Recycling of Primary Lithium Batteries Production Residues.

Production waste of primary lithium batteries constitutes a considerable secondary lithium feedstock. Although the recycling of lithium batteries is a widely studied field of research, the metallic residues of non-rechargeable lithium battery production are disposed of as waste without further recycling. The risks of handling metallic Li on a large scale typically prevent the metal from being recycled. A way out of this situation is to handle Li in an aqueous solution, from where it can be isolated as Li2 CO3 . However, the challenge in hydrometallurgical treatment lies in the high energy release during dissolution and generation of H2 . To reduce these process-related risks, the Li sheet metal punching residues underwent oxidative thermal treatment from 300 to 400 °C prior to dissolution in water. Converting Li metal to Li2 O in this initial process step results in an energy release reduction of ∼70 %. The optimal oxidation conditions have been determined by experimental design varying three factors: temperature, Li metal sheet thickness, and residence time. With 96.9±2.6 % almost the entire Li amount is converted to Li2 O, after 2.5 h treatment at 400 °C for a Li sheet thickness of 1.99 mm. Final precipitation with CO2 yields 85.5±3.0 % Li2 CO3 . Using pure Li sheets, the product Li2 CO3 is obtained in battery-grade quality (>99.5 %). Non-precipitated Li is recirculated into the process on the stage of dissolving Li2 O, thus avoiding loss of material.

Pathway-Based Approaches for Assessing Biological Hazards of Complex Mixtures of Contaminants: A Case Study in the Maumee River.

Assessment of ecological risks of chemicals in the field usually involves complex mixtures of known and unknown compounds. We describe the use of pathway-based chemical and biological approaches to assess the risk of chemical mixtures in the Maumee River (OH, USA), which receives a variety of agricultural and urban inputs. Fathead minnows (Pimephales promelas) were deployed in cages for 4 d at a gradient of sites along the river and adjoining tributaries in 2012 and during 2 periods (April and June) in 2016, in conjunction with an automated system to collect composite water samples. More than 100 industrial chemicals, pharmaceuticals, and pesticides were detected in water at some of the study sites, with the greatest number typically found near domestic wastewater treatment plants. In 2016, there was an increase in concentrations of several herbicides from April to June at upstream agricultural sites. A comparison of chemical concentrations in site water with single chemical data from vitro high-throughput screening (HTS) assays suggested the potential for perturbation of multiple biological pathways, including several associated with induction or inhibition of different cytochrome P450 (CYP) isozymes. This was consistent with direct effects of water extracts in an HTS assay and induction of hepatic CYPs in caged fish. Targeted in vitro assays and measurements in the caged fish suggested minimal effects on endocrine function (e.g., estrogenicity). A nontargeted mass spectroscopy-based analysis suggested that hepatic endogenous metabolite profiles in caged fish covaried strongly with the occurrence of pesticides and pesticide degradates. These studies demonstrate the application of an integrated suite of measurements to help understand the effects of complex chemical mixtures in the field. Environ Toxicol Chem 2021;40:1098-1122. © 2020 SETAC. This article has been contributed to by US Government employees and their work is in the public domain in the USA.

Evaluation of whole-mount in situ hybridization as a tool for pathway-based toxicological research with early-life stage fathead minnows.

