Searching for Life in Hot Spring Carbonate Systems: Investigating Raman Spectra of Carotenoid-Bearing Organic Carbonaceous Inclusions from Travertines of Italy.

Carotenoid pigments provide some of the most common exclusively biogenic markers on Earth, and these organic pigments may be present in extraterrestrial life. Raman spectroscopy can be used to identify carotenoids quickly and accurately through the inelastic scattering of laser light. In this study, we show that Raman spectra of organic matter found in hot spring bacterial assemblages exhibit "spectral overprinting" of the carotenoid spectrum by the carbon spectrum as the organic matter progressively breaks down. Here, we present how, with increasing thermal maturity, the relative intensity of the carotenoid spectrum increases, and as maturity increases a low-intensity carbon spectrum forms in the same region as the carotenoid spectrum. This carbon spectrum increases in intensity as the thermal maturity increases further, progressively obscuring the carotenoid spectrum until only the carbon spectrum can be observed. This means key carotenoid biogenic signatures in hot spring deposits may be hidden within carbon spectra. A detailed study of the transition from carotenoid to carbon, Raman spectra may help develop deconvolution processes that assist in positively identifying biogenic carbon over abiogenic carbon. Our results are relevant for the data analysis from the Raman spectroscopy instruments on the Perseverance (National Aeronautics and Space Administration [NASA]) and Rosalind Franklin (European Space Agency [ESA]) rovers.

Establishment of a whole slide imaging-based frozen section service at a cancer center.

Background: In recent years, there has been a surge of interest in clinical digital pathology (DP). Hardware and software platforms have matured and become more affordable, and advances in artificial intelligence promise to transform the practice of pathology. At our institution, we are launching a stepwise process of DP adoption which will eventually encompass our entire workflow. Out of necessity, we began by establishing a whole slide imaging (WSI)-based frozen section service. Methods: We proceeded in a systematic manner by first assembling a team of key stakeholders. We carefully evaluated the various options for digitizing frozen sections before deciding that a WSI-based solution made the most sense for us. We used a formalized evaluation system to quantify performance metrics that were relevant to us. After deciding on a WSI-based system, we likewise carefully considered the various whole slide scanners and digital slide management systems available before making decisions. Results: During formal evaluation by pathologists, the WSI-based system outperformed competing platforms. Although implementation was relatively complex, we have been happy with the results and have noticed significant improvements in our frozen section turnaround time. Our users have been happy with the slide management system, which we plan on utilizing in future DP efforts. Conclusions: There are various options for digitizing frozen section slides. Although WSI-based systems are more complex and expensive than some alternatives, they perform well and may make sense for institutions with a pre-existing or planned larger DP infrastructure.

Raman spectroscopy: current applications in breast cancer diagnosis, challenges and future prospects.

Despite significant improvements in the way breast cancer is managed and treated, it continues to persist as a leading cause of death worldwide. If detected and diagnosed early, when tumours are small and localised, there is a considerably higher chance of survival. However, current methods for detection and diagnosis lack the required sensitivity and specificity for identifying breast cancer at the asymptomatic or very early stages. Thus, there is a need to develop more rapid and reliable methods, capable of detecting disease earlier, for improved disease management and patient outcome. Raman spectroscopy is a non-destructive analytical technique that can rapidly provide highly specific information on the biochemical composition and molecular structure of samples. In cancer, it has the capacity to probe very early biochemical changes that accompany malignant transformation, even prior to the onset of morphological changes, to produce a fingerprint of disease. This review explores the application of Raman spectroscopy in breast cancer, including discussion on its capabilities in analysing both ex-vivo tissue and liquid biopsy samples, and its potential in vivo applications. The review also addresses current challenges and potential future uses of this technology in cancer research and translational clinical application.

Dubiofossils from a Mars-analogue subsurface palaeoenvironment: The limits of biogenicity criteria.

The search for a fossil record of Earth's deep biosphere, partly motivated by potential analogies with subsurface habitats on Mars, has uncovered numerous assemblages of inorganic microfilaments and tubules inside ancient pores and fractures. Although these enigmatic objects are morphologically similar to mineralized microorganisms (and some contain organic carbon), they also resemble some abiotic structures. Palaeobiologists have responded to this ambiguity by evaluating problematic filaments against checklists of "biogenicity criteria". Here, we describe material that tests the limits of this approach. We sampled Jurassic calcite veins formed through subseafloor serpentinization, a water-rock reaction that can fuel the deep biosphere and is known to have occurred widely on Mars. At two localities ~4 km apart, veins contained curving, branched microfilaments composed of Mg-silicate and Fe-oxide minerals. Using a wide range of analytical techniques including synchrotron X-ray microtomography and scanning transmission electron microscopy, we show that these features meet many published criteria for biogenicity and are comparable to fossilized cryptoendolithic fungi or bacteria. However, we argue that abiotic processes driven by serpentinization could account for the same set of lifelike features, and report a chemical garden experiment that supports this view. These filaments are, therefore, most objectively described as dubiofossils, a designation we here defend from criticism and recommend over alternative approaches, but which nevertheless signifies an impasse. Similar impasses can be anticipated in the future exploration of subsurface palaeo-habitats on Earth and Mars. To avoid them, further studies are required in biomimetic geochemical self-organization, microbial taphonomy and micro-analytical techniques, with a focus on subsurface habitats.

