The nanocomposite scaffold of poly(lactide-co-glycolide) and hydroxyapatite surface-grafted with L-lactic acid oligomer for bone repair

Nanohydroxyapatite (op-HA) surface-modified with L-lactic acid oligomer (LAc oligomer) was prepared by LAc oligomer grafted onto the hydroxyapatite (HA) surface. The nanocomposite of op-HA/PLGA with different op-HA contents of 5, 10, 20 and 40 wt.% in the composite was fabricated into three-dimensional scaffolds by the melt-molding and particulate leaching methods. PLGA and the nanocomposite of HA/PLGA with 10 wt.% of ungrafted hydroxyapatite were used as the controls. The scaffolds were highly porous with evenly distributed and interconnected pore structures, and the porosity was around 90%. Besides the macropores of 100-300 mu m created by the leaching of NaCl particles, the micropores (1-50 mu m) in the pore walls increased with increasing content of op-HA in the composites of op-HA/PLGA. The op-HA particles could disperse more uniformly than those of pure HA in PLGA matrix. The 20 wt.% op-HA/PLGA sample exhibited the maximum mechanical strength, including bending strength (4.14 MPa) and compressive strength (2.31 MPa). The cell viability and the areas of the attached osteoblasts on the films of 10 wt.% op-HA/PLGA and 20 wt.% op-HA/PLGA were evidently higher than those on the other composites.

In Situ Scanning Tunneling Microscopy on the Dynamic Process of the Replacement of Coronene on Au (111) by 6-Mercapto-1-Hexanol

The replacement of coronene monolayer on Au (111) by 6-mercapto-1-hexanol (MHO) was studied by in situ scanning tunneling microscopy (STM) in solutions. It was found that the rate of replacement depends strongly on the concentration of MHO. The replacement finished within a second at a higher concentration of MHO. At a lower concentration, the slow replacement could be followed by in situ STM. The replacement occurred initially near the elbow position of reconstructed Au (111) with the formation of pits in a single or several missing molecules. With the proceeding of replacement, these small pits expanded, and the surrounding coronene molecules were gradually substituted by MHO, which developed into ordered domains within a spatial confined environment. Meanwhile, the reconstruction of Au (111) was lifted. The replacement expanded fast along the reconstruction lines in the domain. For the fast replacement, a (root 3 x root 3) R30 degrees adlattice was observed, while a c(4 x 2) superlattice was observed for the slow replacement.

Diluting thiol-derivatized oligonucleotide monolayers on Au(111) by mercaptohexanol replacement reaction

Surface replacement reaction of thiol-derivatized, single-stranded oligonucleotide (HS-ssDNA) by mercaptohexanol (MCH) is investigated in order to reduce surface density of the HS-ssDNA adsorbed to Au(111) surface. Cyclic voltammograms (CVs) and scanning tunneling microscopy (STM) are employed to assess the composition and state of these mixed monolayers. It is found that each CV of mixed self-assembled monolayers (SAMs) only shows a single reductive desorption peak, which suggests that the resulted, mixed SAMs do not form discernable phase-separated domains. The peak potential gradually shifts to negative direction and the peak area increases step by step over the whole replacement process. By analyzing these peak areas, it is concluded that two MCH molecules will replace one HS-ssDNA molecule and relative coverage can also be estimated as a function of exposing time. The possible mechanism of the replacement reaction is also proposed. The DNA surface density exponentially reduces with the exposing time increasing, in other words, the replacement reaction is very fast in the first several hours and then gradually slows down. Moreover, the morphological change in the process is also followed by STM.

Estimation of individual phases from the four-phase structure invariants in the single isomorphous replacement case

The probability distribution of the four-phase invariants in the case of single isomorphous replacement has been developed to estimate some individual phases. An example of its application to obtain the phases having special values of 0, pi or +/-pi /2 is given for a known protein structure in space group P2(1)2(1)2(1). The phasing procedure includes the determination of starting phases and an iterative calculation. The initial values of starting phases, which are required by the formula, can be obtained from the estimate of one-phase seminvariants and by specifying the origin and enantiomorph. In addition, the calculations lead to two sets of possible phases for each type of reflection by assigning arbitrarily an initial phase value. The present method provides a possibility for the multisolution technique to increase greatly the number of known phases while keeping the number of the trials quite small.