Early-life stage fish can be more sensitive to toxicants than adults, so delineating mechanisms of perturbation of biological pathways by chemicals during this life stage is crucial. Whole-mount in situ hybridization (WISH) paired with quantitative real-time polymerase chain reaction (QPCR) assays can enhance pathway-based analyses through determination of specific tissues where changes in gene expression are occurring. While WISH has frequently been used in zebrafish (Danio rerio), this technology has not previously been applied to fathead minnows (Pimephales promelas), another well-established small fish model species. The objective of the present study was to adapt WISH to fathead minnow embryos and larvae, and use the approach to evaluate the effects of estrone, an environmentally-relevant estrogen receptor (ER) agonist. Embryos were exposed via the water to 0, 18 or 1800 ng estrone/L (0, 0.067 and 6.7nM) for 3 or 6 days in a solvent-free, flow-through test system. Relative transcript abundance of three estrogen-responsive genes, estrogen receptor-α (esr1), cytochrome P450-aromatase B (cyp19b), and vitellogenin (vtg) was examined in pooled whole embryos using QPCR, and the spatial distribution of up-regulated gene transcripts was examined in individual fish using WISH. After 3 days of exposure to 1800 ng estrone/L, esr1 and cyp19b were significantly up-regulated, while vtg mRNA expression was not affected. After 6 days of exposure to 1800 ng estrone/L, transcripts for all three genes were significantly up-regulated. Corresponding WISH assays revealed spatial distribution of esr1 and vtg in the liver region, an observation consistent with activation of the hepatic ER. This study clearly demonstrates the potential utility of WISH, in conjunction with QPCR, to examine the mechanistic basis of the effects of toxicants on early-life stage fathead minnows.

Maggot excretion products from the blowfly Lucilia sericata contain contact phase/intrinsic pathway-like proteases with procoagulant functions.

For centuries, maggots have been used for the treatment of wounds by a variety of ancient cultures, as part of their traditional medicine. With increasing appearance of antimicrobial resistance and in association with diabetic ulcers, maggot therapy was revisited in the 1980s. Three mechanisms by which sterile maggots of the green bottle fly Lucilia sericata may improve healing of chronic wounds have been proposed: Biosurgical debridement, disinfecting properties, and stimulation of the wound healing process. However, the influence of maggot excretion products (MEP) on blood coagulation as part of the wound healing process has not been studied in detail. Here, we demonstrate that specific MEP-derived serine proteases from Lucilia sericata induce clotting of human plasma and whole blood, particularly by activating contact phase proteins factor XII and kininogen as well as factor IX, thereby providing kallikrein-bypassing and factor XIa-like activities, both in plasma and in isolated systems. In plasma samples deficient in contact phase proteins, MEP restored full clotting activity, whereas in plasma deficient in either factor VII, IX, X or II no effect was seen. The observed procoagulant/intrinsic pathway-like activity was mediated by (chymo-) trypsin-like proteases in total MEP, which were significantly blocked by C1-esterase inhibitor or other contact phase-specific protease inhibitors. No significant influence of MEP on platelet activation or fibrinolysis was noted. Together, MEP provides contact phase bypassing procoagulant activity and thereby induces blood clotting in the context of wound healing. Further characterisation of the active serine protease(s) may offer new perspectives for biosurgical treatment of chronic wounds.

Field-based approach for assessing the impact of treated pulp and paper mill effluent on endogenous metabolites of fathead minnows (Pimephales promelas).

A field-based metabolomic study was conducted during a shutdown of a pulp and paper mill (PPM) to assess the impacts of treated PPM effluent on endogenous polar metabolites in fathead minnow (FHM; Pimephales promelas) livers. Caged male and female FHMs were deployed at a Great Lakes area of concern during multiple periods (pre-, during, and post-shutdown) near the outflow for a wastewater treatment plant. Influent to this plant is typically 40% PPM effluent by volume. Additional FHMs were exposed to reference lake water under laboratory conditions. A bioassay using T47D-KBluc cells showed that estrogenic activity of receiving water near the outflow declined by 46% during the shutdown. We then used (1)H NMR spectroscopy and principal component analysis to profile abundances of hepatic endogenous metabolites for FHMs. Profiles for males deployed pre-shutdown in receiving water were significantly different from those for laboratory-control males. Profiles were not significantly different for males deployed during the shutdown, but they were significant again for those deployed post-shutdown. Impacts of treated effluent from this PPM were sex-specific, as differences among profiles of females were largely nonsignificant. Thus, we demonstrate the potential utility of field-based metabolomics for performing biologically based exposure monitoring and evaluating remediation efforts occurring throughout the Great Lakes and other ecosystems.