The Apoptotic Crypt Abscess: An Underappreciated Histologic Finding in Gastrointestinal Pathology.

OBJECTIVES: To differentiate apoptotic crypt abscesses (ACAs) from neutrophilic crypt abscesses (NCAs). METHODS: Cases with crypt abscesses were classified as containing ACAs, NCAs, or mixed crypt abscesses (MCAs) by H&E staining. Sections were stained with cleaved caspase 3 and myeloperoxidase and recategorized. RESULTS: Fifty-nine cases were reviewed: inflammatory bowel disease (IBD; n = 33), acute cellular rejection (n = 5), graft vs host disease (GVHD; n = 14), cytomegalovirus (n = 5), and drug reaction (n = 2). Concordance was seen in 59%, with most reclassifications resulting from a change of ACAs to MCAs. When cases were classified as having NCA vs those with apoptosis (ACA and MCA), there was 85% agreement (P < .01). NCAs were present in IBD (96%) and not in GVHD or drug injury. Crypt abscesses with apoptosis were seen in 18% of IBD and 96% of non-IBD cases. CONCLUSIONS: ACAs and MCAs can be distinguished from NCAs and may be a diagnostically useful finding.

Endocytic recycling protein EHD1 regulates primary cilia morphogenesis and SHH signaling during neural tube development.

Members of the four-member C-terminal EPS15-Homology Domain-containing (EHD) protein family play crucial roles in endocytic recycling of cell surface receptors from endosomes to the plasma membrane. In this study, we show that Ehd1 gene knockout in mice on a predominantly B6 background is embryonic lethal. Ehd1-null embryos die at mid-gestation with a failure to complete key developmental processes including neural tube closure, axial turning and patterning of the neural tube. We found that Ehd1-null embryos display short and stubby cilia on the developing neuroepithelium at embryonic day 9.5 (E9.5). Loss of EHD1 also deregulates the ciliary SHH signaling with Ehd1-null embryos displaying features indicative of increased SHH signaling, including a significant downregulation in the formation of the GLI3 repressor and increase in the ventral neuronal markers specified by SHH. Using Ehd1-null MEFS we found that EHD1 protein co-localizes with the SHH receptor Smoothened in the primary cilia upon ligand stimulation. Under the same conditions, EHD1 was shown to co-traffic with Smoothened into the developing primary cilia and we identify EHD1 as a direct binding partner of Smoothened. Overall, our studies identify the endocytic recycling regulator EHD1 as a novel regulator of the primary cilium-associated trafficking of Smoothened and Hedgehog signaling.

Circulatory changes associated with the closure of the ductus arteriosus in hatching emu (Dromaius novaehollandiae).

In developing avian embryos, the right and left ductus arteriosi (DA) allow for a shunt of systemic venous return away from the lungs to the body and chorioallantoic membrane (CAM). Unlike in mammals where the transition from placental respiration to lung respiration is instantaneous, in birds the transition from embryonic CAM respiration to lung respiration can take over 24h. To understand the physiological consequences of this long transition we examined circulatory changes and DA morphological changes during hatching in the emu (Dromaius novaehollandiae), a primitive ratite bird. By tracking microspheres injected into a CAM vein, we observed no change in DA blood flow between the pre-pipped to internally pipped stages. Two hours after external pipping, however, a significant decrease in DA blood flow occurred, evident from a decreased systemic blood flow and subsequent increased lung blood flow. Upon hatching, the right-to-left shunt disappeared. These physiological changes in DA blood flow correspond with a large decrease in DA lumen diameter from the pre-pipped stages to Day 1 hatchlings. Upon hatching, the right-to-left shunt disappeared and at the same time apoptosis of smooth muscle cells began remodeling the DA for permanent closure. After the initial smooth muscle contraction, the lumen disappeared as intimal cushioning formed, the internal elastic lamina degenerated, and numerous cells underwent regulated apoptosis. The DA closed rapidly between the initiation of external pipping and hatching, resulting in circulatory patterns similar to the adult. This response is most likely produced by increased DA constriction in response to increased arterial oxygen levels and the initiation of vessel remodeling.

