Trajectory of functional outcome and health status after moderate-to-major trauma in Hong Kong: A prospective 5 year cohort study.

BACKGROUND: Trauma care systems in Asia have been developing in recent years, but there has been little long-term outcome data from injured survivors. This study aims to evaluate the trajectory of functional outcome and health status up to five years after moderate to major trauma in Hong Kong. METHODS: We report the five year follow up results of a multicentre, prospective cohort from the trauma registries of three regional trauma centres in Hong Kong. The original cohort recruited 400 adult trauma patients with ISS ≥ 9. Telephone follow up was conducted longitudinally at seven time points, and the extended Glasgow Outcome Scale (GOSE) and Short-Form 36 (SF36) were tracked. RESULTS: 119 out of 309 surviving patients (39%) completed follow up after 5 years. The trajectory of GOSE, PCS and MCS showed gradual improvements over the seven time points. 56/119 (47.1%) patients reported a GOSE = 8 (upper good recovery), and the mean PCS and MCS was 47.8 (95% CI 45.8, 49.9) and 55.8 (95% CI 54.1, 57.5) respectively at five years. Univariate logistic regression showed change in PCS - baseline to 1 year and 1 year to 2 years, and change in MCS - baseline to 1 year were associated with GOSE = 8 at 5 years. Linear mixed effects model showed differences in PCS and MCS were greatest between 1-month and 6-month follow up. CONCLUSIONS: After injury, the most rapid improvement in PCS and MCS occurred in the first six to 12 months, but further recovery was still evident for MCS in patients aged under 65 years for up to five years.

Circulating human leucine-rich α-2-glycoprotein 1 mRNA and protein levels to detect acute appendicitis in patients with acute abdominal pain.

BACKGROUND: Elevated levels of circulating plasma and urine leucine-rich-2-glycoprotein-1 (LRG1) protein has been found in patients with acute appendicitis (AA) and may be useful for diagnosis. This study aimed to investigate whether combined tests including circulating LRG1 mRNA levels improve the early diagnosis of AA. METHODS: Between December 2011 and October 2012, a prospective study was conducted on patients aged 18years or older presenting to the ED with acute abdominal pain (<7days of symptom onset). Levels of whole blood LRG1 mRNA and plasma LRG1 protein taken from these patients within 24h of arrival (mean 12.4h) were analyzed. The primary outcome was AA. RESULTS: Eighty-four patients (40 (47.6%) with AA and 44 (52.4%) without AA; mean age 35years; 41.6% males) were recruited. Median whole blood LRG1 mRNA and plasma LRG1 levels were higher in AA patients than in non-AA. Of 40 AA patients, 13 (32.5%) were diagnosed as complicated AA. In ROC analysis of LRG1 mRNA (normalized to GAPDH), LRG1 protein and Alvarado score for discriminating AA and non-AA, the areas under the curve (AUC) were 0.723, 0.742 and 0.805 respectively. The AUC of combination of normalized LRG1 mRNA, LRG1 protein and Alvarado score was 0.845. CONCLUSION: A combination of modified whole blood LRG1 mRNA levels, plasma LRG1 protein and Alvarado score at the ED may be useful to diagnose simple and complicated AA from other causes of abdominal pain.

Add-on tests for improving risk-stratification in emergency department patients with chest pain who are at low to moderate risk of 30-day major adverse cardiac events.

BACKGROUND: Chest pain patients commonly present to emergency departments (ED), and require either hospital admission and/or lengthy diagnostic protocols to rule-out myocardial infarction. We aimed to identify the best combination of add-on tests to high-sensitivity cardiac troponin (hs-cTnT) for predicting 30-day major adverse cardiac events (MACE) in adult chest pain patients presenting to an ED with suspected acute coronary syndrome. METHODS: This prospective observational study was conducted in the ED of a tertiary university hospital in Hong Kong, recruiting adult patients with chest pain of less than 24h duration, suspected with acute coronary syndrome (ACS), and had no history of coronary artery bypass grafting or stent insertion. Patients underwent triage assessment, electrocardiography, blood sampling for laboratory hs-cTnT, and Thrombolysis in Myocardial Infarction (TIMI) and HEART score assessment. The primary outcome was the number of patients with 30-day MACE. RESULTS: 602 consecutive patients were recruited and completed 30-day follow-up. A 30-day MACE occurred in 42 (7.0%) patients. Out of 12 possible models for stratifying patients at risk of 30-day MACE within 2h of ED arrival, a combination of electrocardiography (ECG) and one-time hs-cTnT (model 5) provided the simplest and most accurate model. A risk score of 0 to 5 was derived from raw coefficients of model 5. The risk score provided excellent calibration (P=0.91) and discrimination (AUC 0.87, 95% CI: 0.82 to 0.93). CONCLUSION: Appropriate early risk-stratification of patients with chest pain and possible ACS using a combination of ECG and one-time hs-cTnT may improve efficiency of care.