Metabolite profiling and a transcriptional activation assay provide direct evidence of androgen receptor antagonism by bisphenol A in fish.

Widespread environmental contamination by bisphenol A (BPA) has created the need to fully define its potential toxic mechanisms of action (MOA) to properly assess human health and ecological risks from exposure. Although long recognized as an estrogen receptor (ER) agonist, some data suggest that BPA may also behave as an androgen receptor (AR) antagonist. However, direct evidence of this activity is deficient. To address this knowledge gap, we employed a metabolomic approach using in vivo exposures of fathead minnows (FHM; Pimephales promelas ) to BPA either alone or in a binary mixture with 17ß-trenbolone (TB), a strong AR agonist. Changes in liver metabolite profiles in female FHM in response to these exposures were determined using high resolution (1)H NMR spectroscopy and multivariate and univariate statistics. Using this approach, we observed clear evidence of the ability of BPA to mitigate the impact of TB, consistent with an antiandrogenic MOA. In addition, a transcriptional activation assay with the FHM AR was used to confirm the AR antagonistic activity of BPA in vitro. The results of these in vivo and in vitro analyses provide strong and direct evidence for ascribing an antiandrogenic MOA to BPA in vertebrates.

Investigating compensation and recovery of fathead minnow (Pimephales promelas) exposed to 17alpha-ethynylestradiol with metabolite profiling.

1H NMR spectroscopy was used to profile metabolite changes in the livers of fathead minnows (Pimephales promelas) exposed to the synthetic estrogen 17alpha-ethynylestradiol (EE2) via a continuous flow water exposure. Fish were exposed to either 10 or 100 ng EE2/L for 8 days, followed by an 8 day depuration phase. Livers were collected after days 1, 4, and 8 of the exposure, and at the end of the depuration phase. Analysis of polar extracts of the liver revealed a greater impact of EE2 on males than females, with metabolite profiles of the former assuming similarities with those of the females (i.e., feminization) early in the exposure. Biochemical effects observed in the males included changes in metabolites relating to energetics (e.g., glycogen, glucose, and lactate) and liver toxicity (creatine and bile acids). In addition, amino acids associated with vitellogenin (VTG) synthesis increased in livers of EE2-exposed males, a finding consistent with increased plasma concentrations of the lipoprotein in the fish. Using partial least-squares discriminant analysis (PLS-DA), the response trajectories of the males at both exposure concentrations were compared. This revealed an apparent ability of the fish to compensate for the presence of EE2 later in the exposure, and to partially recover from its effects after the chemical was removed.

15-year prospective follow-up study of behavioral therapy in a large sample of inpatients with chronic tinnitus.

CONCLUSION: The results of this study are in accordance with the assumption that cognitive-oriented therapy enabling the patient to live with tinnitus is of primary importance to enhance quality of life. BACKGROUND AND OBJECTIVES: This study describes the success of an integrative behavioral-medicine inpatient treatment for complex chronic tinnitus and presents its long-term effects. In 1987 we developed and evaluated a new treatment concept of psychological treatment of complex chronic tinnitus based on international experience and results. To evaluate the influence, effects and individual results of the specific therapy we analyzed the data of 434 consecutively treated patients. To investigate the long-term effects of the treatment, we contacted the patients 15 years after discharge from the hospital. PATIENTS AND METHODS: We used the tinnitus questionnaire (TQ) and visual analog scales (VAS) for specific tinnitus variables (loudness, discomfort, control of tinnitus, stress, general mood). The German version of the Derogatis psychopathology checklist (SCL-90-R) was used to analyze the impact of additional symptoms (depression, anxiety, introversion, etc.). RESULTS: Compared with a control group (patients on a waiting list) significant and clinically relevant effects were found. At the outcome, there were significant improvements in almost all parameters measured. For evaluation of the long-term effect we succeeded in contacting 312 of 434 former patients. Data were assessed using the same questionnaires that had been employed at the first contact. In all, 271 patients (86%) returned the questionnaires. Data for 244 cases (mean age 63 years; 79 females, 165 males) were complete enough to be used for data analysis. The results of the follow-up were as unexpected as clear: 15 years after conclusion of the treatment, the improvements of the tinnitus parameters and additional symptoms were stable when compared with the end of therapy.

