Bowel seromuscular flaps used to repair pelvic defects in patients undergoing reoperative surgery.

BACKGROUND: Pedicled seromuscular bowel flaps may serve as an alternative for pelvic floor reconstruction when conventional omental and muscular flaps are not an option in patients undergoing reoperative abdominopelvic surgery. The aim of this study was to evaluate a unique series of bowel seromuscular flaps used to obliterate intrapelvic defects. METHODS: We conducted a retrospective study on all patients in a single tertiary care institutional database who had undergone pelvic reconstruction with a seromuscular bowel flap from January 2006 to December 2018. The primary outcomes measured were the 30-day morbidity and mortality rates. RESULTS: Twelve patients (6 men 6 women, median age 56.5 years [range 33-77 years]) underwent reoperative abdominopelvic surgery requiring the use of a native small or large seromuscular bowel flap to obliterate pelvic defects. The indications for surgery included chronic infections, fistulizing Crohn's disease, and cancer. In all cases, no residual omentum was available and rectus abdominis muscle flaps were not feasible due to prior operative scars. Thirty-day morbidity occurred in 5 patients (42%), and included urine leak from ureteral injury, anastomotic leak, acute kidney injury, and superficial surgical site infection. No flaps became ischemic or required removal in the postoperative setting. No mortality was recorded. CONCLUSIONS: Bowel seromuscular flaps are a feasible and safe alternative for covering pelvic defects in patients who are undergoing reoperative surgery without the option to use traditional omental and muscular flaps.

Quadrupedal water launch capability demonstrated in small Late Jurassic pterosaurs.

Pterosaurs thrived in and around water for 160 + million years but their take-off from water is poorly understood. A purportedly low floating position and forward centre of gravity barred pterosaurs from a bird-like bipedal running launch. Quadrupedal water launch similar to extant water-feeding birds and bats has been proposed for the largest pterosaurs, such as Anhanguera and Quetzalcoatlus. However, quadrupedal water launch has never been demonstrated in smaller pterosaurs, including those living around the Tethys Sea in the Late Jurassic Solnhofen Lagoon. Using Laser-Stimulated Fluorescence, we singled out aurorazhdarchid specimen MB.R.3531 that alone preserved specific soft tissues among more than a dozen well-preserved Solnhofen pterosaur specimens. These soft tissues pertain to primary propulsive contact surfaces needed for quadrupedal water launch (pedal webbing and soft tissues from an articulated forelimb) that permit robust calculations of its dynamic feasibility without the need to make assumptions about contact areas. A first-principles-based dynamics model of MB.R.3531 reveals that quadrupedal water launch was theoretically feasible and that webbed feet significantly impacted launch performance. Three key factors limiting water launch performance in all pterosaurs are identified, providing a foundation for understanding water launch evolution: available propulsive contact area, forelimb extension range and forelimb extension power about the shoulder.

Effects of alcohol consumption on biomarkers of oxidative damage to DNA and lipids in ethanol-fed pigs.

Chronic alcohol consumption is known to result in tissue injury, particularly in the liver, and is considered a major risk factor for cancers of the upper respiratory tract. Here we assessed the oxidative effects of subchronic ethanol consumption on DNA and lipids by measuring biomarkers 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG) and malondialdehyde (MDA), respectively. Physiological responses of pigs (n = 4) administered ethanol in drinking water for 39 days were compared with those of water-fed pigs (n = 4). Alcoholisation resulted in serum ethanol concentration of 1.90 g L(-1) and in a moderate but significant increase in alanine aminotransferase activity, an index of liver injury. However, between the alcoholised and control groups there were no significant differences in the levels of 8-oxodG (8-oxodG per 10(6) 2'deoxyguanosine) from leucocytes (2.52 ± 0.42 Vs 2.39 ± 0.34) or from target organs, liver, cardia and oesophagus. Serum MDA levels were also similar in ethanol-fed pigs (0.33 ± 0.04 µM) and controls (0.28 ± 0.03 µM). Interestingly, levels of 8-oxodG in cardia were positively correlated with those in oesophagus (Spearman correlation coefficient R = 1, P < 0.0001). Our results suggest that alcohol consumption may not cause oxidative damage to DNA and lipids as measured by 8-oxodG and MDA, respectively. The duration of alcoholisation and the potential alcohol-induced nutritional deficiency may be critical determinants of ethanol toxicity. Relevant biomarkers, such as factors involved in sensitization to ethanol-induced oxidative stress are required to better elucidate the relationship between alcohol consumption, oxidative stress and carcinogenesis.