Metallothionein induces a regenerative reactive astrocyte phenotype via JAK/STAT and RhoA signalling pathways.

Following central nervous system injury, astrocytes rapidly respond by undergoing a stereotypical pattern of molecular and morphological alterations termed "reactive" astrogliosis. We have reported previously that metallothioneins (MTs) are rapidly expressed by reactive astrocytes and that their secretion and subsequent interaction with injured neurons leads to improved neuroregeneration. We now demonstrate that exogenous MT induces a reactive morphology and elevated GFAP expression in cultured astrocytes. Furthermore, these astrogliotic hallmarks were mediated via JAK/STAT and RhoA signalling pathways. However, rather than being inhibitory, MT induced a form of astrogliosis that was permissive to neurite outgrowth and which was associated with decreased chondroitin sulphate proteoglycan (CSPG) expression. The results suggest that MT has an important role in mediating permissive astrocytic responses to traumatic brain injury.

Metallothionein treatment attenuates microglial activation and expression of neurotoxic quinolinic acid following traumatic brain injury.

The kynurenine pathway has been implicated as a major component of the neuroinflammatory response to brain injury and neurodegeneration. We found that the neurotoxic kynurenine pathway intermediate quinolinic acid (QUIN) is rapidly expressed, within 24 h, by reactive microglia following traumatic injury to the rodent neocortex. Furthermore, administration of the astrocytic protein metallothionein attenuated this neuroinflammatory response by reducing microglial activation (by approximately 30%) and QUIN expression. The suppressive effect of MT was confirmed upon cultured cortical microglia, with 1 mug/ml MT almost completely blocking interferon-gamma induced activation of microglia and QUIN expression. These results demonstrate the neuroimmunomodulatory properties of MT, which may have therapeutic applications for the treatment of traumatic brain injury.

Metallothionein expression by NG2 glial cells following CNS injury.

Metallothionein (MT) expression is rapidly up-regulated following CNS injury, and there is a strong correlation between the presence or absence of MTand improved or impaired (respectively) recovery from such trauma.We now report that a distinct subset of NG2-positive, GFAP-negative glial cells bordering the injury tract express MT following focal injury to the adult rat neocortex. To confirm the ability of these NG2 glial cells to express MT, we have isolated and cultured them and identified that they can express MT following stimulation with zinc. To investigate the functional importance of MT expression by NG2 glial cells, we plated cortical neurons onto these cells and found that expression of MT enhanced the permissivity of NG2 glial cells to neurite outgrowth. Our data suggest that expression of MT by NG2 glial cells may contribute to the overall permissiveness of these cells to axon regeneration.

AP-2 regulates the transcription of estrogen receptor (ER)-beta by acting through a methylation hotspot of the 0N promoter in prostate cancer cells.

We reported previously that the loss of expression of estrogen receptor (ER)-beta during the development of prostate cancer (PCa) is associated with methylation of a CpG island located in the 5'-flanking sequence of the 0N promoter. Three methylation hotspots, referred to as centers 1, 2 and 3, were identified in the CpG island. In this study, we demonstrated that a 581-bp region with these three centers within it is sufficient for the promoter activity in PCa cells. Deletion analyses indicated that center 1 (16 bp), with a putative activator protein-2 (AP-2) binding site, is essential for gene transactivation. Chromatin immunoprecipitation assays showed that AP-2alpha occupies a short sequence containing center 1. Forced expression of AP-2alpha or -2gamma, but not -2beta, increased activity of the ERbeta 0N promoter and the accumulation of mRNA. Conversely, siRNA-mediated AP-2alpha and -2gamma knockdown reduced levels of ERbeta transcript and promoter activity. Quantitative reverse transcription-PCR showed that AP-2alpha and -2gamma are the predominant transcripts expressed in PCa cells, and levels of ERbeta transcript correlate with levels of these AP-2 transcripts among different PCa cell lines. These results provide the first evidence that ERbeta is an AP-2-regulated gene. They also support the hypothesis that certain cis-acting elements are methylation hotspots susceptible to epigenetic modifications during cancer progression.

Purification and properties of ferrochelatase from Chironomidae larvae.

