Clonal dynamics in early human embryogenesis inferred from somatic mutation.

Cellular dynamics and fate decision in early human embryogenesis remain largely unknown owing to the challenges of performing studies in human embryos1. Here, we explored whole-genomes of 334 single-cell colonies and targeted deep sequences of 379 bulk tissues obtained from various anatomical locations of seven recently deceased adult human donors. Using somatic mutations as an intrinsic barcode, we reconstructed early cellular phylogenies that demonstrate (1) an endogenous mutational rate that is higher in the first cell division but decreases to approximately one per cell per cell division later in life; (2) universal unequal contribution of early cells to embryo proper, resulting from early cellular bottlenecks that stochastically set aside epiblast cells within the embryo; (3) examples of varying degrees of early clonal imbalances between tissues on the left and right sides of the body, different germ layers and specific anatomical parts and organs; (4) emergence of a few ancestral cells that will substantially contribute to adult cell pools in blood and liver; and (5) presence of mitochondrial DNA heteroplasmy in the fertilized egg. Our approach also provides insights into the age-related mutational processes and loss of sex chromosomes in normal somatic cells. In sum, this study provides a foundation for future studies to complete cellular phylogenies in human embryogenesis.

Ultraviolet light screen using cholesteric liquid crystal capsules on the basis of selective reflection.

Sunscreen can protect human skin from sunlight by decreasing exposure to ultraviolet (UV) light, specifically UV-B and UV-A. In this study, a new type of UV screen system is proposed using cholesteric liquid crystal (CLC) capable of selectively reflecting UV-A within the human skin temperature range of 32-36 °C. Polycaprolactone (PCL) capsules with CLC mixture which had a helical chiral pitch corresponding to the wavelength of UV light were made by a solvent evaporation method. The average diameter of the capsules was about 34 µm. Consequently, it was confirmed that the CLC mixture (COC : CN = 80 : 20) could reflect UV-A light over 350-380 nm within the human skin temperature range. Also, it was confirmed that the CLC/PCL microcapsules could block UV light over 290-400 nm by about 6%.

Enhancement of osseointegration of artificial ligament by nano-hydroxyapatite and bone morphogenic protein-2 into the rabbit femur.

The MTT assay showed that the cell proliferation on hydroxyapatite (HAp) and HAp/bone morphogenic protein (BMP) coated group was better than the control and BMP coated groups at 5 days. And after 7 days of culture, the mRNA expression levels of type I collagen, osteonectin, osteopontin, bonesialoprotein, BMP-2, alkaline phosphatase (ALP) and Runx-2 in the HAp/BMP coated group were significantly higher than the other groups. Also, in this group showed the most significant induction of osteogenic gene expression compared to mesenchymal stem cells (MSCs) grown on the other groups. In addition, the cells in the HAp/BMP coated group delivered higher levels of ALP than the other three groups. Also, silk scaffolds were implanted as artificial ligaments in knees of rabbits, and they were harvested 1 and 3 months after implantation. On gross examination, HE staining showed that new bone tissue formation was more observed in the HAp/BMP coated group 3 weeks postoperatively. And masson staining showed that in the HAp/BMP coated group, the silk fibers were encircled by osteoblast, chondrocyte, and collagen. Furthermore, the analysis showed that the width of the graft-bone interface in the HAp and HAp/BMP coated group was narrower than that in the other two groups 3 weeks postoperatively. So, it is concluded that BMP incorporated HAp coated silk scaffold can be enhanced osseointegration and osteogenesis in bone tunnel. As a result, these experimental designs have been demonstrated to be effective in the acceleration of graft-to-bone healing by increasing new bone or fibrocartilage formation at the interface between graft and bone.

Macrolide resistance of Mycoplasma pneumoniae and its detection rate by real-time PCR in primary and tertiary care hospitals.

