The use of autologous platelet-rich plasma (platelet gel) and autologous platelet-poor plasma (fibrin glue) in cosmetic surgery

The purpose of this study was to evaluate a new technique of harvesting and preparing autologous platelet gel and autologous fibrin glue (body glue) and to evaluate their effectiveness in stopping capillary bleeding in the surgical flaps of patients undergoing cosmetic surgery. A convenience sample of 20 patients ranging from 25 to 76 years of age undergoing cosmetic surgery involving the creation of a surgical flap were included in the study. The types of surgical procedures included face lifts, breast augmentations, breast reductions, and neck lifts. Platelet-poor and platelet-rich plasma were prepared during the procedure from autologous blood using a compact, tabletop, automated autologous platelet concentrate system (SmartPReP, Harvest Autologous Hemobiologics, Norwell, Mass.). The platelet-poor and platelet-rich plasma were combined with a thrombin-calcium chloride solution to produce autologous fibrin glue and autologous platelet gel, respectively. Capillary bed bleeding was present in all cases and effectively sealed within 3 minutes following the application of platelet gel and fibrin glue. The technique for making the solution and for evaluating its effectiveness in achieving and maintaining hemostasis during cosmetic surgical procedures is described. Autologous platelet gel and fibrin glue prepared by the automated concentrate system are compared with autotransfusor-prepared platelet gel and Tisseel (Baxter Healthcare Corp.), a commercially prepared fibrin sealant preparation.     

Effectiveness of evaluating bulls according to their progeny by the "daughters-mothers" and "daughters-peers" methods

The repeatability coefficient of sire evaluation by daughters performance in different lactations was 0.38-0.70, that suggests the necessity to specify annually the previous sire evaluation on receiving new daughters performance data. When using the "daughters-mothers" method the repeatability coefficients of sire evaluation by dairy indexes, milk fat content and total milk fat production (0.41, 0.52 and 0.45 respectively) were significantly higher than those obtained by the "daughters-herdmates" method (0.25, 0.30 and 0.28 respectively). Thus, sire progeny testing evaluation should be carried out simultaneously by both methods. Nevertheless, the "daughters-mothers" method is preferable.     

The "echis carinatus venom" prothrombin assay in coumarin treated patients. A comparison with one-stage and immunological assays.

Prothrombin (factor II) was assayed in a group of coumarin treated patients using the Echis carinatus venom as thromboplastin. The levels obtained were comparable to those observed using the classical one-stage method. A good correlation was in fact observed between the two methods. The levels observed by the Echis carinatus method were definitely lower than those obtained using two immunological methods indicating that Echis carinatus venom activated, in our system, only normal prothrombin. However, even the levels obtained immunologically were slightly decreased, regardless of the method used, as compared to pooled normal plasma. In congenital prothrombin deficiency (homozygotes and heterozygotes) the level obtained by the Echis carinatus method was comparable to that observed by the one-stage method. On the contrary, in a congenital dysprothrombinemia (prothrombin Padua) a normal level was observed whereas the one-stage and two-stage methods yielded constantly levels of about 50% of normal.     

Interhemispheric asymmetry of "emotional resonance" in the rat

Hemispheric asymmetry of "emotional resonance" elaborated by the method of P.V. Simonov was studied in Wistar rats. Inactivation of hemispheres was carried out by means of spreading depression. When using as "victims" and "recipients" the animals of the same sex, lateralization of emotional resonance was found to depend upon the velocity of reaction elaboration. In rats rapidly elaborating avoidance reaction the right hemisphere dominated during its performance and so did the left one in animals learning after additional training. When using the animals of different sex as "victims" and "recipients", the right hemisphere dominated in "emotional resonance" performance. Hemispheric asymmetry of "emotional" resonance was more expressed in males than in females.     