ESTIMATION OF THE ONE-PHASE STRUCTURE SEMINVARIANTS IN THE SINGLE ISOMORPHOUS REPLACEMENT CASE - AN APPLICATION OF HAUPTMANS DISTRIBUTION

A method for estimating the one-phase structure seminvariants (OPSSs) having values of 0 or pi has been proposed on the basis of the probabilistic theory of the three-phase structure invariants for a pair of isomorphous structures [Hauptman (1982). Acta Cryst. A38, 289-294]. The test calculations using error-free diffraction data of protein cytochrome c(550) and its PtCl42- derivative show that reliable estimates of a number of the OPSSs can be obtained. The reliability of the estimation increases with the increase of the differences between diffraction intensities of the native protein and its heavy-atom derivative. A means to estimate the parameters of the distribution from the diffraction ratio is suggested.

Behavioural responses by yaks in different physiological states (lactating, dry or replacement heifers), when grazing natural pasture in the spring (dry and germinating) season on the Qinghai-Tibetan plateau

Using heterogeneous vegetation in alpine grassland through grazing is a necessary component of deintensification of livestock systems and conservation of natural environments. However, better understanding of the dynamics of animal feeding behaviour would improve pasture and livestock grazing managements, particularly in the early part of the spring season when forage is scarce. The changes in behaviour may improve the use of poor pastures. Then, enhancing management practices may conserve pasture and improve animal productivity. Grazing behaviour over 24 In periods by yaks in different physiological states (lactating, dry and replacement heifers) was recorded in the early, dry and later, germinating period of the spring season. Under conditions of inadequate forage, the physiological state of yaks was not the primary factor affecting their grazing and ruminating behaviour. Forage and sward state affected yaks' grazing and ruminating behaviour to a greater extent. Generally, yaks had higher intake and spent more time grazing and ruminating during the later part of the spring season, following germination of forage, than during the earlier dry part of the season. However, the live weight of yaks was less during pasture germination than during the early dry part of the season because the herbage mass is low, and the yaks have to expend much energy to seek feed at this particular time. (c) 2007 Elsevier B.V. All rights reserved.