Xenoendocrine disrupters-tiered screening and testing: filling key data gaps.

The US Environmental Protection Agency (EPA) is developing a screening and testing program for endocrine disrupting chemicals (EDCs) to detect alterations of hypothalamic-pituitary-gonadal (HPG) function, estrogen (ER), androgen (AR) and thyroid hormone synthesis and AR and ER receptor-mediated effects in mammals and other animals. High priority chemicals would be evaluated in the Tier 1 Screening (T1S) battery and chemicals positive in T1S would then be tested (Tier 2). T1S includes in vitro ER and AR receptor binding and/or gene expression, an assessment of steroidogenesis and mammalian (rat) and nonmammalian in vivo assays (Table 1). In vivo, the uterotropic assay detects estrogens and antiestrogens, while steroidogenesis, antithyroid activity, (anti)estrogenicity and HPG function are assessed in a 'Pubertal Female Assay'. (Anti-) androgens are detected in the Hershberger Assay (weight of AR-dependent tissues in castrate-immature-male rats). Fish and amphibian assays also are being developed. The fathead minnow assay can identify EDCs displaying several mechanisms of concern, including AR and ER receptor agonists and antagonists and inhibitors of steroid hormone synthesis. An amphibian metamorphosis assay is being developed to detect thyroid-active substances. Several alternative mammalian in vivo assays have been proposed. Of these, a short-term pubertal male rat assay appears most promising. An in utero-lactational screening protocol also is being evaluated. For Tier 2, the numbers of endocrine sensitive endpoints and offspring (F1) examined in multigenerational tests need to be expanded for EDCs. Consideration should be given to tailoring T2, based on the results of T1S. Tier 1 and 2 also should examine relevant mixtures of EDCs. Toxicants that induce malformations in AR-dependent tissues produce cumulative effects even when two chemicals act via different mechanisms of action.

Plasma concentration of ropivacaine after intercostal blocks for video-assisted thoracic surgery.

BACKGROUND: Absorption of local anaesthetics following intercostal blocks is rapid. Therefore, plasma concentrations of ropivacaine during intercostal blocks with ropivacaine 2, 5, 7.5 and 10 mg ml-1 (ropivacaine 5 ml injected into each of four intercostal spaces) in patients undergoing video-assisted thoracic surgery were determined. METHODS: After informed consent and ethics committee approval, 64 patients were randomly allocated to four groups for intercostal nerve block (ropivacaine 2, 5, 7.5 or 10 mg ml-1 at the end of surgery). Central (mixed) venous and arterial plasma samples were collected before the start of intercostal application, and 2, 5, 10, 15, 20, 30, 45, 60 and 90 min afterwards. Plasma concentrations of ropivacaine were measured by high performance liquid chromatography. RESULTS: Maximum venous plasma concentrations occurred after the mean times of 10.7 (range, 5-15), 10.8 (5-20), 11.3 (5-20) and 12.2 (5-45) min, respectively for each group. The groups had mean concentrations of 1.3 (SD, 0.6; range, 0.3-2.3), 2.1 (1.0; 0.5-4.5), 2.4 (1.0; 1.2-5.1) and 2.5 (0.9; 1.7-5.6) micrograms ml-1, respectively. Maximum arterial plasma concentration following 1.0% ropivacaine occurred after 16 (5-45) min with a mean of 2.3 (0.6; 1.5-3.6) micrograms ml-1. No signs of central nervous system or cardiac toxicity were observed. CONCLUSIONS: After intercostal blocks the absorption of ropivacaine is rapid compared with other techniques for regional anaesthesia and results in relatively high venous and arterial plasma concentrations, especially if a dose of 100 mg or more is used.