BACKGROUND: This study aimed to evaluate the prevalence of Mycoplasma pneumoniae in primary and tertiary care hospitals and its macrolide resistance rate. METHODS: Nasopharyngeal swabs were collected from 195 pediatric patients in primary and tertiary care hospitals from October to November 2010. The AccuPower MP real-time PCR kit (Bioneer, Korea) was used for the detection of M. pneumoniae. Direct amplicon sequencing was performed to detect point mutations conferring resistance to macrolides in the 23S rRNA gene. RESULTS: Among the 195 specimens, 17 (8.7%) were M. pneumoniae positive, and 3 of the strains (17.6%) obtained from these 17 specimens displayed the A2063G mutation in 23S rRNA. Three macrolide-resistant M. pneumoniae isolates were isolated from patients hospitalized at the primary care hospital. The positive rates of M. pneumoniae for the primary and tertiary care hospitals were 12.1% (15/124) and 2.8% (2/71), respectively (P=0.033). CONCLUSIONS: The positive rate of M. pneumoniae in the primary care hospital was higher than that in the tertiary care hospital. Simultaneous detection of M. pneumoniae and macrolide-resistant mutation genes in the 23S rRNA by real-time PCR is needed for rapid diagnosis and therapy of M. pneumoniae infections.

Two cases of Mycoplasma pneumoniae pneumonia with A2063G mutation in the 23S rRNA gene in siblings.

We describe 2 cases of pneumonia caused by the same macrolide-resistant Mycoplasma pneumoniae in siblings. M. pneumoniae was identified using real-time PCR. Direct sequence analysis of the 23S rRNA gene revealed a point mutation in V domain (A2063G) of the 23S rRNA gene.

Biochemical Characterization of an Extracellular ß-Glucosidase from the Fungus, Penicillium italicum, Isolated from Rotten Citrus Peel.

A ß-glucosidase from Penicillium italicum was purified with a specific activity of 61.8 U/mg, using a chromatography system. The native form of the enzyme was an 88.5-kDa tetramer with a molecular mass of 354 kDa. Optimum activity was observed at pH 4.5 and 60℃, and the half-lives were 1,737, 330, 34, and 1 hr at 50, 55, 60, and 65℃, respectively. Its activity was inhibited by 47% by 5 mM Ni(2+). The enzyme exhibited hydrolytic activity for p-nitrophenyl-ß-D-glucopyranoside (pNP-Glu), p-nitrophenyl-ß-D-cellobioside, p-nitrophenyl-ß-D-xyloside, and cellobiose, however, no activity was observed for p-nitrophenyl-ß-D-lactopyranoside, p-nitrophenyl-ß-D-galactopyranoside, carboxymetyl cellulose, xylan, and cellulose, indicating that the enzyme was a ß-glucosidase. The k(cat)/K(m) (s(-1) mM(-1)) values for pNP-Glu and cellobiose were 15,770.4 mM and 6,361.4 mM, respectively. These values were the highest reported for ß-glucosidases. Non-competitive inhibition of the enzyme by both glucose (K(i) = 8.9 mM) and glucono-δ-lactone (K(i) = 11.3 mM) was observed when pNP-Glu was used as the substrate. This is the first report of non-competitive inhibition of ß-glucosidase by glucose and glucono-δ-lactone.

Increase of BM-MSC proliferation using L-DOPA on titanium surface in vitro.

In order to increase biocompatibility, many dental implants have been studied by immobilization of biomolecules on biomaterials. We used l-3,4-dihydroxyphenylalanine (L-DOPA) as a biomolecule for surface-modified titanium. Water contact angles of nontreated titanium discs (negative control), etched titanium discs (positive control), and titanium discs treated with L-DOPA following the etching process (experimental group) were 82.4 ± 5.7°, 67.1 ± 0.56°, and 44.15 ± 0.91°, respectively. Using atomic force microscopy images, we were able to find L-DOPA, which adhered to the titanium surface. The number of human bone marrow mesenchymal stem cells (BM-MSCs) in the experimental group was much higher than that of cells in any other group. Quantification values of amine groups in the positive control and experimental groups were approximately 3 and 7.5 µg, respectively. Therefore, findings from our research suggested the possibility of a causal link between increased L-DOPA content and cell proliferation in BM-MSCs. Moreover, coating of the discs with L-DOPA can result in greater hydrophilicity of the titanium surface and enhancement of cell adhesion and mitochondrial activity.