Human body mass estimation: a comparison of "morphometric" and "mechanical" methods

In the past, body mass was reconstructed from hominin skeletal remains using both "mechanical" methods which rely on the support of body mass by weight-bearing skeletal elements, and "morphometric" methods which reconstruct body mass through direct assessment of body size and shape. A previous comparison of two such techniques, using femoral head breadth (mechanical) and stature and bi-iliac breadth (morphometric), indicated a good general correspondence between them (Ruff et al. [1997] Nature 387:173-176). However, the two techniques were never systematically compared across a large group of modern humans of diverse body form. This study incorporates skeletal measures taken from 1,173 Holocene adult individuals, representing diverse geographic origins, body sizes, and body shapes. Femoral head breadth, bi-iliac breadth (after pelvic rearticulation), and long bone lengths were measured on each individual. Statures were estimated from long bone lengths using appropriate reference samples. Body masses were calculated using three available femoral head breadth (FH) formulae and the stature/bi-iliac breadth (STBIB) formula, and compared. All methods yielded similar results. Correlations between FH estimates and STBIB estimates are 0.74-0.81. Slight differences in results between the three FH estimates can be attributed to sampling differences in the original reference samples, and in particular, the body-size ranges included in those samples. There is no evidence for systematic differences in results due to differences in body proportions. Since the STBIB method was validated on other samples, and the FH methods produced similar estimates, this argues that either may be applied to skeletal remains with some confidence.     

Forceful large-scale expression of "problematic" membrane proteins

We developed an Escherichia coli expression system for overproduction of a highly toxic membrane protein that is impossible to overexpress by traditionally used approaches. The method is based on combination of the genetic modifications of a bicistronic expression plasmid, stabilization of a synthesized protein, and selection of a compatible expression host. This enabled us to enhance the expression level of a toxic membrane protein 30-50 times compared with expression in the native state and to obtain 3-5mg of a highly purified functionally active protein per liter of culture. We describe the method for the amplified expression of membrane proteins, using the Pseudomonas aeruginosa multidrug resistance protein, MexY, as an example. The amplified MexY was correctly folded in the cytoplasmic membrane of the E. coli without forming inclusion bodies. This method can be applicable to the large-scale expression of the other problematic membrane proteins that are otherwise extremely difficult to overproduce.     

Estimation of guanine deaminase using guanosine as a "prosubstrate"

Plasma guanine deaminase (guanase; GD) is well established as an indicator of hepatocellular disease, recently being applied in the detection of hepatitis C in donor blood and in the diagnosis of hepatoma. No totally efficient, simple method for the estimation of plasma GD activity is routine since both guanine and 8-azaguanine, the substrates of the enzyme, are scarcely soluble in water. This difficulty in preparing stable substrates of sufficient concentration has resulted in methods that are both troublesome and inaccurate. Here we describe the development of new colorimetric and high-performance liquid chromatography (HPLC) methods utilizing guanosine as a "prosubstrate." After an initial breakdown of the guanosine to guanine using purine nucleoside phosphorylase, the ammonia formed as a result of the breakdown of the guanine by GD was estimated colorimetrically by the Berthelot reaction. As an alternative or a complementary assay, the xanthine also formed was measured using an isocratic HPLC method. These methods are suitable for routine assays for measuring plasma GD over a wide range of activities.     

Nondestructive quantification of neutral lipids by thin-layer chromatography and laser-fluorescent scanning: suitable methods for "lipidome" analysis

A simple method for separation and quantification of neutral lipids was developed using thin-layer chromatography (TLC) and high-performance fluorescent scanning. Neutral lipid classes were separated using the double-developing TLC method and detected by rhodamine 6G and a laser-excited fluorescent scanner. The amount of lipids applied correlated with scanned intensity volume in a dose-dependent manner. The mass of each neutral lipid band was determined by comparing band intensities of unknown samples to dilution curves of authentic standards. After scanning the dye-sprayed TLC, acyl chain species of triglyceride (TG) extracted from TLC could be determined by gas chromatography. Using this method, we quantified the amounts of TG in mouse liver and found that the measured total mass of TG correlated with that obtained by enzymatic methods. Our method should provide the basic technique for "lipidome" analysis, designed to determine and compare total lipid classes and mass present in biological samples.     

"Cystope tagging" for labeling and detection of recombinant protein expression

A labeling and detection method, based on the addition of a single cysteine residue at the C terminus of a recombinant protein and the subsequent sulfhydryl-specific Michael addition to the double bond of maleimide and its derivatives, was developed. The method was named "cystope tagging." Sorbit dehydrogenase (SDH) from Rhodobacter sphaeroides, a member of the short-chain dehydrogenase family of proteins that contains three inherent cysteines, was used as a model recombinant protein. By labeling with fluorescein-maleimide, it was demonstrated that only the single accessory cysteine is accessible under nonreducing conditions. After the addition of beta-mercaptoethanol, the inherent cysteines of SDH were also detectable by coupling to fluorescein-maleimide. The data were obtained using Autodisplay, an efficient surface expression system in Escherichia coli, but the method presented in this article represents a rather general solution for analyzing the expression of recombinant proteins, irrespective of the expression system used. The authors conclude that cystope tagging is an interesting alternative to other tagging methods applied in recombinant protein techniques.     