Raster image correlation spectroscopy (RICS) for measuring fast protein dynamics and concentrations with a commercial laser scanning confocal microscope.

Raster image correlation spectroscopy (RICS) is a new and novel technique for measuring molecular dynamics and concentrations from fluorescence confocal images. The RICS technique extracts information about molecular dynamics and concentrations from images of living cells taken on commercial confocal systems. Here we develop guidelines for performing the RICS analysis on an analogue commercial laser scanning confocal microscope. Guidelines for typical instrument settings, image acquisition settings and analogue detector characterization are presented. Using appropriate instrument/acquisition parameters, diffusion coefficients and concentrations can be determined, even for highly dynamic dye molecules in solution. Standard curves presented herein demonstrate the ability to detect protein concentrations as low as approximately 2 nM. Additionally, cellular measurements give accurate values for the diffusion of paxillin-enhanced-green fluorescent protein (EGFP), an adhesion adaptor molecule, in the cytosol of the cell and also show slower paxillin dynamics near adhesions where paxillin interacts with immobile adhesion components. Methods are presented to account for bright immobile structures within the cell that dominate spatial correlation functions; allowing the extraction of fast protein dynamics within and near these structures. A running average algorithm is also presented to address slow cellular movement or movement of cellular features such as adhesions. Finally, methods to determine protein concentration in the presence of immobile structures within the cell are presented. A table is presented giving guidelines for instrument and imaging setting when performing RICS on the Olympus FV300 confocal and these guidelines are a starting point for performing the analysis on other commercial confocal systems.

Blockade of endothelin ET(A)/ET(B) receptors favors a role for endothelin during acute Trypanosoma cruzi infection in rats.

Endothelin has been implicated in the pathogenesis of experimental and human Chagas' disease (American trypanosomiasis). In the present study, we tested the effect of bosentan, an antagonist of both ET(A) and ET(B) endothelin receptors, on parasitemia, histopathology (heart and diaphragm), heart levels of tumor necrosis factor (TNF)-alpha, interleukin (IL)-10, interferon (IFN)-gamma, CCL2, CCL3 and CCL5, and the serum levels of nitrate/nitrite (NOx). Bosentan treatment was accompanied by a significant increase in parasitemia and tissue parasitism or inflammation. In vehicle-treated rats, Trypanosoma cruzi infection increased the cardiac levels of TNF-alpha, IFN-gamma and IL-10, at day 9 post inoculation, and the TNF-alpha remained elevated until day 13. The infection also caused a significant increase in the cardiac levels of the chemokines CCL2 (9, 13 and 18 days) and CCL3 (13 and 18 days). Bosentan-treatment had no significant effect on the infection-associated increase in IFN-gamma and chemokine concentrations. There was a lower increase in IL-10 at day 9 and this was mirrored by a greater increase of TNF-alpha at day 13, in comparison with vehicle-treated rats. These latter findings correlated well with the enhanced inflammatory process in hearts of bosentan-treated infected rats. Bosentan treatment reduced the infection-associated increase in NOx serum concentration. Altogether, our data suggest that ET action on ET(A) and ET(B) receptors may play a role in the initial control of T. cruzi infection in rats probably by interfering in NO production.

Interaction of coxsackievirus A21 with its cellular receptor, ICAM-1.

