Background: This study aims to investigate the impact of test conditions on the results of the compression testing of microneedle arrays (MNAs). Methods: Uniaxial compression tests were conducted on polyglycolic acid-fabricated biodegradable MNAs. Load–displacement curves were obtained for varying conditions, including the number of microneedles (MNs) compressed simultaneously, compression speeds, and compression angles. Subsequently, the buckling load and stiffness were calculated, and the MN deformation during compression was observed. Results: The buckling load and stiffness per MN decreased significantly with a simultaneous increase in compressed MNs. The mean buckling load and stiffness of 52 MNs in single-needle compression tests were 0.211 ± 0.008 N and 13.9 ± 1.3 N/mm, respectively, with no variation among the three MNAs. However, a significant difference in buckling load and stiffness was observed among the MNs within the MNAs. Additionally, buckling loads and stiffnesses were significantly lower in certain MNs at the same location in different MNAs. Buckling load and stiffness decreased significantly during inclined compression compared to during vertical compression. While the tests evaluate the mechanical properties of MNAs, test results may vary depending on test conditions. Conclusions: Compression testing of the individual MNs comprising an MNA helps evaluate the mechanical properties of MNs and ensure the quality of MNAs.

Multipotent mesenchymal stem cells (MSCs) are widespread in adult organisms and are implicated in tissue maintenance and repair, regulation of hematopoiesis, and immunologic responses. Human (h)MSCs have applications in tissue engineering, cell-based therapy, and medical devices but it is unclear how they respond to unfavorable conditions, such as hypoxia or inflammation after transplantation in vivo. Although endotoxin testing is required for evaluating the quality and safety of transplanted MSCs, no reports on their dose response to endotoxins are available to establish the limits for in vitro MSC culture systems. In the present study, we aimed to accurately quantify the risk of endotoxin contamination in cell culture systems to establish an acceptable endotoxin limit for the differentiation of hMSC osteoblasts and adipocytes. Three types of bone marrow-derived hMSCs (hMSC-1: 21-year-old, M/B; hMSC-2: 36-year-old, M/B; hMSC-3: 43-year-old, M/C) and adipose-derived stem cells (ADSCs; StemPro Human) were cultured in osteogenic or adipogenic differentiation media, respectively, from commercial kits, containing various concentrations of endotoxin (0.01-100 ng/ml). The degree of adipocyte and osteoblast differentiation was estimated by fluorescent staining of lipid droplets and hydroxyapatite, respectively. To clarify the molecular mechanism underlying the effect of endotoxin on hMSC differentiation, cellular proteins were extracted from cultured cells and subjected to liquid chromatograph-tandem mass spectrometry shotgun proteomics analysis. Although endotoxin did not effect the adipocyte differentiation of hMSCs, osteoblast differentiation was enhanced by various endotoxin concentrations: over 1 ng/ml, for hMSC-1; 10 ng/ml, for hMSC-2; and 100 ng/ml, for hMSC-3. Proteomic analysis of hMSC-1 cells revealed up-regulation of many proteins related to bone formation. These results suggested that endotoxin enhances the osteoblast differentiation of MSCs depending on the cell type. Since endotoxins can affect various cellular functions, an endotoxin limit should be established for in vitro MSC cultures. Its no-observed-adverse-effect level was 0.1 ng/ml based on the effect on the hMSC osteoblast differentiation, but it may not necessarily be the limit for ADSCs.

To develop a high-affinity aptamer against AML1 Runt domain, two aptamers were conjugated based on their structural information. The newly designed aptamer Apt14 was generated by the conjugation of two RNA aptamers (Apt1 and Apt4) obtained by SELEX against AML1 Runt domain, resulting in improvement in its binding performance. The residues of AML1 Runt domain in contact with Apt14 were predicted in silico and confirmed by mutation and NMR analyses. It was suggested that the conjugated internal loop renders additional contacts and is responsible for the enhancement in the binding affinity. Conjugation of two aptamers that bind to different sites of the target protein is a facile and robust strategy to develop an aptamer with higher performance.

In vivo skin sensitization tests are required to evaluate the biological safety of medical devices in contact with living organisms to provide safe medical care to patients. Negative and positive reference materials have been developed for biological tests of cytotoxicity, implantation, hemolysis, and in vitro skin irritation. However, skin sensitization tests are lacking. In this study, polyurethane sheets containing 1 wt/wt % 2,4‐dinitrochlorobenzene (DNCB‐PU) were developed and evaluated as a positive reference material for skin sensitization tests. DNCB‐PU sheet extracts prepared with sesame oil elicited positive sensitization responses for in vivo sensitization potential in the guinea pig maximization test and the local lymph node assay. Furthermore, DNCB‐PU sheet extracts prepared with water and acetonitrile, 10% fetal bovine serum‐containing medium, or sesame oil elicited positive sensitization responses as alternatives to animal testing based on the amino acid derivative reactivity assay, human cell line activation test, and epidermal sensitization assay, respectively. These data suggest that the DNCB‐PU sheet is an effective extractable positive reference material for in vivo and in vitro skin sensitization testing in medical devices. The formulation of this reference material will lead to the development of safer medical devices that contribute to patient safety.