[Postoperative pain therapy in minimally invasive direct coronary arterial bypass surgery. I.v. opioid patient-controlled analgesia versus intercostal block]. / Postoperative Schmerztherapie bei minimal-invasiver direkter koronararterieller Bypass-Chirurgie (MIDCAB). I.v.-Opioid-PCA versus Interkostalblockaden.

OBJECTIVE: Lately introduced cardiosurgical procedures such as MIDCAB enable an early extubation immediately after surgery. This also requires an adequate anesthesia regime and especially a sufficient postoperative analgesia. Patient controlled analgesia (PCA) and intercostal nerve blockade (ICB) were evaluated for their suitability for postoperative pain relief in patients undergoing a MIDCAB procedure. MATERIAL AND METHODS: After approval by the local ethic committee and obtaining written informed consent 43 patients were included in this study. Anesthesia was induced and maintained in a total intravenous standardised manner with propofol, remifentanil, cisatracurium and additionally glyceroltrinitrate, clonidine and esmolol were given as needed. After revascularisation patients were randomly assigned to one of two groups receiving either 7.5 mg piritramid i.v. before extubation and continuing a PCA with 2 mg boli and a 10 min lockout, or an ICB with ropivacaine 1% (4 times 5 ml). Additionally all patients received 1 g paracetamol rectally before induction of anesthesia and 1 g metamizol i.v. at the end of surgery. A rescue medication of 3.75 mg piritramid i.v. was allowed. A pain score (NRS 0-10), the Aldrete score (AS 0-12) and oxygen saturation were obtained 1, 4, and 8 h after extubation. RESULTS: The ICB group showed a significantly greater pain reduction in the first (5.8 +/- 1.8 vs. 7.3 +/- 1.9; P < 0.02) and fourth h (3.6 +/- 1.3 vs. 4.6 +/- 1.4; P < 0.02), respectively. Transfer to an intermediate care ward one hr after extubation was achieved more often in the ICB group according to the AS (ICB 9.6 +/- 1.5 vs. PCA 8.9 +/- 1.2; P < 0.05), too. There was no difference with respect to the oxygen saturation. The additional piritramid demand was 9.3 mg in the ICP group and 5 mg in the PCA group in the first 8 hours postoperative. CONCLUSION: ICB gives a better pain relief in the early postoperative phase after MIDCAB procedures compared to a PCA. Both regimes are adequate in order to provide a sufficient pain relief and help to avoid prolonged postoperative mechanical ventilation. These will enable an early transfer of patients to an intermediate care station and save ICU capacity.

Quantitative competitive NASBA for measuring mRNA expression levels of the immediate early 1, late pp67, and immune evasion genes US3, US6 and US11 in cells infected with human cytomegalovirus.

Different cell types were infected with human cytomegalovirus (HCMV) and RNA expression dynamics were analyzed by quantitative NASBA assays for IE1 (UL123), pp67 (UL65) and the immune evasion genes (US3, US6 and US11). The quantitative NASBA assays gave reproducible quantification in the range of 10(3)-10(6) RNA copies for IE1 and pp67 RNA, from 3x10(3) to 1x10(6) RNA copies for US6 and US11 RNA, and from 1x10(4) to 1x10(6) RNA copies for US3 RNA. SMC, HAEC and HUVEC cells infected with an, in endothelial cells, propagated HCMV strain (VHL/E) showed similar RNA expression dynamics for the analyzed genes. Expression of all genes studied was observed within the first 4 h post-infection. The first gene for which expression could be detected was IE1, followed by US3, US11, pp67 and US6. Fibroblasts infected with HCMV strain AD169 showed a different RNA expression pattern for US3. Translation of the mRNA studied was demonstrated by detection of the proteins 48 h post-infection by immunofluorescence.

