivary gland organoids, in which we could easily produce uniform size of 3D organoids. This 3D culture system supports spherical organization, gene and protein expression of acinar markers, TJ proteins, adherence, and cytoskeletal proteins, as well as to promote epithelial structural integrity and acinar secretory functions, and results showed superior efficiency relative to Matrigel and nanofibrous scaffold culture. This 3D culture system has benefits in terms of inert, non-animal and serum-free culture conditions, as well as controllable spheroid size and scalable production of functional SG organoids and is applicable to bioengineering approaches for an artificial bio-gland, as well as to investigations of salivary gland physiology and regeneration.
... The gland-like structure also showed the production of human salivary a-amylase (acinar cell protein). [23,24] Gene therapy is another potential approach for salivary gland regeneration. It relies on the using of a recombinant adenovirus to deliver a water-channel protein gene (aquaporin) to the surviving ductal epithelial cells. ...
... 3,4 To accomplish these complex biological processes, several parallel lines of research have focused on identifying and isolating salivary gland stem cells/progenitor cells, 5,6 elucidating pathways and factors associated with salivary gland development, 4 and developing appropriate biomaterial scaffolds to support the proliferation and differentiation of salivary gland progenitors. 7,8 Extracellular matrix (ECM) is an important component of the cellular niche and supplies critical biochemical and physical signals to initiate or sustain cellular functions in the tissue. [9][10][11] Within the rapidly developing field of tissue engineering, various biomaterial scaffolds have been developed for tissue regeneration or repair that mimic some of the critical properties of the ECM. 12 Examples of biomaterials that have been shown to support the proliferation and differentiation of salivary gland progenitors include chitosan, polyglycolic acid (PGA), poly-(L)-lactic acid (PLLA), poly (lactic-co-glycolic acid) (PLGA), and poly(ethylene glycol)terephthalate/poly(butylene)terephthalate (PEGT/PBT) block co-polymers. ...
... [9][10][11] Within the rapidly developing field of tissue engineering, various biomaterial scaffolds have been developed for tissue regeneration or repair that mimic some of the critical properties of the ECM. 12 Examples of biomaterials that have been shown to support the proliferation and differentiation of salivary gland progenitors include chitosan, polyglycolic acid (PGA), poly-(L)-lactic acid (PLLA), poly (lactic-co-glycolic acid) (PLGA), and poly(ethylene glycol)terephthalate/poly(butylene)terephthalate (PEGT/PBT) block co-polymers. 6,8,13 However, these polymeric scaffolds have the potential to induce inflammation in vivo due to the acidity of their degradation products. 14,15 Another potential scaffold material, Matrigel, which contains basement membrane proteins secreted by EHS mouse sarcoma cells, has been used to grow primary salivary gland epithelial cells (pSGECs) in culture. ...
Salivary gland hypofunction often results from a number of causes, including the use of various medications, radiation for head and neck tumors, autoimmune diseases, diabetes, and aging. Since treatments for this condition are lacking and adult salivary glands have little regenerative capacity, there is a need for cell-based therapies to restore salivary gland function. Development of these treatment strategies requires the establishment of a system that is capable of replicating the salivary gland cell “niche” to support the proliferation and differentiation of salivary gland progenitor cells. In this study, a culture system using three-dimensional silk fibroin scaffolds (SFS) and primary salivary gland epithelial cells (pSGECs) from rat submandibular (SM) gland and parotid gland (PG) was established and characterized. pSGECs grown on SFS, but not tissue culture plastic (TCP), formed aggregates of cells with morphological features resembling secretory acini. High levels of amylase were released into the media by both cell types after extended periods in culture on SFS. Remarkably, cultures of PG-derived cells on SFS, but not SM cells, responded to isoproterenol, a β-adrenergic receptor agonist, with increased enzyme release. This behavior mimics that of the salivary glands in vivo. Decellularized extracellular matrix (ECM) formed by pSGECs in culture on SFS contained type IV collagen, a major component of the basement membrane. These results demonstrate that pSGECs grown on SFS, but not TCP, retain important functional and structural features of differentiated salivary glands and produce an ECM that mimics the native salivary gland cell niche. These results demonstrate that SFS has potential as a scaffold for creating the salivary gland cell niche in vitro and may provide an approach for inducing multipotent stem cells to provide therapeutically meaningful numbers of salivary gland progenitor cells for regenerating these tissues in patients.
... For example, previous studies have shown that it is difficult to maintain the same in vivo expression levels of amylase in cultured human salivary gland acinar cells without the addition of stimulants or the formation of 3D aggregates in vitro. 6,8,9 In order to circumvent these issues, it is important to mimic the in vivo cellular environment, whereby the salivary gland acinar cells are surrounded by a complex stromal environment, in which fibroblasts are the main cell type in proximity to the gland cells. Therefore, coculturing the salivary gland cells with fibroblasts in conditioned medium has allowed researchers to create artificial microenvironments mimicking the in vivo situation. ...
Background:
Artificial salivary gland replacement would be an ideal treatment for xerostomia. In vivo, salivary gland cells are surrounded by a complex stromal environment in which fibroblasts are the main cell type in proximity to the gland cells. However, very little is known about the relationship between these fibroblasts and the gland cells.
Methods:
Parotid gland acinar cells (PGACs) and fibroblasts from the same human gland were cocultured. PGAC function-related protein expression was investigated.
Results:
The expression of α-amylase in PGACs was increased in a fibroblast ratio-dependent manner. Both fibroblast-conditioned medium and direct coculture also significantly enhanced the PGAC expression of α-amylase. Basic fibroblast growth factor (bFGF) seems to be a regulator of α-amylase expression in PGACs.
