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Biomedical Engineering Education

Biomedical Engineering Education - Poster Session B

Poster E10 - Design of microfluidic device with silk constructs that replicate the progression of periodontal disease.

Thursday, October 24, 2024
2:45 PM - 3:45 PM EST
Location: Exhibit Hall E, F & G

Introduction:  : Emerging evidence highlights a strong correlation between periodontal conditions, such as gingivitis or periodontitis, and systemic diseases (i.e., cardiovascular disease, diabetes, and respiratory infections); this relationship is primarily due to an unbalanced microbiome originating from the oral cavity. A critical indicator of periodontal disease is the depth of this sulcus, ranging from 2 mm in healthy gums to up to 8 mm in severe periodontitis.
Clinical investigations have shown that during periodontal disease, the cyto-anatomical architecture of the gingival sulcus, which is the ecological niche of the oral microbiota, is compromised due to a dysbiotic microbiota and exacerbated inflammation driven by immune cells (neutrophils). Disrupting this balance hampers the existence of commensal bacteria and facilitates the growth of disease-causing pathogens. Understanding host-microbiome-immune interactions is crucial, as an imbalanced oral microbiome can drive systemic inflammation and disease.
Yet knowledge about microbiome-immune interactions in the healthy gingival sulcus is still limited, and the mechanisms leading to disease progression are even less understood. Conventional methods for studying periodontal conditions cannot often capture the complex interactions within the gingival sulcus, focusing mainly on individual pathogens rather than polymicrobial communities and their interactions with the host's immune system. Therefore, an advanced system is needed to fill these gaps in understanding the oral microbiota-immune relationship in the gingival sulcus. This project aims to develop a 3D microfluidic device that mimics a tooth-entity unit, incorporating immune cells, human subgingival plaque samples, and artificial saliva to observe inflammatory marker modulation and humanized responses in periodontal disease progression.

Materials and

Methods:  To understand periodontal disease development, silk fibroin constructs of 8, 4, and 2 mm were created to replicate oral clinical manifestations. First, the silk fibroin was processed as stipulated by the protocol (Rockwood, D. N.; Preda, R. C.; Yucel, T.; Wang, X. Q.; Lovett, M. L.; Kaplan, D. L., Materials fabrication from Bombyx mori silk fibroin. Nat. Protoc. 2011). (Fig. 1-i). SolidWorks was used to design molds mimicking the gingival sulcus depth, which were then 3D printed in SLA resin. These resin molds were used to create polydimethylsiloxane (PDMS) molds, as PDMS is ideal for molding and compatible with silk manufacturing. The top and base molds ensured the abutments were hollow (Fig. 1-ii). The 4% silk fibroin solution was poured into the assembled PDMS molds and freeze-dried to create a porous structure. Finally, the constructs were autoclaved at 121°C for 1 hour to achieve insolubility and sterility (Fig. 1-iii). Structural-mechanical characterization included compression tests and porosity analysis (Fig. 1-iv). For compression tests, 3-mm freeze-dried silk constructs were prepared. Using the UniVert system (CellScale biomaterials testing), compression tests were performed in displacement control mode at 0.01 mm/s. Samples were compressed up to 80% of their original height, identified as the failure point. For porosity analysis, 3-mm biopsies of lyophilized constructs were immersed in a 0.05% crystal violet solution (Sigma Aldrich) and imaged with a fluorescence microscope. Images were processed using MATLAB software to determine the equivalent radius and area of each pore.

Results, Conclusions, and Discussions: To generate the silk constructs, prototypes were designed using SOLIDWORKS, including both the top and the base, as shown in Fig. 2 (i) and (ii). The prototypes aimed to produce impressions with five 8 mm high pillars for both parts (top and base), with lengths of 55 mm for the base part and 52 mm for the top part. Once the 3D printed parts were produced using SLA, their dimensions were measured, and the height of the pillars (8mm) and the lengths of both parts were verified to be as stipulated. In addition, consistent resin quality was obtained in all prints. These were then used to create PDMS molds for the base and top part, as illustrated in Fig. 2 (iii) and (iv), to generate 8 mm deep holes in the base mold and 8 mm long pillars in the top part. Molds were obtained with dimensions of 58 mm in length, 15 mm in width for the base part mold, and 55 mm long for the top part mold. These were then assembled to insert silk between them and create the hollow silk construction. Following the procedures described in the Materials and Methods section, the resulting silk constructions were hollow structures as shown in Fig. 2 (v), with consistent measurements of 8 mm in height for each of the five pillars, a width of 1 mm for each pillar, and a total construction length of 55 mm.

For mechanical characterization, biopsies of lyophilized silk constructs of 2 mm thickness were used for porosity and compression testing. Graph 1(i) shows the compression test results plotted as strain (%) versus stress (kPa), giving a compression modulus of 4.5 kPa (Graph 1(ii)), within the range of mammalian soft tissues (0.1-100 kPa). Porosity testing, focusing on open porosity essential for optimal cell maintenance, used an inverted microscope to capture images analyzed via MATLAB. The analysis converted original images to grayscale and then to porous segmentation images, culminating in a porosity percentage of 62.33%. This high porosity supports cellular activity, confirming the construct's quality for studying cellular behavior in periodontal disease progression

Acknowledgements (Optional):  This work was conducted at the University of Massachusetts Lowell’s. REU Site: Inclusion and Innovation in Medical Devices, funded by the Science Foundation (Award #2243722).
C.E. Ghezzi, A.R. Diaz, M. Adelfio and G.E Cullen (Umass Lowell) for their guidance, contributions and support for this research.