Coxsackievirus A21 (CAV21), like human rhinoviruses (HRVs), is a causative agent of the common cold. It uses the same cellular receptor, intercellular adhesion molecule 1 (ICAM-1), as does the major group of HRVs; unlike HRVs, however, it is stable at acid pH. The cryoelectron microscopy (cryoEM) image reconstruction of CAV21 is consistent with the highly homologous crystal structure of poliovirus 1; like other enteroviruses and HRVs, CAV21 has a canyon-like depression around each of the 12 fivefold vertices. A cryoEM reconstruction of CAV21 complexed with ICAM-1 shows all five domains of the extracellular component of ICAM-1. The known atomic structure of the ICAM-1 amino-terminal domains D1 and D2 has been fitted into the cryoEM density of the complex. The site of ICAM-1 binding within the canyon of CAV21 overlaps the site of receptor recognition utilized by rhinoviruses and polioviruses. Interactions within this common region may be essential for triggering viral destabilization after attachment to susceptible cells.

Host range and variability of calcium binding by surface loops in the capsids of canine and feline parvoviruses.

Canine parvovirus (CPV) emerged in 1978 as a host range variant of feline panleukopenia virus (FPV). This change of host was mediated by the mutation of five residues on the surface of the capsid. CPV and FPV enter cells by endocytosis and can be taken up by many non-permissive cell lines, showing that their host range and tissue specificity are largely determined by events occurring after cell entry. We have determined the structures of a variety of strains of CPV and FPV at various pH values and in the presence or absence of Ca(2+). The largest structural difference was found to occur in a flexible surface loop, consisting of residues 359 to 375 of the capsid protein. This loop binds a divalent calcium ion in FPV and is adjacent to a double Ca(2+)-binding site, both in CPV and FPV. Residues within the loop and those associated with the double Ca(2+)-binding site were found to be essential for virus infectivity. The residues involved in the double Ca(2+)-binding site are conserved only in FPV and CPV. Our results show that the loop conformation and the associated Ca(2+)-binding are influenced by the Ca(2+) concentration, as well as pH. These changes are correlated with the ability of the virus to hemagglutinate erythrocytes. The co-localization of hemagglutinating activity and host range determinants on the virus surface implies that these properties may be functionally linked. We speculate that the flexible loop and surrounding regions are involved in binding an as yet unidentified host molecule and that this interaction influences host range.

Reduced cholesterol is associated with recent minor illness: the CARDIA Study. Coronary Artery Risk Development in Young Adults.

Lower levels of plasma total cholesterol have been observed during severe infection, but it is not known whether the minor illnesses encountered in the general population are also associated with reduced cholesterol. This paper examines the relation between minor illness and plasma lipids, using 7- and 10-year follow-up data from more than 3,000 generally healthy participants in the Coronary Artery Risk Development in Young Adults (CARDIA) Study. At both 7 and 10 years, approximately 8.5% of participants stated they had been "ill with cold, flu, fever, or vomiting in the past 24 hours." In both cross-sectional and longitudinal analyses, the plasma total cholesterol was about 5 mg/dl lower (p < 0.006) and high density lipoprotein cholesterol about 1.2 mg/dl lower (p < 0.12) in those who reported minor illness than in those who did not. Plasma triglycerides did not vary with minor illness. The authors conclude that reductions in plasma total, low density, and high density lipoprotein cholesterol mark an acute phase response even during minor illness. These reductions may bias surveys over a limited geographic area during a short period because the proportion with minor illness may vary locally. Because this effect should be stronger with more precise illness diagnosis, clinicians should avoid making measurements for cholesterol management when illness may alter plasma lipid levels and the resulting decisions.

Epidemiology of low total plasma cholesterol concentration among young adults: the CARDIA study. Coronary Artery Risk Development in Young Adults.

