Octavia M. Peck Palmer

Metallothionein-induced zinc partitioning exacerbates hyperoxic acute lung injury.

Hypozincemia, with hepatic zinc accumulation at the expense of other organs, occurs in infection, inflammation, and aseptic lung injury. Mechanisms underlying zinc partitioning or its impact on extrahepatic organs are unclear. Here we show that the major zinc-binding protein, metallothionein (MT), is critical for zinc transmigration from lung to liver during hyperoxia and preservation of intrapulmonary zinc during hyperoxia is associated with an injury-resistant phenotype in MT-null mice. Particularly, lung-to-liver zinc ratios decreased in wild-type (WT) and increased significantly in MT-null mice breathing 95% oxygen for 72 h. Compared with female adult WT mice, MT-null mice were significantly protected against hyperoxic lung injury indicated by reduced inflammation and interstitial edema, fewer necrotic changes to distal airway epithelium, and sustained lung function at 72 h hyperoxia. Lungs of MT-null mice showed decreased levels of immunoreactive LC3, an autophagy marker, compared with WT mice. Analysis of superoxide dismutase (SOD) activity in the lungs revealed similar levels of manganese-SOD activity between strains under normoxia and hyperoxia. Lung extracellular SOD activity decreased significantly in both strains at 72 h of hyperoxia, although there was no difference between strains. Copper-zinc-SOD activity was ~4× higher under normoxic conditions in MT-null compared with WT mice but was not affected in either group by hyperoxia. Collectively the data suggest that genetic deletion of MT-I/II in mice is associated with compensatory increase in copper-zinc-SOD activity, prevention of hyperoxia-induced zinc transmigration from lung to liver, and hyperoxia-resistant phenotype strongly associated with differences in zinc homeostasis during hyperoxic acute lung injury.

Effects of Transport Temperature on the Stability of Inflammatory, Hemostasis, Endothelial Function, and Oxidative Stress Plasma Biomarker Concentrations

Background: A number of studies in critically ill patients are conducted outside the hospital. Specimens should ideally be transported from out-of-hospital setting to a laboratory using dry ice, but this approach is expensive and may not be feasible in some circumstances. We, therefore, examined the impact of temperature during transport of specimens on the precision of biomarker concentrations. Objective: To determine the effects of transport temperature conditions on biomarker concentrations in specimens processed within 1 h of collection. Methods/Patients: We simulated transport by storing specimens at four temperature conditions: packaged at −80°C (control), on dry ice (−79°C), on cold gel packs (4°C), and at room temperature (RT, 21°C). We examined eight biomarkers spanning four signaling domains— inflammation, hemostasis, endothelial dysfunction, and oxidative stress. We calculated mean, median, and percent difference for each biomarker concentration compared with the control transport temperature at −80°C in 26 subjects (16 hospitalized with severe sepsis and 10 non-hospitalized volunteers). Results: Patients with severe sepsis had log-fold higher median concentrations of IL-6, hs-CRP, D-dimer, E-selectin, sICAM-1, and sVCAM-1 compared with non-hospitalized volunteers (P <0.05). When specimens were combined, we observed a ⩽7% difference in the mean and median IL-6, hs-CRP, D-dimer, PAI-1, E-selectin, s-ICAM, s-VCAM, and nitrite concentrations for dry ice and cold gel packs transport compared with transport at −80°C (P>0.05). Larger differences (up to 12%) were observed when biomarker concentrations for PAI-1 and s-VCAM at room temperature were compared with transport at −80°C (P >0.05). Conclusions: Select inflammatory, coagulation, endothelial dysfunction, and oxidative stress biomarkers can be transported at 4°C on gel packs for 24 h with minimal effects on precision.