河流成因储层沉积微相与剩余油分布及形成机理研究

Based on the study of fluvial sandstone reservoir in upper of Guantao group in Gudao and Gudong oilfields, this paper first introduces A.D.Miall's(1996a) architectural-element analysis method that was summarized from ground outcrop scale into the reservoir formation research of the study area, more subtly divides sedimentary microfacies and establishes sedimentary model of research area.on this base, this paper summarizes the laws of residual oil distribution of fluvial formation and the control effect of sedimentary microfacies to residual oil distribution, and reveals residual oil formation mechanism. These results have been applied to residual oil production, and the economic effect is good. This paper will be useful for residual oil research and production and enhancement of oil recovery in similar reservoir. The major conclusions of this paper are as follows. 1. Using the architectural-element analysis method to the core data, a interfacial division scheme of the first to the dixth scale is established for the studied fluvial formation. 2.Seven architectural-elements are divided in upper of Guantao group of study area. The sandstone group 5～1+2 of Neogene upper Gutao group belongs to high sinuous fine grain meandering river, and the sandstone group 6 is sandy braided river. 3. Inter layer, the residual oil saturation of "non-main layer" is higher than "main layer", but the residual recoverable reserve of former is larger. Therefore, "main layer" is the main body of residual oil distribution. The upper and middle part of inner layer has lower permeability and strong seeping resistance. Addition to gravity effect in process of driving, its driving efficiency is low; residual oil saturation is high. Because of controlling of inside non-permeable interlayer or sedimentary construction, the residual oil saturation of non-driving or lower driving efficiency position also is high. On plane, the position of high residual oil saturation mostly is at element LV, CS, CH (FF), FF etc, Which has lower porosity and permeability, as well as lens sand-body and sand-body edge that is not controlled by well-net, non-perfect area of injection and production, lower press difference resort area of inter-well diffiuent-line and shelter from fault, local high position of small structure. 4.Microscopic residual oil mainly includes the non-moved oil in the structure of fine pore network, oil in fine pore and path, oil segment in pore and path vertical to flow direction, oil spot or oil film in big pore, residual oil in non-connective pore. 5.The most essential and internal controlling factor of fluvial formation residual oil distribution is sedimentary microfacies. Status of injection and production is the exterior controlling factor of residual oil distribution. 6. The controlling effect of formation sedimentary microfacies to residual oil distribution indicates inter-layer vertical sedimentary facies change in scale of injection and production layer-series, planar sedimentary face change and inner-layer vertical sedimentary rhythm and interbed in single layer to residual oil distribution. 7. It is difficult to clear up the inter-layer difference in scale of injection and production layer-series. The using status of minor layer is not good and its residual oil saturation is high relatively. It is obvious that inter-layer vertical sedimentary facies changes control inter-layer residual oil distribution at the same or similar conditions of injection and production. For fluvial formation, this vertical sedimentary facies change mainly is positive gyration. Namely, from down to top, channel sediment (element CHL, LA) changes into over-bank sediment (element LV, CR, CS). 8. In water-injection developing process of transverse connecting fluvial sandstone oil formation, injection water always comes into channel nearby, and breaks through along channel and orientation of high pressure gradient, does not expand into side of channel until pressure gradient of channel orientation changes into low. It brings about that water-driving status of over-bank sedimentary element formation (LV, CR, CS) is not good, residual oil saturation is high. In non-connective abandoned channel element (CH) formation with channel, because this reverse is difficult to control by injection and production well-series, its using status is not good, even terribly not good, residual oil is enrichment. 9. The rhythm and sedimentary structure, sedimentary facies change in single sand body brings about vertical changes of formation character, growth character of inner layer interbed. These are important factor of controlling and affecting vertical water spread volume and inner layer residual oil forming and distribution in single sand body. Positive rhythm, is the principal part of fluvial sandstone inner layer sedimentary rhythm. Namely, from down to upside, rock grain granularity changes from coarse to fine, seeping ability changes from strong to feebleness. It brings about that water-driving status of inner layer upside is not good, residual oil saturation is high. Inner layer interbed has different degree affecting and controlling effect to seeping of oil and water. Its affecting degree lies on interbed thickness, extending scale, position, and jeted segment of production or injection well. The effect of interbed at upside of oil formation to oil and water seeping is less; the effect of interbed at middle of oil formation to oil and water seeping is more. 10. Indoor experiment and research indicate that wettability, permeability step, vertical permeability, position of Kmax and ratio of oil viscousity and water viscousity all have great effect on the water-driving recovery ratio. 11. Microscopic residual oil distribution is affected and controlled by formation pore network structure, pressure field distribution, and oil characteristic. 12.The residual oil forming mechanism: the over-bank sedimentary element and upper part of a positive rhythm sandstone have fine pore and throat network, permeability is low, displacement pressure of pore and throat is high. The water-driving power usually falls short of displacement pressure that brings about injection water does not spread into these pore and throat network, thereby immovable oil area, namely residual oil, is formed. At underside of channel sedimentary element and positive rhythm sandstone, porosity and permeability is relatively high, connecting degree of pore and throat is high, displacement pressure of pore and throat is low. Thereby injection water is easy to enter into pore and throat, driving oil in them. Because the pore space is irregular, the surface of pore wall is coarse and non-flat. That the oil locate on concave hole of pore wall and the dead angle of pore, and the oil attaches on surface of pore wall by surface tension, are difficult to be peeled off, becoming water-driving residual oil (remaining oil). On the other hand, Because flowing section lessens, flowing resistance increase, action of capillary fore, or seeping speed decreases at process of transfer at pass narrow throat path in the course carried by driving water. The "oil drop", "oil bead", or "oil segment" peeled off by driving water is difficult to carry and to drive out by water at less pressure difference. Thereby they are enclosed in pore to form discontinuous residual oil. 13.This results described above have been applied in nine develop blocks of Gudao and Gudong oilfield. Its applying effect is marked through local injection production adjustment, deploying replacement well, repair hole, replacement envelop, block off water and profile control etc. Relative method and technology can be applied to other oil production area of Shengli oilfield, and obtain better economic and societal effect.