Ferrochelatase with an Mr of 42,700 Da and a pI of 7.35 has been purified to homogeneity from chironomidae larvae. The activity of the enzyme reached maximum at pH 7.8 and decreased with the increase of pH. The enzyme activity varied with temperature and showed maximum activity around 37 degrees C. The purified enzyme was active towards protoporphyrin but inactive towards other porphyrins. The specific enzyme activity of ferrochelatase from chironomidae is about 10-fold higher than that of the rat. Electrophoresis of the purified fractions shows that the enzyme contains only one single polypeptide. The soluble ferrochelatase contained one mole of iron in each mole of the enzyme. The N-terminal sequence analysis of the enzyme shows a high percentage of conserved regions of the enzyme among other species. The enzyme properties are similar to those of the mammalian ferrochelatases except with slightly higher specific activity. Chironomidae ferrochelatase appeared to be more heat resistant and less susceptible than its mammalian equivalent to inhibition by lead.

Effects of vitamins and common drugs on reduction of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone in rat microsomes.

4-(Methylnitrosamino)-1-(3-pyridyl)-butanone (NNK) is a tobacco-specific nitrosamino that requires metabolic activation by cytochrome P450 enzymes. The activation of NNK by cytochrome P450 enzymes leads to the formation of different metabolites. Detoxification of NNK usually occurs via carbonyl reduction to its hydroxyl product, 4-(methylnitrosamino)-1-(3-pyridyl)-butanol (NNAL). In the present study, the influences of common vitamins and P450 modulators on the reduction of NNK by rat microsomes were studied. The formation of NNAL but not other metabolites was detected by the described HPLC method. Among the vitamins tested, vitamins E, A (retinol), B6 and B5 were found to be marginal effective upon reduction of NNK while vitamins A (cis-acid), A (trans-acid), D2, D3, K1, K3, B1 and A (crocetin) increased the formation of NNAL from 3 to 21%. The effect of vitamin C-palmitate (<10 microM) was most pronounced followed by crocetin upon reduction of NNK. Clonidine, tolbutamide and atropine slightly increased the reduction of NNK while cimetidine showed no effects. The modulation of NNK reduction could reduce the carcinogenic potential of NNK, since the main detoxification pathway of NNK involves carbonyl reduction.

Modulation of reduction of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone by vitamin C-palmitate.

An in vitro study of effects of vitamin C-palmitate on the metabolism of 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) in rat microsomes was performed. A sensitive assay method has been developed for the detection of metabolites of NNK in microsomes. Only the reduced metabolite of NNK, 4-(methylnitrosamino)-1-(3-pyridyl)-butanol (NNAL), was detected and measured in a time-course study. Vitamin C-palmitate enhanced the reduction of NNK in a concentration-dependent manner. The results indicate a significant increase in Vmax and K(m) in the presence of vitamin C. However, the rate of formation of NNAL at low substrate concentration varied. The ratio of Vmax to K(m) decreases. The results suggest that the kinetics are accounted for best by an uncompetitive activator binding model at low concentration of vitamin C. The uncompetitive binding model becomes sketchy at higher concentration of vitamin C. These observations infer that vitamin C loosely binds to the substrate-enzyme complex. Furthermore, the nature of the binding would facilitate the modulation of NNK biotransformation leading to the formation of NNAL. The results also show that vitamin C-palmitate is a potent activator of NNK reduction in rat liver microsomes. Thus, vitamin C-palmitate would mediate the metabolism of NNK through reduction. The resulting NNAL-glucuronide is more readily eliminated in urine.

Expression of grass carp growth hormone by baculovirus in silkworm larvae.

A total of five recombinant Bombyx mori nuclear polyhedrosis viruses (BMNPV) carrying the grass carp (Ctenopharyngodon idellus) growth hormone (GH) cDNA were constructed in this study. Two of them were able to express the hormone up to a level of 12 microgram/ml medium when cultured B. mori cells were infected for 4 days. Inoculation of the viruses into silkworm (B. mori) host significantly increased the level of GH achievable. The amount of hormone produced per larva was estimated to be around 1 mg. The recombinant grass carp GH had immunological and biological activities similar to the native hormone. The N-terminal sequence of the recombinant hormone was the same as the native one, indicating that the fish signal peptide was correctly processed by the insect cells. Silkworm powder prepared from larvae infected with the recombinant virus was used as food supplement for fish. Compared with the control, this dietary supplement was effective in increasing the growth rate of juvenile carp.

Inhibitory effect of nicotine and its metabolites on tolbutamide hydroxylation in rat liver microsomes.