Description and evaluation of a short-term reproduction test with the fathead minnow (Pimephales promelas).

Due to the time and expense associated with full life-cycle testing, most current toxicity tests with fish do not explicitly consider reproductive output as an endpoint but, rather, focus on early life-stage survival and development. However, some classes of chemicals could adversely impact reproduction at concentrations below those that affect development. Further, estimates of the effects of toxic compounds on reproductive output can be critical to the ecological risk assessment process. In this manuscript, we describe a short-term reproduction test with the fathead minnow (Pimephales promelas) and evaluate the test using two model reproductive toxicants, methoxychlor (an estrogenic compound) and methyltestosterone (an androgenic chemical). The test is initiated with reproductively mature animals and is comprised of a pre-exposure phase of 14 to 21 d, followed by a chemical exposure of up to 21 d. During and at completion of the test, several endpoints related to reproductive fitness and endocrine function are assessed. Both chemicals evaluated in our study caused a significant decrease in fecundity of the fish at nominal concentrations of 5.0 micrograms/L (methoxychlor) and 0.2 mg/L (methyltestosterone). Methoxychlor decreased plasma concentrations of one or more steroids (testosterone, 11-ketotestosterone, beta-estradiol) in both sexes and caused a significant induction of plasma vitellogenin in males, a response consistent with activation of the estrogen receptor by the pesticide (or its metabolites). Methyltestosterone decreased plasma concentrations of sex steroids and adversely affected gonadal status (as evaluated by relative weight and histopathology) in both sexes. The androgenic nature of methyltestosterone was clearly expressed as masculinization of exposed females via formation of nuptial tubercles, structures normally present only in reproductively active males. The chemical also caused a significant induction of plasma vitellogenin in both males and females; this unexpected estrogenic response was most likely due to aromatization of the androgen to a form capable of binding to the estrogen receptor. These studies demonstrate the utility of this short-term assay for identifying chemicals that exert reproductive toxicity through alterations in endocrine systems controlled by estrogens and androgens.

Aspects of basic reproductive biology and endocrinology in the fathead minnow (Pimephales promelas).

The fathead minnow (Pimephales promelas) has been proposed as a model species for assessing the adverse effects of endocrine-disrupting chemicals (EDCs) on reproduction and development. The purpose of these studies was to develop baseline reproductive biology and endocrinology data for this species to support interpretation of tests with potential EDCs. Pairs of reproductively-active fathead minnows (n=70) were evaluated with respect to reproductive cyclicity in terms of spawning interval and fecundity. The mode and mean (+/-SE) spawning intervals for the fish in this study were 3.0 and 3.7+/-0.1 days, respectively. The mean number of eggs produced per spawn was 85+/-2.8. Animals were sacrificed at periodic intervals during the established spawning cycle and measurements made of gonadal condition (gonadosomatic index [GSI], histopathology) and plasma concentrations of vitellogenin and sex steroids (beta-estradiol, testosterone, 11-ketotestosterone). The GSI in females varied significantly as a function of spawning interval, with the largest values occurring day 2 post-spawn, just prior to the interval of maximum spawning activity. Plasma beta-estradiol concentrations in females also varied significantly relative to peak values in the GSI and spawning activity. Vitellogenin concentrations in the female, and male GSI and steroid concentrations did not vary significantly relative to position in the spawning cycle. Concentrations of beta-estradiol in females and 11-ketotestosterone in males were positively correlated with testosterone concentrations.

Tropism of human cytomegalovirus for endothelial cells is determined by a post-entry step dependent on efficient translocation to the nucleus.