Challenges and Opportunities in Implementing Total Laboratory Automation

In total laboratory automation (TLA)10, preanalytic, analytic, and postanalytic phases of laboratory testing may be combined into an integrated system such that specimens are processed, tested, and even stored with minimal user intervention. Given the pressures of an ongoing workforce shortage of laboratory professionals, laboratory automation offers an attractive, albeit expensive, solution that laboratories are increasingly considering in planning for future growth and work flow requirements. In an ideal system, TLA handles routine, repetitive steps—leveraging the quality and efficiency obtainable in the manufacturing industry and freeing operators to focus on specialized testing that benefits from their unique training and expertise. A variety of laboratory automation solutions have been available globally for decades, with technologies that have advanced based on engineering innovation and practical trial and error. To address the difficulties and benefits involved in implementing and sustaining TLA in a clinical laboratory setting, we invited a group of 5 experts to share their perspectives on laboratory automation and provide real-world advice based on their experiences with TLA at their respective facilities. In what ways has automation impacted your clinical laboratory? Giuseppe Lippi: Automation has completely revolutionized the organization of my laboratory. The main advantages that we have observed are improved standardization, more simplified and efficient approaches for managing work flow, improvement in the performance of complex tests, better high-volume test management, shorter turnaround time (TAT) by elimination of some manually intensive preanalytical steps, provision of valuable walk-away time, and reduction of personnel costs (i.e., laboratory technicians and subsidiary staff), along with notable reductions in errors and biological risks attributable to manual handling of specimens. Another important advantage is represented by more efficient management of reruns and reflex testing, which can now be performed automatically with little user action needed. Carey-Ann Burnham: Historically, the clinical microbiology laboratory has lagged …

Ranitidine interference with standard amphetamine immunoassay.

BACKGROUND We recently encountered several cases of possible false positive results of amphetamine on the Beckman Coulter AMPH assay, but not on the Siemens EMIT II Plus assay. Our clinical chart review suggested that ranitidine interference may be responsible for the false positive results of the AMPH assays. METHODS Blank urine specimens spiked with ranitidine concentrations ranging from 5μg/ml to 5mg/ml were analyzed on both the AMPH and EMIT II Plus assays. To examine if the false positive results were due to assay specific reagent/sample ratios, we prepared 3 different sample to reagent ratios and analyzed them for amphetamine reaction rates on both assays. RESULTS Ranitidine at 160μg/ml caused a positive interference on the AMPH assay. No interference was observed by ranitidine on the EMIT II Plus assay. Specifically, the sample to reagent ratios tested neither eliminated the positive inference on the AMP assay nor introduced an interference on the EMIT II Plus assay. CONCLUSIONS Unlike the EMIT II Plus assay, the AMPH assay has cross-activity with ranitidine, which is independent of sample to reagent ratio.

Elaidate, an 18-Carbon Trans-monoenoic Fatty Acid, but not Physiological Fatty Acids Increases Intracellular Zn2+ in Human Macrophages

Artificial trans fatty acids promote atherosclerosis by blocking macrophage clearance of cell debris. Classical fatty‐acid response mechanisms include TLR4‐NF‐κB activation, and Erk1/2 phosphorylation, but these may not indicate long‐term mechanisms. Indeed, nuclear NF‐κB was increased by 60 min treatment by 30 μM of the 18 carbon trans unsaturated fatty acid elaidic acid (elaidate), the physiological cis‐unsaturated fatty acid oleic acid (oleate), and the 18 or 16 carbon saturated fatty acids stearic and palmitic acid (stearate or palmitate). However, except for stearate, effects on related pathways were minimal at 44 h. To determine longer term effects of trans fatty acids, we compared mRNA expression profiles of (trans) elaidate to (cis) oleate, 30 μM, at 44 h in human macrophages. We found that elaidate changed Zn2+‐homeostasis gene mRNAs markedly. This might be important because Zn2+ is a major regulator of macrophage activity. Messenger RNAs of seven Zn2+‐binding metallothioneins decreased 2–4‐fold; the zinc importer SLC39A10 increased twofold, in elaidate relative to oleate‐treated cells. Results were followed by quantitative PCR comparing cis, trans, and saturated fatty acid effects on Zn2+‐homeostasis gene mRNAs. This confirmed that elaidate uniquely decreased metallothionein expression and increased SLC39A10 at 44 h. Further, intracellular Zn2+ was measured using N‐(carboxymethyl)‐N‐[2‐[2‐[2(carboxymethyl) amino]‐5‐(2,7,‐difluoro‐6‐hydroxy‐3‐oxo‐3H‐xanthen‐9‐yl)‐phenoxy]‐ethoxy]‐4‐methoxyphenyl]glycine, acetoxymethyl ester (FluoZin‐3‐AM). This showed that, at 44 h, only cells treated with elaidate had increased Zn2+. The durable effect of elaidate on Zn2+ activation is a novel and specific effect of trans fatty acids on peripheral macrophage metabolism. J. Cell. Biochem. 116: 524–532, 2015.