The use of transvaginal synthetic mesh for anterior vaginal wall prolapse repair: a randomized controlled trial

The aim of the study was to compare the efficacy and safety of transvaginal trocar-guided polypropylene mesh insertion with traditional colporrhaphy for treatment of anterior vaginal wall prolapse.This is a randomized controlled trial in which women with advanced anterior vaginal wall prolapse, at least stage II with Ba a parts per thousand yenaEuro parts per thousand+1 cm according to the Pelvic Organ Prolapse Quantification (POP-Q) classification, were randomly assigned to have either anterior colporrhaphy (n = 39) or repair using trocar-guided transvaginal mesh (n = 40). the primary outcome was objective cure rate of the anterior compartment (point Ba) assessed at the 12-month follow-up visit, with stages 0 and I defined as anatomical success. Secondary outcomes included quantification of other vaginal compartments (POP-Q points), comparison of quality of life by the prolapse quality of life (P-QOL) questionnaire, and complication rate between the groups after 1 year. Study power was fixed as 80 % with 5 % cutoff point (p < 0.05) for statistical significance.The groups were similar regarding demographic and clinical preoperative parameters. Anatomical success rates for colporrhaphy and repair with mesh placement groups were 56.4 vs 82.5 % (95 % confidence interval 0.068-0.54), respectively, and the difference between the groups was statistically significant (p = 0.018). Similar total complication rates were observed in both groups, with tape exposure observed in 5 % of the patients. There was a significant improvement in all P-QOL domains as a result of both procedures (p < 0.001), but they were not distinct between groups (p > 0.05).Trocar-guided transvaginal synthetic mesh for advanced anterior POP repair is associated with a higher anatomical success rate for the anterior compartment compared with traditional colporrhaphy. Quality of life equally improved after both techniques. However, the trial failed to detect differences in P-QOL scores and complication rates between the groups.

Valve surgery in active infective endocarditis: a simple score to predict in-hospital prognosis

Aims Surgery for infective endocarditis (IE) is associated with high mortality. Our objectives were to describe the experience with surgical treatment for IE in Spain, and to identify predictors of in-hospital mortality. Methods Prospective cohort of 1000 consecutive patients with IE. Data were collected in 26 Spanish hospitals. Results Surgery was performed in 437 patients (43.7%). Patients treated with surgery were younger and predominantly male. They presented fewer comorbid conditions and more often had negative blood cultures and heart failure. In-hospital mortality after surgery was lower than in the medical therapy group (24.3 vs 30.7%, p = 0.02). In patients treated with surgery, endocarditis involved a native valve in 267 patients (61.1%), a prosthetic valve in 122 (27.9%), and a pacemaker lead with no clear further valve involvement in 48 (11.0%). The most common aetiologies were Staphylococcus (186, 42.6%), Streptococcus (97, 22.2%), and Enterococcus (49, 11.2%). The main indications for surgery were heart failure and severe valve regurgitation. A risk score for in-hospital mortality was developed using 7 prognostic variables with a similar predictive value (OR between 1.7 and 2.3): PALSUSE: prosthetic valve, age ≥ 70, large intracardiac destruction, Staphylococcus spp, urgent surgery, sex [female], EuroSCORE ≥ 10. In-hospital mortality ranged from 0% in patients with a PALSUSE score of 0 to 45.4% in patients with PALSUSE score > 3. Conclusions The prognosis of IE surgery is highly variable. The PALSUSE score could help to identify patients with higher in-hospital mortality.