BACKGROUND: Data on stability of plasma total cholesterol levels and its determinants among young adults are lacking. Knowledge of factors associated with low levels of plasma total cholesterol during young adulthood may help clarify the nature of associations between hypocholesterolemia and health or illness. METHODS: Tracking of plasma total cholesterol was investigated using data from the baseline (1985-1986), Year 5 (1990-1991), and Year 7 (1992-1993) examinations of the Coronary Artery Risk Development in Young Adults Study. Lifestyle (including dietary), physiological, medical, and psychological correlates of plasma total cholesterol were examined cross-sectionally at baseline using ANCOVA and multivariate logistic regression. The attributes of participants with persistently low plasma total cholesterol level after 7 years (i.e., remaining below the 10th percentile of sex- and race-specific distributions) were also examined. RESULTS: The cohort in this analysis comprised 720 black men, 922 white men, 899 black women, and 944 white women who were between the ages of 18 and 30 years at baseline. Between 44 and 52% of those with plasma total cholesterol levels below the 10th percentile remained below the same percentile 7 years later. Among black men, a difference of 1 SD in age [3.7 years; odds ratio (OR) = 0.69; 95% CI = 0.52-0.91] and a difference of 1 SD in systolic blood pressure (10.5 mm Hg; OR = 0.73; 95% CI = 0.54-0.97) were independently associated with lower odds, respectively, of being in the lowest 10th percentile of the plasma total cholesterol distribution. Also among black men, current smoking and more calories from carbohydrates were associated with nonsignificantly higher odds of low total cholesterol level. Among white men, a 1 SD older age (3.4 years; OR = 0.78; 95% CI = 0.61-1.00) and a 1 SD higher physical fitness (118 sec; OR = 1.41; 95% CI = 1.09-1.82) predicted lower and higher odds, respectively, of low plasma total cholesterol. Among black women, a 1 SD difference in albumin (0.3 g/dL; OR = 0.80; 95% CI = 0.63-1.03) was related to lower odds of low plasma total cholesterol. Among white women, the factors independently associated with low plasma total cholesterol were body mass index (OR for a difference in 4.0 kg/m2 = 0.73; 95% CI = 0.54-1.00) and gamma-glutamyl transferase (OR for an increase in 9.6 IU/L = 0.41; 95% CI = 0.18-0.93). The independent predictive factors of stably low total cholesterol levels were age and uric acid among black men (both inversely related) and age, Framingham Type A Behavior (inversely), and calories from carbohydrates (positively related) among white men. CONCLUSION: Young adults with low plasma total cholesterol level have characteristics generally associated with good cardiovascular health. However, adverse attributes such as current cigarette smoking (notably among black men) may confound future associations between low total cholesterol and disease.

Genome organization of the Kresse strain of porcine parvovirus: identification of the allotropic determinant and comparison with those of NADL-2 and field isolates.

The Kresse strain of porcine parvovirus (PPV) was cloned into pUC19, and independent infectious clones were sequenced. The PPV Kresse and NADL-2 strains, which have different pathogenicities, shared an identical genomic organization and a high degree of sequence identity. Partial genomes (1.5 or 1.6 kb) of 15 field isolates were also amplified by PCR in regions with significant sequence differences between the laboratory strains. Five amino acid differences were consistently present within the VP1/VP2 coding region of the Kresse strain and virulent field isolates. A number of inconsistent point mutations were also found throughout the genomes of field isolates. In addition, among those with the vaccine amino acid profile, all but one isolate (IAF-3) contained a 127-bp noncoding direct repeat downstream of the capsid protein gene. The one exception was also the only vaccine-type PPV obtained from a mummified fetus. In order to identify genetic elements responsible for the distinct tropism (and possibly the pathology) of the Kresse strain, in vitro cell systems which differentiated the virulent from the vaccinal strains were established. Subsequently, chimeric infectious clones of the Kresse and NADL-2 strains were used to identify the allotropic determinant located in the VP1/VP2 region. The transfer of the BglII fragment of the Kresse genome, containing three amino acid differences, into the NADL-2 background, or the opposite construct, caused the phenotype of the target genome to revert to that of the parent strain of the BglII fragment. Prediction of the localization of amino acid differences on the basis of canine parvovirus capsid structure indicates that each is located on or near the outer surface of the virion. In particular, the position of one mutation (S-436-->P) maps by analogy to the threefold spike, the most accessible region of the capsid.