Marked interstrain differences in the endothelial cell (EC) tropism of human cytomegalovirus (HCMV) isolates have been described. This study aimed to define the step during the replicative cycle of HCMV that determines this phenotype. The infection efficiency of various HCMV strains in EC versus fibroblasts was quantified by immunodetection of immediate early (IE), early and late viral antigens. Adsorption and penetration were analysed by radiolabelled virus binding assays and competitive HCMV-DNA-PCR. The translocation of penetrated viral DNA to the nucleus of infected cells was quantified by competitive HCMV-DNA-PCR in pure nuclear fractions. The intracytoplasmic translocation of capsids that had penetrated was followed by immunostaining of virus particles on a single particle level; this was correlated with the initiation of viral gene expression by simultaneous immunostaining of viral IE antigens. The infectivity of nonendotheliotropic HCMV strains in EC was found to be 100-1000-fold lower when compared to endotheliotropic strains. The manifestation of this phenotype at the level of IE gene expression indicated the importance of initial replication events. Surprisingly, no interstrain differences were detected during virus entry. However, dramatic interstrain differences were found regarding the nuclear translocation of penetrated viral DNA. With nonendotheliotropic strains, the content of viral DNA in the cell nucleus was 100-1000-fold lower in EC when compared to endotheliotropic strains, thereby reflecting the strain differences in IE gene expression. Simultaneous staining of viral particles and viral IE antigen revealed that interstrain differences in the transport of penetrated capsids towards the nucleus of endothelial cells determine the EC tropism of HCMV.

Efficient lytic infection of human arterial endothelial cells by human cytomegalovirus strains.

Endothelial cells (EC) are common targets of permissive infection by human cytomegalovirus (HCMV) in vivo during acute disease. However, studies of HCMV-EC interactions in vitro have generated discordant results. While lytic infection of cultured venous EC has been well established, Fish et al. (K. N. Fish, C. Soderberg Naucler, L. K. Mills, S. Stenglein, and J. A. Nelson, J. Virol. 72:5661-5668) have reported noncytopathic persistence of the virus in cultured aortic EC. We propose that interstrain differences in viral host cell tropism rather than the vascular bed of origin of infected EC might account for these discrepancies. In the present investigation we compared the responses of EC derived from human adult iliac artery, placental microvasculature, and umbilical vein to infection with various HCMV strains. Regardless of the vascular bed of origin, infection with EC-propagated HCMV strains induced 100% efficient cytopathic change progressing to complete lysis of inoculated monolayers. While fibroblast-propagated strains persisted at low titer in infected arterial EC cultures, they were also cytolytic for individual infected cells. The finding of cytopathic lytic infection of arterial EC by HCMV implicates a mechanism of vascular injury in the pathogenesis of HCMV infection.

Prospective evaluation of morphology, function, and quality of life after extracorporeal shockwave lithotripsy and endoscopic treatment of chronic calcific pancreatitis.

OBJECTIVE: Therapeutic endoscopy may be effective in selected patients with chronic calcific pancreatitis (CCP). We prospectively evaluated the early outcome of extracorporeal shockwave lithotripsy (ESWL) in combination with interventional endoscopy, using broad inclusion criteria. METHODS: A total of 48 consecutive patients (35 male, 13 female) were recruited for ESWL and endoscopic therapy of symptomatic CCP. Symptoms, quality of life, pancreatic morphology and function were assessed before and after. RESULTS: Multiple stones (n = 43), strictures (n = 34), and pancreas divisum (n = 11) were found. A median of 13 ESWL sessions (range 2-74) with a median of 22,100 shockwaves (1,700-150,900) were required. Endoscopic pancreatic sphincterotomy (n = 48), stricture dilation (n = 12), and/or stenting (n = 27) were performed. After therapy, drainage of the pancreatic duct system was achieved in 36, complete stone clearance in 21 patients. Follow-up (n = 38) at 7 months (range 5-9) showed a significant decrease in pancreatic duct diameter (p < 0.001) and pain score (p < 0.0001) whereas complete pain relief was observed in 45% of cases. Several quality of life scores improved significantly. Weight gain occurred in 68% of patients. Normalization of fasting blood glucose and HbA1c levels were observed in four patients, without modifying their treatment. Improvement in pain score correlated with weight gain and decrease in pancreatic duct diameter. Nonalcoholic etiology was associated with a better chance for improvement in pain score and decrease in pancreatic duct diameter. The presence of strictures did not deteriorate the clinical outcome. CONCLUSIONS: Besides pain relief, ESWL in combination with interventional endoscopy resulted in pancreatic ductal decompression, weight gain, and improvement in quality of life in a considerable number of patients with advanced CCP.