A new approach to gene mutation analysis using "GFP-Display"

The unique behavior of green fluorescent protein (GFP) on SDS-PAGE was applied to the detection of a single amino acid substitution in GFP-tagged polypeptides. This simple detection method using SDS/urea gels was designated GFP-display. The N-terminal 18 or 37 amino acids of K-Ras was used as a model GFP-tagged polypeptide. K-ras exon 1 was fused to a gfp cDNA at each end and expressed in Escherichia coli. Amino acid number 12 of K-Ras (wild type; Gly) was changed to Ser, Arg, Cys, Asp, Ala, or Val, and the mobility shift of the greenish fluorescent bands in the SDS/urea gel was analyzed. These mutants were easily detected by GFP-display; however, detection depended strongly on the urea concentration and electrophoresis temperature. Subsequently, GFP-display was applied to the 36 amino acids encoding human p53 exon 7. Amino acid number 248 (wild type; Arg) was changed to Gly, Trp, Gln, Pro, or Leu, and similar mobility shifts were observed. GFP-display could be coupled with an in vitro translation system. Fluorescent active GFP and GFP-Ras fusion proteins were synthesized within a few hours. GFP-display shows potential as a modern approach to gene mutation analysis at the protein level, and is a useful method for protein engineering studies.     

"Reverse ecology" and the power of population genomics

Rapid and inexpensive sequencing technologies are making it possible to collect whole genome sequence data on multiple individuals from a population. This type of data can be used to quickly identify genes that control important ecological and evolutionary phenotypes by finding the targets of adaptive natural selection, and we therefore refer to such approaches as "reverse ecology." To quantify the power gained in detecting positive selection using population genomic data, we compare three statistical methods for identifying targets of selection: the McDonald-Kreitman test, the mkprf method, and a likelihood implementation for detecting d(N)/d(S) > 1. Because the first two methods use polymorphism data we expect them to have more power to detect selection. However, when applied to population genomic datasets from human, fly, and yeast, the tests using polymorphism data were actually weaker in two of the three datasets. We explore reasons why the simpler comparative method has identified more genes under selection, and suggest that the different methods may really be detecting different signals from the same sequence data. Finally, we find several statistical anomalies associated with the mkprf method, including an almost linear dependence between the number of positively selected genes identified and the prior distributions used. We conclude that interpreting the results produced by this method should be done with some caution.     

Evolution of recombination in "host-parasite" systems. Multilocus models

Evolution of the recombination system caused by antagonistic species interactions (host and parasite, for example) was studied. The genetic structure of host population as well as that of parasite is explicitly present in our models. The selection intensity depends on host's resistance and parasite's virulence, both controlled by polygenic systems. The rec-system dynamics was numerically studied using the genetic operators method. It is shown that high-recombination alleles of the rec-modificator can have a short-term selective advantage in both interacting populations simultaneously.     

Determining dual Euler angles of the ankle complex in vivo using "flock of birds" electromagnetic tracking device

The dual Euler angles method has been proposed as an alternative approach to describe the general spatial human joint motion. In this study, the dual Euler angles method was applied to study the three-dimensional motion of the ankle complex. The methodology for obtaining dual Euler angles of the ankle complex was developed by using a "Flock of Birds" electromagnetic tracking device. The repeatability of the methodology was studied based on the intertester and intratester variability analysis. Finally kinematic coupling characteristics of the ankle complex during dorsiflexion-plantarflexion, eversion-inversion, and abduction-adduction were analyzed according to the parameters of the dual Euler angles.     

Cellulose in algal cell wall: an "in situ" localization.

In order to ascertain the presence and the in muro localization of cellulose, the enzyme-gold affinity test was applied to algal cell walls. The high specificity of affinity cytochemistry allowed us, by using the enzyme cellulase, to confirm the available biochemical data and to give a map of the cellulose localization in different algal groups. Taking into account the complex skeletal polysaccharide structure and composition of the algal cell walls, this method proved to be a reliable tool in this field.     