A simple HPLC/fluorescence method to detect hydroxytolbutamide (a major metabolite of the anti-diabetic drug tolbutamide) has been developed. The effects of nicotine and some of its metabolites on tolbutamide hydroxylation is described. An extraction procedure with diethyl ether was followed by isocratic HPLC analysis of tolbutamide hydroxylation with a binary mobile phase composed of 10 mM monobasic sodium phosphate in methanol (45:55, v/v, apparent pH 2.28). A detection limit of sub-nanogram amounts (0.353 ng) of hydroxytolbutamide was obtained with fluorescence detection at 226 nm for excitation and 318 nm for emission. Overall precision values for hydroxytolbutamide was determined with coefficients of variation of 1.4-4.6% when nanogram levels of the metabolite were analyzed. Differential inhibitory responses were demonstrated for tolbutamide hydroxylation to nicotine and its metabolites. Tolbutamide hydroxylation was apparently inhibited by cotinine and relatively less inhibited by nicotine. Nornicotine, however, caused very little inhibition of tolbutamide hydroxylation. The implication is that nornicotine may not share similar affinity for the substrate binding site for tolbutamide. The results also suggest that heavy smokers may experience reduction in tolbutamide metabolism. The assay system itself will be useful for future studies of tolbutamide, and possibly related sulfonylureas.

Characteristics of frequent pediatric emergency department users.

The purpose of this study was to examine the medical and demographic characteristics of patients who frequently seek emergency care at a pediatric emergency department (ED). Registration information of ED visits during the study period from 11/1/87 to 5/31/92 (4.6 years) was stored in a data base. Patients with 10 or more ED visits during this study period were considered to be "frequent" ED users. Outpatient and inpatient medical records of these patients were manually reviewed. Demographics, chronic conditions, and the acute conditions for each ED visit were coded and analyzed. During the study period, there were 79,049 ED patient visits under 21 years of age. Of the patients born after 1970, there were 47,451 visits by patients seen one or two times, 25,883 visits by patients seen three to nine times, and 5178 visits by 357 patients seen in the ED ten times or more. Ninety-nine patients were seen more than 15 times, 39 patients were seen more than 20 times, 17 patients were seen more than 25 times, and 10 patients were seen more than 30 times. Two hundred sixty-five of the 357 frequent ED users (74%) had chronic disease conditions. Two hundred and twenty-three of them had good functional status, 25 had mild or moderate impairment in carrying out activities of daily living, and 17 had severe impairment of function. The most common chronic medical conditions were recurrent wheezing (226), neurologic conditions (33), gastrointestinal conditions (13), cardiac conditions (12), and endocrine conditions (9). The other 92 were assessed as healthy children. Patients' immunization status were up to date as of the last ED visit during the study period in 329 patients (92%). Pediatricians were the primary care providers in 339 patients (95%). Medical insurance status of patients follows: private insurance (38%), military (0.3%), Medicaid or state assistance (60%), and no insurance (1.4%). Polynesian ethnic groups were over-represented in the cohort of frequent ED users. We conclude that cultural differences appeared to be an important factor associated with frequent ED use by healthy persons. Medical care resources as measured by immunizations, insurance, and identification of a primary care physician did not appear to be deficient in this cohort of frequent ED users. Since recurrent wheezing is a dominant chronic condition among frequent ED users, pediatric emergency medicine training programs may consider the inclusion of the chronic management of wheezing in their curriculum.

Verapamil decreases ethanol-induced gastric acid secretion in rats.

The present study examined the effect of verapamil, a calcium channel blocker, on gastric acid secretion and circulating gastrin levels in rats after ethanol challenge. Normal saline or verapamil were given intraperitoneally to different groups of rats at 1 min, 1 h or 2 h before the administration of ethanol. One hour later, gastrin and gastric acid concentrations were determined. Regression analysis revealed the relationship between gastric acid output and serum gastrin levels in the group receiving saline intraperitoneally and ethanol orogastrically and the group receiving saline both intraperitoneally and orogastrically is similar. The slope of the regression line of the ethanol-challenged group treated with verapamil, however, was significantly lower than the slopes of the other two groups (P less than 0.001). Furthermore, verapamil decreased gastrin levels and acid output significantly in the ethanol-challenged group (P less than 0.01). When given 10 min prior to ethanol challenge, verapamil had a greater acid suppression effect than when given 60 or 120 min before the challenge. Verapamil at 20 mg/kg was more effective in acid secretion than at 10 mg/kg bodyweight, when administered 60 min before ethanol challenge.

Kinetics of the multi-step catalytic degradation of a polymer to its constituent subunits.