Conclusion:
An appropriate number of fibroblasts in contact with the PGACs is necessary to promote PGAC function. Fibroblast-secreted bFGF may play a paracrine signaling role in the regulation of α-amylase expression in PGACs. © 2015 Wiley Periodicals, Inc. Head Neck, 2015.
... The use of 2D scaffold is however still possible to engineer salivary glands. 116 Primary human salivary gland cells seeded biodegradable polymer scaffolds showed the formation of postconfluence structure in vitro by enhancing E-cadherin expression 117 and acinar gland-like structure after 4 weeks of subcutaneous implantation in athymic mice. The gland-like structure also showed the production of human salivary aamylase (acinar cell protein). ...
The objective
of this review is to inform practitioners with the most updated information on tissue engineering and its potential applications in dentistry.
Data
The authors used “PUBMED” to find relevant literature written in English and published from the beginning of tissue engineering until today. A combination of key words was used as the search terms e.g., “tissue engineering”, “approaches”, “strategies” “dentistry”, “dental stem cells”, “dentino-pulp complex”, “guided tissue regeneration”, “whole tooth”, “TMJ”, “condyle”, “salivary glands”, and “oral mucosa”.
Sources
Abstracts and full text articles were used to identify causes of craniofacial tissue loss, different approaches for craniofacial reconstructions, how the tissue engineering emerges, different strategies of tissue engineering, biomaterials employed for this purpose, the major attempts to engineer different dental structures, finally challenges and future of tissue engineering in dentistry.
Study selection
Only those articles that dealt with the tissue engineering in dentistry were selected.
Conclusions
There has been a recent surge in guided tissue engineering methods to manage periodontal diseases beyond the traditional approaches. However, the predictable reconstruction of the innate organisation and function of whole teeth as well as their periodontal structures remains challenging. Despite some limited progress and minor successes, there remain distinct and important challenges in the development of reproducible and clinically safe approaches for oral tissue repair and regeneration. Clearly, there is a convincing body of evidence which confirms the need for this type of treatment, and public health data worldwide indicates a more than adequate patient resource. The future of these therapies involving more biological approaches and the use of dental tissue stem cells is promising and advancing. Also there may be a significant interest of their application and wider potential to treat disorders beyond the craniofacial region.
... Additionally, Matrigel is nonimplantable in humans. Other approaches have explored the use of flat synthetic and/or natural substrates to organize salivary gland cells into a monolayer [11][12][13]. Salivary gland cells grown on flat polymeric substrates often fail to assemble tight junctions, which are needed for directional secretion of saliva [14][15][16], and are unable to form a complex 3-D branching structure, a requirement for producing sufficient surface area to produce adequate amounts of saliva [17]. By adding physical complexity to substrates, it may be possible to achieve acinar cell organization and differentiation which more closely resembles cells grown in 3-D environments, while still achieving an interconnected monolayer of epithelial cells capable of directing saliva. ...
There is a need for an artificial salivary gland as a long-term remedy for patients suffering from salivary hypofunction, a leading cause of chronic xerostomia (dry mouth). Current salivary gland tissue engineering approaches are limited in that they either lack sufficient physical cues and surface area needed to facilitate epithelial cell differentiation, or they fail to provide a mechanism for assembling an interconnected branched network of cells. We have developed highly-ordered arrays of curved hemispherical "craters" in polydimethylsiloxane (PDMS) using wafer-level integrated circuit (IC) fabrication processes, and lined them with electrospun poly-lactic-co-glycolic acid (PLGA) nanofibers, designed to mimic the three-dimensional (3-D) in vivo architecture of the basement membrane surrounding spherical acini of salivary gland epithelial cells. These micropatterned scaffolds provide a method for engineering increased surface area and were additionally investigated for their ability to promote cell polarization. Two immortalized salivary gland cell lines (SIMS, ductal and Par-C10, acinar) were cultured on fibrous crater arrays of various radii and compared with those grown on flat PLGA nanofiber substrates, and in 3-D Matrigel. It was found that by increasing crater curvature, the average height of the cell monolayer of SIMS cells and to a lesser extent, Par-C10 cells, increased to a maximum similar to that seen in cells grown in 3-D Matrigel. Increasing curvature resulted in higher expression levels of tight junction protein occludin in both cell lines, but did not induce a change in expression of adherens junction protein E-cadherin. Additionally, increasing curvature promoted polarity of both cell lines, as a greater apical localization of occludin was seen in cells on substrates of higher curvature. Lastly, substrate curvature increased expression of the water channel protein aquaporin-5 (Aqp-5) in Par-C10 cells, suggesting that curved nanofiber substrates are more suitable for promoting differentiation of salivary gland cells.
Salivary gland (SG) dysfunction impairs the life quality of many patients, such as patients with radiation therapy for head and neck cancer and patients with Sjögren’s syndrome. Multiple SG engineering strategies have been considered for SG regeneration, repair, or whole organ replacement. An in-depth understanding of the development and differentiation of epithelial stem and progenitor cells niche during SG branching morphogenesis and signaling pathways involved in cell–cell communication constitute a prerequisite to the development of suitable bioengineering solutions. This review summarizes the essential bioengineering features to be considered to fabricate an engineered functional SG model using various cell types, biomaterials, active agents, and matrix fabrication methods. Furthermore, recent innovative and promising approaches to engineering SG models are described. Finally, this review discusses the different challenges and future perspectives in SG bioengineering.
