Identification of Critical Amino Acids of Coxsackievirus A10 Associated with Cell Tropism and Viral RNA Release during Uncoating.

Coxsackievirus A10 (CV-A10) is a prevailing causative agent of hand-foot-mouth disease, necessitating the isolation and adaptation of appropriate strains in cells allowed for human vaccine development. In this study, amino acid sequences of CV-A10 strains with different cell tropism on RD and Vero cells were compared. Various amino acids on the structural and non-structural proteins related to cell tropism were identified. The reverse genetic systems of several CV-A10 strains with RD+/Vero- and RD+/Vero+ cell tropism were developed, and a set of CV-A10 recombinants were produced. The binding, entry, uncoating, and proliferation steps in the life cycle of these viruses were evaluated. P1 replacement of CV-A10 strains with different cell tropism revealed the pivotal role of the structural proteins in cell tropism. Further, seven amino acid substitutions in VP2 and VP1 were introduced to further investigate their roles played in cell tropism. These mutations cooperated in the growth of CV-A10 in Vero cells. Particularly, the valine to isoleucine mutation at the position VP1-236 (V1236I) was found to significantly restrict viral uncoating in Vero cells. Co-immunoprecipitation assays showed that the release of viral RNA from the KREMEN1 receptor-binding virions was restricted in r0195-V1236I compared with the parental strain r0195 (a RD+/Vero+ strain). Overall, this study highlights the dominant effect of structural proteins in CV-A10 adaption in Vero cells and the importance of V1236 in viral uncoating, providing a foundation for the mechanism study of CV-A10 cell tropism, and facilitating the development of vaccine candidates.

[Comparison of mechanical properties of three machinable resin ceramic composite materials].

PURPOSE: This in vitro study was to compare the flexural properties, fracture toughness and hardness of three machinable composite materials. METHODS: Three kinds of resin composite ceramic Upcera Hyramic, 3M Lava Ultimate, Vita Enamic and a glass ceramic Vitablocs Mark II were chosen for the study. Bar-shaped specimens (16 mm×4 mm×1 mm, 2 mm) were prepared for flexural strength experiment; specimens (17 mm×4 mm×3 mm) were prepared for fracture toughness experiment and specimens of 4 mm thickness were prepared for hardness test. Flexural test and fracture toughness experiment were performed with an universal testing machine at a cross-head speed of 0.5 mm/min. Hardness test was performed with an micro hardness tester.Scanning electron microscope was used to observe the roughness of fracture surface. One-way variance analysis was used to determine the statistical differences with SPSS 17.0 software package. RESULTS: The mean flexural strength of the tested blocks at 1 mm thickness was Hyramic（207.7515±13.12）MPa＞Vita Enamic（182.0286±15.18）MPa＞Lava Ultimate（145.8469±8.98）MPa＞Vitablocs MarkⅡ（103.0542±18.19）MPa. The mean flexural modulus were Vitablocs MarkⅡ（49.49±5.50）GPa＞Vita Enamic（40.65±3.80）GPa＞Hyramic（14.89±2.38）GPa＞Lava Ultimate（7.09±1.24）GPa. The mean flexural strength of the tested blocks at 2 mm thickness was Hyramic（208.1986±25.07）MPa＞Lava Ultimate（172.9297±12.73）MPa＞Vitablocs MarkⅡ（158.6587±15.37） MPa＞Vita Enamic（155.3670±13.77）MPa. The mean flexural modulus were Vitablocs MarkⅡ（24.07±1.86）GPa＞Vita Enamic（19.64±0.98）GPa＞Hyramic（10.35±0.87）GPa＞Lava Ultimate（8.68±0.86）GPa. The mean fracture toughness was Vita Enamic（1.6357±0.16）MPa·m1/2＞Lava Ultimate（1.4286±0.11）MPa·m1/2>Vitablocs MarkII（1.3233±0.10）MPa·m1/2>Hyramic（1.0614±0.09）MPa·m1/2. The hardness of the experimental group was significantly lower than that of the control group. CONCLUSIONS: According to ISO 6872/2008, three kinds of machinable resin ceramic composites meet the needs of clinical strength.Hyramic showed higher flexural strength at different thickness, it is an ideal material for dental restoration. Vita Enamic has not only higher flexural strength at the thickness of 1 mm, but also good toughness, it is suitable for repair of patients that have limited occlusal space and great bite force, named occlusal veneer.