Measuring "free" iron levels in Caenorhabditis elegans using low-temperature Fe(III) electron paramagnetic resonance spectroscopy

Oxidative stress, caused by free radicals within the body, has been associated with the process of aging and many human diseases. Because free radicals, in particular superoxide, are difficult to measure, an alternative indirect method for measuring oxidative stress levels has been used successfully in Escherichia coli and yeast. This method is based on a proposed connection between elevated superoxide levels and release of iron from solvent-exposed [4Fe-4S] enzyme clusters that eventually leads to an increase in hydroxyl radical production. In past studies using bacteria and yeast, a positive correlation was found between superoxide production or oxidative stress due to superoxide within the organism and electron paramagnetic resonance (EPR) detectable "free" iron levels. In the current study, we have developed a reliable and efficient method for measuring "free" iron levels in Caenorhabditis elegans using low-temperature Fe(III) EPR at g=4.3. This method uses synchronized worm cultures grown on plates that are homogenized and treated with desferrioxamine, an Fe(III) chelator, prior to packing the EPR tube. Homogenization was found not to alter "free" iron levels, whereas desferrioxamine treatment significantly raised these levels, indicating the presence of both Fe(II) and Fe(III) in the "free" iron pool. The correlation between free radical levels and the observed "free" iron levels was examined by using heat stress and paraquat treatment. The intensity of the Fe(III) EPR signal, and thus the concentration of the "free" iron pool, varied with the treatments that altered radical levels without changing the total iron levels. This study provides the groundwork needed to uncover the correlation among oxidative stress, "free" iron levels, and longevity in C. elegans.     

Development of a multiplex amplification refractory mutation system for simultaneous authentication of Korean ginseng cultivars "Gumpoong" and "Chungsun"

BACKGROUND AND AIMS. Molecular authentication of Korean ginseng cultivars was investigated using the mitochondrial nicotinamide adenine dinucleotide (NADH) dehydrogenase subunit 7 (nad7) intron 3 region. MATERIALS AND METHODS. A mutation site specific to Panax ginseng "Gumpoong" and "Chungsun" cultivars was detected within the sequence data. Based on this mutation site and the "Gumpoong"-specific single nucleotide polymorphism site reported in 26S rDNA, two modified allele-specific primer pairs were designed and a multiplex amplification refractory mutation system (MARMS) was applied to identify "Gumpoong" and "Chungsun." RESULTS. The results showed that "Gumpoong" and "Chungsun" can be clearly discriminated from the other Korean ginseng cultivars by simultaneously identifying the haplotype of "Gumpoong" and the specific allele of "Chungsun" by applying the MARMS. CONCLUSION. This study, therefore, provides a simple and reliable method for simultaneous authentication of "Gumpoong" and "Chungsun" cultivars.     

The "jam roll" flap for fingertip reconstruction

A method of fingertip reconstruction using a deepithelialized cross-finger flap in "jam roll" fashion is described. The technique has found relatively frequent indications.     

"PCR-karyotype" of human chromosomes in somatic cell hybrids

Amplification of human DNA sequences in 16 monochromosomal somatic cell hybrids containing different human chromosomes were performed by the polymerase chain reaction (PCR) using primer directed at human-specific regions of Alu or L1, the two major classes of interspersed repetitive sequences (IRS-PCR). A chromosome-specific pattern of amplification products was observed on agarose gels run with ethidium bromide, producing a "PCR-karyotype." This simple gel analysis provides a rapid method for identifying and monitoring the human chromosomal content of monochromosomal somatic cell hybrids without conventional cytogenetic analysis. Hybrids containing multiple human chromosome produce complex gel patterns, but identification of chromosome content can be achieved by hybridization of PCR products against a reference panel of monochromosomal or highly reduced hybrids representing each human chromosome. This dot-blot method also enables identification of human marker chromosomes or translocated pieces in hybrids that are not identifiable by cytogenetic methods. These IRS-PCR methods should greatly reduce the need for more laborious cytogenetic, isozyme, and Southern blot characterizations of human-rodent cell hybrids.     

"Malignant" contracture of the eye socket

A new method for reconstruction of "malignant" contracture of the eye socket is described using a simple procedure based on the principle of epithelial inlay. The lining consists of a free skin graft. No cumbersome external appliances for the prevention of contraction of the graft are used; hence the hospitalization is minimized. The results have been satisfactory.