The stepwise catalytic degradation of a polymer to its constituent subunits results in many molecules of polymers of shorter chain length. Using the Law of Mass Action, ordinary differential equations for each species can be obtained from the set of elementary reactions and these can be integrated to obtain species concentrations as a function of time. A special case of catalytic degradation of polymers, in which one subunit is released at each step, is examined. A mathematical dissertation on the kinetics of the series of reactions involved is presented. The solution of the set of differential equations involve tedious computations and cannot be easily adapted to the analysis of experimental data. Two additional methods of analysis are described. The first one makes use of the property that for any intermediary polymer whose initial and final concentrations are zero, the integral of its concentration with respect to time is inversely proportional to the degradation rate constant. Stepwise calculation of the rate constants can thus be obtained by determining the area under the concentration-time trajectories while the degradation rate constant of the first polymer is known. The second method makes use of the property that the maximum concentration attained by any individual intermediary polymer is inversely proportional to its degradation constant. These methods have the advantage of being simple since the degradation constants can be obtained stepwise from experimental data.

Medium-chain acyl CoA dehydrogenase deficiency: electron microscopic differentiation from Reye syndrome.

Inborn errors involving the oxidative metabolism of fatty acids may present clinically with a Reye syndrome-like picture. This case report of a patient with medium-chain acyl CoA dehydrogenase (MCAD) deficiency illustrates that electron microscopy may help to differentiate this disorder from Reye syndrome even if a liver biopsy is performed in a patient who recovered from an acute metabolic decompensation. Together with this case, a review of the few reports in the literature of pathological findings in MCAD deficiency is given. Changes uncharacteristic for Reye syndrome are a large-droplet steatosis and the presence of distinctive mitochondrial abnormalities on electron microscopy. The detection of an electron dense mitochondrial matrix and a widened space of inner mitochondrial membranes rules out Reye syndrome and is suggestive of a disorder of mitochondrial fatty acid oxidation.

Effect of thyroxine on the maturation of cholecystokinin (CCK) receptors in pancreatic acini of neonatal rats.

Neonatal rat pancreata are not responsive to stimulation by cholecystokinin (CCK) and this has been shown to be due partly to low binding of CCK to pancreatic acinar cells of rats at this age. The effect of thyroxine on the maturation of CCK receptor binding and enzyme secretion is studied. One-day-old rat pups were injected daily with thyroxine (0.1 microgram/g of body weight) for 3 days and killed on day 5. Control littermates were injected with normal saline at the same volume and schedule as the thyroxine group. The pancreatic weight and amylase activity were significantly higher in pups from the thyroxine group. Amylase release after stimulation with various concentrations of CCK was also higher in the thyroxine group. The maximal binding to [125I]BH-CCK-8 was significantly higher in dispersed acini from the thyroxine group when compared to the control group (5.2 vs. 2.0%). Analysis of binding data showed that the higher binding was due to a higher maximal binding capacity in the thyroxine group (1.1 +/- 0.41 vs. 5.2 +/- 1.4 fmol/mg of protein). Thyroxine, therefore, induces a precocious maturation of the secretory function of the pancreatic acini, specifically by modulating the maximal binding capacity of the high-affinity binding sites.

The role of carbohydrate moieties of cholecystokinin receptors in cholecystokinin octapeptide binding: alteration of binding data by specific lectins.

Cholecystokinin (CCK) receptors on rat pancreatic acini have been demonstrated to be glycoproteins. In order to study whether their carbohydrate moieties play a role in ligand binding, membrane preparations (adjusted to 0.2 mg protein me) were incubated with 20 pM 125-I-CCK octapeptide (125I-CCK8) for 4 h at 30 degrees C in the presence of lectins with different sugar specificities. Concanavalin A, soy-bean agglutinin, and peanut agglutinin in concentrations up to 1 mM did not alter specific 125I-CCK8 binding. Ulex europeus lectin I showed a dose-dependent enhancement of CCK binding up to 150% of controls at a concentration of 1 mM. Wheat-germ agglutinin (WGA) was the only lectin found to have an inhibitory effect. Inhibition was dose-dependent, with maximal reduction attained at 42 nM, but CCK binding was only partially inhibited to 66.2 +/- 4.4%. Inhibition by WGA was prevented by the presence of N-acetyl-D-glucosamine or N,N',N"-triacetylchitotriose, sugars that are specific for WGA. The inhibitory effect of WGA was not due to an increase in non-specific binding, increased CCK degradation, or CCK binding to WGA. Binding data indicated that the presence of WGA resulted in a decrease in receptor affinity (Kd = 567 +/- 191 v. 299 +/- 50 pM). No significant change in the number of available binding sites was observed. This suggests that WGA is not binding to the active binding site. It is conceivable that binding of WGA to N-acetyl-D-glucosamine or its polymers can lead to a conformational change in the receptor protein, and that this carbohydrate moiety is essential for optimal receptor-ligand interaction.