BackgroundThis study aimed to investigate the effects of vitrification and slow freezing on actin, tubulin, and nuclei of in vivo preimplantation murine embryos at various developmental stages using a Confocal Laser Scanning Microscope (CLSM).Material/MethodsFifty female mice, aged 4–6 weeks, were used in this study. Animals were superovulated, cohabitated overnight, and sacrificed. Fallopian tubes were excised and flushed. Embryos at the 2-cell stage were collected and cultured to obtain 4- and 8-cell stages before being cryopreserved using vitrification and slow freezing. Fixed embryos were stained with fluorescence-labelled antibodies against actin and tubulin, as well as DAPI for staining the nucleus. Labelled embryos were scanned using CLSM and images were analyzed with Q-Win software V3.ResultsThe fluorescence intensity of both vitrified and slow-frozen embryos was significantly lower for tubulin, actin, and nucleus as compared to non-cryopreserved embryos (p<0.001). Intensities of tubulin, actin, and nucleus in each stage were also decreased in vitrified and slow-frozen groups as compared to non-cryopreserved embryos.ConclusionsCryopreservation of mouse embryos by slow freezing had a more detrimental effect on the actin, tubulin, and nucleus structure of the embryos compared to vitrification. Vitrification is therefore superior to slow freezing in terms of embryonic cryotolerance.

Eight human thyroid cancers (four papillary, and four lymph node metastases of papillary cancers) were studied at the ultrastructural level. The most characteristic anomalies affect the nucleus: "ground glass nuclei", highly indented nuclear membrane with formation of nuclear pseudoinclusions, nuclear bodies probably of nucleolar origin, fractionation of the periphery of the nucleus into multiple lobes joined by thin bridges of nuclear substance. Other abnormalities related to the mitochondria with decrease of size and number of cristae, the rough endoplasmic reticulum which was segregated in parallel saccules and the basal lamina was often reduplicated. The significance of these anomalies is discussed.

Morphological and phenotypical signs of cultured readaptation osteoblasts were studied after a short-term space mission. The ultrastructure and phenotype of human osteoblasts after Soyuz TMA-11 space flight (2007) were evaluated by scanning electron microscopy, laser confocal microscopy, and ELISA. The morphofunctional changes in cell cultures persisted after 12 passages. Osteoblasts retained the drastic changes in their shape and size, contour deformation, disorganization of the microtubular network, redistribution of organelles and specialized structures of the plasmalemma in comparison with the ground control cells. On the other hand, the expression of osteoprotegerin and osteocalcin (bone metabolism markers) increased; the expression of bone resorption markers ICAM-1 and IL-6 also increased, while the expression of VCAM-1 decreased. Hence, space flight led to the development of persistent shifts in cultured osteoblasts indicating injuries to the cytoskeleton and the phenotype changes, indicating modulation of bone metabolism biomarkers.

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