Engenharia de Tecidos na substituição de tecido ósseo

Projeto de Pós-Graduação/Dissertação apresentado à Universidade Fernando Pessoa como parte dos requisitos para obtenção do grau de Mestre em Ciências Farmacêuticas

Zinc finger nuclease gene repair as a treatment for cystinosis

Cystinosis is a multi-system autosomal recessive disorder caused by mutations and/or deletions in both alleles of CTNS, a gene encoding for the low pH dependent lysosomal cystine exporter cystinosin. Cystinosis occurs in approximately 1:200,000 newborns worldwide and is characterised by an accumulation of cystine in the lysosomes. The most severe form of the disorder is nephropathic cystinosis presenting Fanconi syndrome and leads without treatment to an end-stage renal failure before the age of ten. The only treatment available so far is cysteamine therapy, which delays disease progression by five years, but does not provide a cure for cystinosis patients. Current gene and cell based therapeutic approaches have not yet provided a suitable alternative. A potentially approach for a long-term treatment could be to generate autologous gene–modified stem cells by repairing the gene. Zinc Finger Nucleases (ZFNs) serve as a tool to increase HDR up to a 200,000-fold by introducing a double-stranded break (DSB). Thus, simple mutations in the CTNS gene could be corrected by introduction of a double-stranded break using ZFNs to boost the process of HDR with a suitable donor DNA sequence. A permanent repair of the most common lesion CTNS, a 57 kb deletion, could be achieved by ZFN-mediated HDR using a minigene CTNS promoter/cDNA construct. The thesis describes the design and testing of seven zinc finger nuclease pairs for their cleavage activity in vitro and in cellulo.. A highly sensitive assay to detect even low levels of ZFN-mediated HDR was also developed. Finally, to further investigate the role of autophagy in tissue injury in cystinotic cells an assay to monitor autophagy levels in the cells was successfully developed. This assay provides the opportunity to demonstrate functional restoration of CTNS after successful ZFN-HDR in cystinotic cells.

Cystic fibrosis gene repair: correction of ΔF508 using ZFN and CRISPR/Cas9 guide RNA gene editing tools

Cystic Fibrosis (CF) is an autosomal recessive monogenic disorder caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene with the ΔF508 mutation accounting for approximately 70% of all CF cases worldwide. This thesis investigates whether existing zinc finger nucleases designed in this lab and CRISPR/gRNAs designed in this thesis can mediate efficient homology-directed repair (HDR) with appropriate donor repair plasmids to correct CF-causing mutations in a CF cell line. Firstly, the most common mutation, ΔF508, was corrected using a pair of existing ZFNs, which cleave in intron 9, and the donor repair plasmid pITR-donor-XC, which contains the correct CTT sequence and two unique restriction sites. HDR was initially determined to be <1% but further analysis by next generation sequencing (NGS) revealed HDR occurred at a level of 2%. This relatively low level of repair was determined to be a consequence of distance from the cut site to the mutation and so rather than designing a new pair of ZFNs, the position of the existing intron 9 ZFNs was exploited and attempts made to correct >80% of CF-causing mutations. The ZFN cut site was used as the site for HDR of a mini-gene construct comprising exons 10-24 from CFTR cDNA (with appropriate splice acceptor and poly A sites) to allow production of full length corrected CFTR mRNA. Finally, the ability to cleave closer to the mutation and mediate repair of CFTR using the latest gene editing tool CRISPR/Cas9 was explored. Two CRISPR gRNAs were tested; CRISPR ex10 was shown to cleave at an efficiency of 15% and CRISPR in9 cleaved at 3%. Both CRISPR gRNAs mediated HDR with appropriate donor plasmids at a rate of ~1% as determined by NGS. This is the first evidence of CRISPR induced HDR in CF cell lines.