Plasma Abnormalities Following Overdose

A 62-year-old female with a history of schizophrenia was found unresponsive after ingesting supratherapeutic amounts of diltiazem and valsartan. At presentation to the emergency department, she was hypotensive and bradycardic. Eight hours later, a blood sample (Fig. 1A) was drawn for chemistry analysis and ultracentrifuged to clarify the turbidity (Fig. 1B). However, the lab was unable to assay …

Graph Theoretical Analysis of Genome-Scale Data: Examination of Gene Activation Occurring in the Setting of Community-Acquired Pneumonia

Introduction: We have previously reported evidence that Black individuals appear to have a significantly higher incidence of infection-related hospitalizations compared with White individuals. It is possible that the host immune response is responsible for this vital difference. In support of such a hypothesis, the aim of this study was to determine whether Black and White individuals exhibit differential whole blood gene network activation. Methods: We examined whole blood network activation in a subset of patients (n = 22 pairs, propensity score matched (1:1) Black and White patients) with community-acquired pneumonia (CAP) from the Genetic and Inflammatory Markers of Sepsis study. We employed day one whole blood transcriptomic data generated from this cohort and constructed co-expression graphs for each racial group. Pearson correlation coefficients were used to weight edges. Spectral thresholding was applied to ascribe significance. Innovative graph theoretical methods were then invoked to detect densely connected gene networks and provide differential structural analysis. Results: Propensity matching was employed to reduce potential bias due to confounding variables. Although Black and White patients had similar socio- and clinical demographics, we identified novel differences in molecular network activation—dense subgraphs known as paracliques that displayed complete gene connection for both White (three paracliques) and Black patients (one paraclique). Specifically, the genes that comprised the paracliques in the White patients include circadian loop, cell adhesion, mobility, proliferation, tumor suppression, NF&kgr;B, and chemokine signaling. However, the genes that comprised the paracliques in the Black patients include DNA and messenger RNA processes, and apoptosis signaling. We investigated the distribution of Black paracliques across White paracliques. Black patients had five paracliques (with almost complete connection) comprised of genes that are critical for host immune response widely distributed across 22 parcliques in the White population. Anchoring the analysis on two critical inflammatory mediators, interleukin (IL)-6 and IL-10 identified further differential network activation among the White and Black patient populations. Conclusions: These results demonstrate that, at the molecular level, Black and White individuals may experience different activation patterns with CAP. Further validation of the gene networks we have identified may help pinpoint genetic factors that increase host susceptibility to community-acquired pneumonia, and may lay the groundwork for personalized management of CAP.

Assuming It Was There, Where Did It Go?

A 60-year-old woman with epilepsy and a long-term alcohol addiction presented to the emergency department with a 2-day history of nausea and vomiting. On day 2 of hospitalization, the patient complained of muscle weakness, dizziness, and intermittent abdominal pain. Laboratory results are shown in Table 1 …

A 72-Year-Old Woman with Markedly Increased Ferritin

A 72-year-old woman with a history of adult-onset Still disease and congestive heart failure was transferred from an outside hospital. She had a 2-week history of fevers, diarrhea, nausea, sore throat, and myalgias. At presentation, she had a markedly increased plasma ferritin concentration [238 380 ng/mL (reference intervals shown in Table 1)], low hemoglobin …

Cyst, What Art Thou?

A 51-year-old woman, with a medical history of abnormal Pap smears, extended menstrual cycles, and Barretts esophagus, underwent an upper gastrointestinal endoscopic ultrasound for abnormal findings on a computed tomography scan. The ultrasound characterized a peripherally calcified, multicystic, and septated lesion that was close to both the pancreatic tail and the upper pole of the left …