Fair-play rules and injury reduction in ice hockey.

OBJECTIVE: To determine the rate, type, and severity of injuries incurred and penalties assessed during the qualifying fair-play (points for playing without excessive penalties) and championship "regular" rules (winner advances) portions of a 1994 Junior Gold ice hockey tournament. DESIGN: A prospective evaluation of injuries by certified athletic trainers at the tournament site. SETTING: A community-organized, 3-day, 31-game tournament in Minnesota. PARTICIPANTS: Two hundred seventy-three male players, younger than 20 years and in high school. MEASUREMENTS/MAIN RESULTS: Injuries were recorded by an on-site certified athletic trainer, and the penalties were tallied from the score sheets. The injury rates for the total number of injuries were 26.4 injuries per 1000 athlete exposures and 273.8 injuries per 1000 player hours. When only notable injuries (concussion, facial laceration, or moderate level of severity and above) were considered, the injury rates were 10 per 1000 athlete exposures and 103.9 per 1000 player hours. The ratio of notable fair-play to notable regular-rules injuries was 1:4.8. The number of penalties assessed per game averaged 7.1 penalties during fair-play rules and 13 penalties during the regular-rules competition. Penalties related to rough play and injury occurred four times more frequently during games with regular rules than those with fair-play rules. CONCLUSIONS: The fair-play concept can reduce injury rates, penalty rates, and severity of penalties and should be considered for ice hockey at all levels of play. The fair-play concept could be applied to other contact sports to reduce injury rates and rules infractions.

Protein tyrosine kinases transcribed in a murine thymic medullary epithelial cell line.

Medullary epithelial cells of the thymus can be activated by contact with thymocytes. We identified a number of protein tyrosine kinases (PTKs) that could be responsible for the tyrosine phosphorylation observed in a murine thymic medullary epithelial cell line (E-5) following complex formation with thymocytes. Degenerate oligodeoxyribonucleotides (oligos) derived from the amino acid (aa) sequence motifs of PTK catalytic domains were used as oligo primers for PCR amplification to determine the PTK genes which are normally transcribed in the E-5 cell line. Amplicons were cloned, sequenced and the deduced aa sequences were compared to known PTK sequences. Among the 13 distinct PTK catalytic domains identified in E-5 cells, two were novel: they were encoded by eteck, a member of the eph sub-family of PTKs, and thy, a member of the src sub-family of PTKs.

Increased PCR sensitivity by using paraffin wax as a reaction mix overlay.

The use of mineral oil as a reaction mix overlay in conventional PCR may lead to problems. In addition to more difficult handling, traces of mineral oil in amplicon suspensions have been shown to decrease the efficiency of post-PCR manipulations. Commercial alternatives aimed at resolving the problem more than double the cost of an amplification. This is an important drawback when dealing with a large number of clinical samples. The use of inexpensive paraffin wax as a reaction mix overlay eliminated problems associated with the presence of mineral oil while being more practical and safer in handling potentially contaminated clinical samples. Moreover, when used in conjunction with a modified hot start technique, the use of paraffin wax increased the specificity and sensitivity of PCR amplifications over mineral oil in similar hot start conditions. Using the aforementioned method, the increase in specificity and sensitivity has enabled specific detection of viral DNA in clinical samples which the conventional PCR method failed to detect.

Influences of anabolic androgens on cardiac growth and metabolism in the rat.

Testosterone, and to a lesser degree methandrostenolone, was shown to influence cardiac growth in immature male rats by affecting protein synthesis and degradation. The nature of cardiac responses to androgen appear to depend on the prevailing experimental conditions. Protein synthesis was inhibited in the castrate rat and was stimulated by subsequent treatment with androgen. Under conditions of induced overgrowth of the ventricles, androgens gave rise to an attenuation of the effects of aortic constriction on ventricular mass and blood pressure involving smaller changes in protein synthesis and proteolysis. Concentrations of testosterone receptors in ventricular cytosol further indicated that the myocardium is more sensitive to androgen action during the prepubertal phase of the life-span. Changes in amount and properties of the receptors showed them to be functional and responsive to castration, aortic constriction, and administration of the androgens. The androgens affected cardiac protein balance by stimulating the incorporation of radiolabelled amino acid into protein in vivo. They also appeared to influence proteolytic processes involving lysosomal hydrolase activities, but their actions were either stimulatory or inhibitory depending on the internal environment. The heart is a target organ for several hormones including androgen, and our findings fortify the notion that hormone action needs to be investigated alone and in association with other endocrines.

