Optimizing decellularization protocols for human thyroid tissues: a step towards tissue engineering and transplantation.

Hypothyroidism is caused by insufficient stimulation or disruption of the thyroid. However, the drawbacks of thyroid transplantation have led to the search for new treatments. Decellularization allows tissue transplants to maintain their biomimetic structures while preserving cell adhesion, proliferation, and differentiation. This study aimed to decellularize human thyroid tissues using a structure-preserving optimization strategy and present preliminary data on recellularization. Nine methods were used for physical and chemical decellularization. Quantitative and immunohistochemical analyses were performed to investigate the DNA and extracellular matrix components of the tissues. Biomechanical properties were determined by compression test, and cell viability was examined after seeding MDA-T32 papillary thyroid cancer (PTC) cells onto the decellularized tissues. Decellularized tissues exhibited a notable decrease (< 50 ng/mg DNA, except for Group 2) compared to the native thyroid tissue. Nonetheless, collagen and glycosaminoglycans were shown to be conserved in all decellularized tissues. Laminin and fibronectin were preserved at comparatively higher levels, and Young's modulus was elevated when decellularization included SDS. It was observed that the strain value in Group 1 (1.63 ± 0.14 MPa) was significantly greater than that in the decellularized tissues between Groups 2-9, ranging from 0.13 ± 0.03 to 0.72 ± 0.29 MPa. Finally, viability assessment demonstrated that PTC cells within the recellularized tissue groups successfully attached to the 3D scaffolds and sustained metabolic activity throughout the incubation period. We successfully established a decellularization optimization for human thyroid tissues, which has potential applications in tissue engineering and transplantation research. Our next goal is to conduct recellularization using the methods utilized in Group 1 and transplant the primary thyroid follicular cell-seeded tissues into an in vivo animal model, particularly due to their remarkable 3D structural preservation and cell adhesion-promoting properties.

Computational drug repurposing for primary hyperparathyroidism.

In hyperparathyroidism (hyperPTH), excessive amounts of PTH are secreted, interfering with calcium regulation in the body. Several drugs can control the disease's side effects, but none of them is an alternative treatment to surgery. Therefore, new drug candidates are necessary. In this study, three computationally repositioned drugs, DG 041, IMD 0354, and cucurbitacin I, are evaluated in an in vitro model of hyperPTH. First, we integrated publicly available transcriptomics datasets to propose drug candidates. Using 3D spheroids derived from a single primary hyperPTH patient, we assessed their in vitro efficacy. None of the proposed drugs affected the viability of healthy cell control (HEK293) or overactive parathyroid cells at the level of toxicity. This behavior was attributed to the non-cancerous nature of the parathyroid cells, establishing the hyperPTH disease model. Cucurbitacin I and IMD 0354 exhibited a slight inverse relationship between increased drug concentrations and cell viability, whereas DG 041 increased viability. Based on these results, further studies are needed on the mechanism of action of the repurposed drugs, including determining the effects of these drugs on cellular PTH synthesis and secretion and on the metabolic pathways that regulate PTH secretion.

Combined approaches for detecting polypropylene microplastics in crop plants.

Microplastics (MPs) pollution in the terrestrial environment causes accumulation in crop plants. Consumption of these plants may have negative effects on human health. Therefore, it is crucial to analyze MPs accumulation in the plants. The aim of this study is to determine polypropylene (PP) particles in plants exposed to label-free PP for 75 days. In order to extract PP from organic matter, a two-step alkaline and wet peroxide oxidation chemical digestion method was applied to the roots, stems, and leaves of maize and wheat. The PP particles in the digested solutions were detected by the Nile red staining method, which has not been used previously in the detection of MPs in plants. Nile red stained PP particles mostly accumulated in the roots of wheat and the stems of maize plants. Statistical analysis revealed that the maize deposited more and larger PP particles regardless of the location. Moreover, the presence of PP particles in the digestion solutions was proved by the heating method. The PP particles on the glass slides were transformed into different shapes due to melting.

Production of parathyroid-like cells from thyroid stem cells in co-culture environment.