Modification of human cytomegalovirus tropism through propagation in vitro is associated with changes in the viral genome.

Following extensive propagation in fibroblasts, human cytomegalovirus (HCMV) loses tropism for a number of otherwise natural host cells, in particular, endothelial cells. In this study, the hypothesis was tested that loss of endothelial tropism is associated with the appearance of genomic variants. Initial quantitative focus expansion assays on endothelial monolayers demonstrated that, while the laboratory strains AD169 and Towne failed to form detectable foci, 29 out of 30 recent clinical HCMV isolates had the potential to expand in endothelial cell culture. By long-term adaptation in fibroblast cultures, nonendotheliotropic strains could be selected from clinical HCMV isolates, while long-term endothelial-adapted strains of the same isolates retained both fibroblast tropism and endothelial tropism. Such differentially adapted isolate pairs always displayed genomic differences in restriction fragment length analyses. Coinfection of endothelial cells by two nonendotheliotropic HCMV strains yielded an endotheliotropic recombinant HCMV variant combining portions of the genomes of both parental viruses. When DNA purified from various isolates was transfected into fibroblasts, progeny virus retained the specific tropism of parental virus from which the DNA was isolated. These findings demonstrate that endothelial tropism is an inherent property of most clinical HCMV isolates and is determined by the viral genome. Although the specific determinants of HCMV cell tropism are still unknown, this study provides the first evidence for a genetic contribution.

A simple and rapid method for preparation of viral DNA from cell associated cytomegalovirus.

In the field of human cytomegalovirus pathogenesis there is growing interest in analyzing recent clinical isolates rather than cell culture adapted laboratory strains. However, true low passage isolates are strictly cell associated prior to cell culture adaptation and only a minor fraction of cells are infected at low passage number. Both conditions hinder the preparation of pure viral DNA. To date, genetic analyses had been carried out mostly with supernatant associated cytomegalovirus. A rapid and simple method is described for preparation of viral DNA from low passage cell associated isolates with little cytopathogenic effect. The protocol is based on a combination of Triton X-100 lysis, nuclease treatment, and subsequent phenol chloroform extraction. Cellular background was reduced significantly to enable clear detection of all viral DNA fragments in restriction fragment length analysis. The method yielded DNA which was suitable for downstream applications like cloning of viral DNA fragments or transfection of genomic viral DNA. This method may facilitate genomic analyses of pathogenic cell associated recent cytomegalovirus isolates.

Saturation units for use in aquatic bioassays.

Methods were developed for preparing liquid/liquid and glass wool column saturators for generating chemical stock solutions for conducting aquatic bioassays. Exposures have been conducted using several species of fish, invertebrate, and mollusks in static and flow-through conditions using these methods. Stock solutions for 82 organic chemicals were prepared using these saturation units. The primary purpose of stock generation was to provide a continuous and consistent amount of toxicant laden solution at a measured analytical level which would be available to test organisms for the test duration. In the present study, the glass wool column and liquid/liquid saturators were used to provide consistent stock concentrations, at times approaching saturation, for fathead minnow (Pimephales promelas) acute exposures. Attempts were made to achieve the maximum solubility of these compounds for comparison purposes to water solubility values available in the literature. Literature solubility values from a database by Yalkowsky et al. [1] provided information on temperatures and data quality which allowed comparison to values obtained from the present study. Twenty four compounds were identified and analyzed for the comparison of maximum obtainable solubility levels. Maximum saturator stock water concentrations were generally lower (R = 0.98) but were in close agreement with published water solubility values.