Development of the coronary vasculature in hypoxic fetal rats treated with a purified perfluorocarbon emulsion.

OBJECTIVE: To quantitatively define the coronary vascular bed in the 21-day-old rat fetus with gestational normoxia and hypoxia; to determine if maternal supplemental oxygen and/or oxygen-carrying perfluorocarbons (PFCs) influence development of coronary vessels; and to compare the results using purified and unpurified PFC treatment. DESIGN: Unilateral uterine artery ligation was introduced on gestational day 17 in pregnant animals. Control fetuses were from unligated uterine horns. Experimental intervention occurred during gestational days 17 to 21, with fetuses recovered on day 21. Developing coronary vessels were analyzed quantitatively via light microscopy. ANIMALS: Pregnant Sprague Dawley rats. INTERVENTIONS: Following ligation, pregnant rats received no further treatment, supplemental oxygen inhalation alone, or daily intravenous purified PFC treatment, with or without supplemental oxygen. MAIN RESULTS: Hypoxia caused an increase in resorptions (P less than 0.001), and decreased fetal body weight (P less than 0.001) and heart weight (P less than 0.05). Although the area occupied by developing coronary vessels (sinusoids) was substantially increased, maturation was unchanged. Oxygen supplementation alone did not appreciably influence fetal resorptions or body weight in ligated horns, but did increase fetal heart weight. Sinusoidal area decreased (P less than 0.01), with no effect on sinusoidal maturity. Purified PFC treatment did not alter maternal weight gain or fetal body weight, and moderately decreased resorptions in ligated horns. Fetal heart weight was augmented with purified perflurochemical, while unpurified perfluorochemical treatment diminished heart weight. Both PFC emulsions substantially decreased sinusoidal area. CONCLUSIONS: Perflurocarbon treatment associated with supplemental oxygen is capable of improving the hypoxic effects on fetal heart and coronary vessel development if the emulsion used is appropriately purified.

N-glycosylation and in vitro enzymatic activity of human recombinant tissue plasminogen activator expressed in Chinese hamster ovary cells and a murine cell line.

To probe the effects of N-glycosylation on the fibrin-dependent plasminogenolytic activity of tissue-type plasminogen activator (t-PA), we have expressed a human recombinant t-PA (rt-PA) gene in Chinese hamster ovary (CHO) cells and in a murine C127 cell line. The resulting rt-PA glycoproteins were isolated and their associated N-linked oligosaccharide structures determined by using a combination of high-resolution Bio-Gel P-4 gel filtration chromatography, sequential exoglycosidase digestion, and methylation analysis. The results show that CHO rt-PA is N-glycosylated differently from murine C127 derived rt-PA. Further, both rt-PA's are N-glycosylated differently from t-PA derived from a human colon fibroblast and the Bowes melanoma cell line (Parekh et al., 1989), confirming that N-glycosylation of the human t-PA polypeptide is cell-type-specific. Both CHO and murine rt-PA were fractionated on lysine-Sepharose chromatography. The N-glycosylation of the major forms was analyzed and their fibrin-dependent plasminogenolytic activity determined by using an indirect amidolytic assay with Glu-plasminogen and a chromogenic plasmin substrate. The results suggest that the various forms of rt-PA differ from one another with respect to the kinetics of their fibrin-dependent activation of plasminogen. Together, these data support the notion (Wittwer et al., 1989) that N-glycosylation influences the fibrin-dependent catalytic activity of t-PA and that t-PA when expressed in different cell lines may consist of kinetically and structurally distinct glycoforms.