BACKGROUND: Parathyroid-like cells were aimed to be developed using cells isolated from thyroid since their embryological origins are the same. METHOD: Activin A and sonic hedgehog (Shh) are the proteins used in differentiation (dif) medium. Parathyroid and thyroid cells were cultured in a 3-dimensional environment and divided into five groups: thyroid standard (st) medium, thyroid dif medium, parathyroid st medium, thyroid-parathyroid co-culture st medium, and thyroid-parathyroid co-culture dif medium. Throughout 28 days of incubation, groups were investigated by carrying out the live dead assay, confocal microscopy, real-time PCR, immunohistochemistry and biochemical assays. RESULTS: Thyroid-parathyroid co-culture cells grown in dif medium exhibited upregulated expressions of parathormone (PTH) (5.1-fold), PTH1R (3.6-fold), calcium sensing receptor (CaSR) (8.8-fold), and loss of thyroid-specific thyroid transcription factor 1 (TTF1) expression when compared to the thyroid st medium group. PTH secretion decreased by 35% in the parathyroid st medium group and 99.9% in the thyroid-parathyroid co-culture st medium group but decreased only 3.5% in the thyroid-parathyroid co-culture dif medium group on day 28. CONCLUSION: Using Activin A and Shh proteins, thyroid stem/progenitor cells were differentiated to parathyroid-like cells successfully in a co-culture environment. A potentially effective novel method for cell differenatiation is co-culture of cells having the same embryological origin.

Chondro-inductive hyaluronic acid/chitosan coacervate-based scaffolds for cartilage tissue engineering.

Injuries related to articular cartilage are among the most challenging musculoskeletal problems because of poor repair capacity of this tissue. The lack of efficient treatments for chondral defects has stimulated research on cartilage tissue engineering applications combining porous biocompatible scaffolds with stem cells in the presence of external stimuli. This work presents the role of rat bone marrow mesenchymal stem cell (BMSC) encapsulated-novel three-dimensional (3D) coacervate scaffolds prepared through complex coacervation between different chitosan salts (CHI) and sodium hyaluronate (HA). The 3D architecture of BMSC encapsulated scaffolds (HA/CHI) was shown by scanning electron microscopy (SEM) to have an interconnected structure to allow cell-cell and cell-matrix interactions. Chondrogenic induction of encapsulated BMSCs within HA/CHI coacervates demonstrated remarkable cellular viability in addition to the elevated expression levels of chondrogenic markers such as sex determining region Y-box 9 protein (SOX9), aggrecan (ACAN), cartilage oligomeric matrix protein (COMP) and collagen type II (COL2A1) by immunofluorescence staining, qPCR and ELISA test. Collectively, HA/CHI coacervates are promising candidates for future use of these scaffolds in cartilage tissue engineering applications.

Hyaluronic Acid/Chitosan Coacervate-Based Scaffolds.

Chitosan-chloride (CHI) and sodium hyaluronate (HA), two semiflexible biopolymers, self-assemble to form nonstoichiometric coacervates. The effect of counterions was briefly investigated by preparing HA/CHI coacervates in either CaCl2 or NaCl solutions to find only a small difference in their tendency to coacervate. Higher water content in coacervates within CaCl2 was attributed to the chaotropic nature of Ca2+ ions. This effect was also evidenced with smaller pore sizes for coacervates in NaCl. Besides, for coacervation of chitosan-glutamate (CHI-G) with HA, dynamic light scattering at different charge ratios indicated a wider coacervation region for the HA/CHI-G pair than the HA/CHI. This was attributed to the chaotropic and "soft" ion nature of glutamate compared to chloride as a counterion of chitosan. Positive zeta potential values for both coacervate suspensions were explained by the contribution of charge mismatch, chain semiflexibility, and intra- and intercomplex disproportionation. Finally, HA/CHI coacervates were used to encapsulate bone marrow stem cells. While cell viabilities in HA/CHI coacervates were remarkable up to 21 days, their well-spread morphology has proved that HA/CHI coacervates are promising scaffolds for cartilage tissue engineering.