Restoration of compact Golgi morphology in advanced prostate cancer enhances susceptibility to galectin-1-induced apoptosis by modifying mucin O-glycan synthesis.

UNLABELLED: Prostate cancer progression is associated with upregulation of sialyl-T antigen produced by ß-galactoside α-2,3-sialyltransferase-1 (ST3Gal1) but not with core 2-associated polylactosamine despite expression of core 2 N-acetylglucosaminyltransferase-L (C2GnT-L/GCNT1). This property allows androgen-refractory prostate cancer cells to evade galectin-1 (LGALS1)-induced apoptosis, but the mechanism is not known. We have recently reported that Golgi targeting of glycosyltransferases is mediated by golgins: giantin (GOLGB1) for C2GnT-M (GCNT3) and GM130 (GOLGA2)-GRASP65 (GORASP1) or GM130-giantin for core 1 synthase. Here, we show that for Golgi targeting, C2GnT-L also uses giantin exclusively whereas ST3Gal1 uses either giantin or GM130-GRASP65. In addition, the compact Golgi morphology is detected in both androgen-sensitive prostate cancer and normal prostate cells, but fragmented Golgi and mislocalization of C2GnT-L are found in androgen-refractory cells as well as primary prostate tumors (Gleason grade 2-4). Furthermore, failure of giantin monomers to be phosphorylated and dimerized prevents Golgi from forming compact morphology and C2GnT-L from targeting the Golgi. On the other hand, ST3Gal1 reaches the Golgi by an alternate site, GM130-GRASP65. Interestingly, inhibition or knockdown of non-muscle myosin IIA (MYH9) motor protein frees up Rab6a GTPase to promote phosphorylation of giantin by polo-like kinase 3 (PLK3), which is followed by dimerization of giantin assisted by protein disulfide isomerase A3 (PDIA3), and restoration of compact Golgi morphology and targeting of C2GnT-L. Finally, the Golgi relocation of C2GnT-L in androgen-refractory cells results in their increased susceptibility to galectin-1-induced apoptosis by replacing sialyl-T antigen with polylactosamine. IMPLICATIONS: This study demonstrates the importance of Golgi morphology and regulation of glycosylation and provides insight into how the Golgi influences cancer progression and metastasis.

Improvements in cerebral blood flow and recanalization rates with transcranial diagnostic ultrasound and intravenous microbubbles after acute cerebral emboli.

OBJECTIVES: Intravenous microbubbles (MBs) and transcutaneous ultrasound have been used to recanalize intra-arterial thrombi without the use of tissue plasminogen activator. In the setting of acute ischemic stroke, it was our objective to determine whether skull attenuation would limit the ability of ultrasound alone to induce the type and level of cavitation required to dissolve thrombi and improve cerebral blood flow (CBF) in acute ischemic stroke. MATERIALS AND METHODS: In 40 pigs, bilateral internal carotid artery occlusions were created with 4-hour-old thrombi. Pigs were then randomized to high-mechanical index (MI = 2.4) short-pulse (5 microseconds) transcranial ultrasound (TUS) alone or a systemic MB infusion (3% Definity) with customized cavitation detection and imaging system transmitting either high-MI (2.4) short pulses (5 microseconds) or intermediate-MI (1.7) long pulses (20 microseconds). Angiographic recanalization rates of both internal carotids were compared in 24 of the pigs (8 per group), and quantitative analysis of CBF with perfusion magnetic resonance imaging was measured before, immediately after, and at 24 hours using T2* intensity versus time curves in 16 pigs. RESULTS: Complete angiographic recanalization was achieved in 100% (8/8) of pigs treated with image-guided high-MI TUS and MBs, but in only 4 of 8 treated with high-MI TUS alone or 3 of 8 pigs treated with image-guided intermediate-MI TUS and MBs (both P < 0.05). Ipsilateral and contralateral CBF improved at 24 hours only after 2.4-MI 5-microsecond pulse treatments in the presence of MB (P < 0.005). There was no evidence of microvascular or macrovascular hemorrhage with any treatment. CONCLUSIONS: Guided high-MI impulses from an ultrasound imaging system produce sustained improvements in ipsilateral and contralateral CBF after acute cerebral emboli.

Myeloid-derived suppressor cells contribute to Staphylococcus aureus orthopedic biofilm infection.

Myeloid-derived suppressor cells (MDSCs) are a heterogeneous population of immature monocytes and granulocytes that are potent inhibitors of T cell activation. A role for MDSCs in bacterial infections has only recently emerged, and nothing is known about MDSC function in the context of Staphylococcus aureus infection. Because S. aureus biofilms are capable of subverting immune-mediated clearance, we examined whether MDSCs could play a role in this process. CD11b(+)Gr-1(+) MDSCs represented the main cellular infiltrate during S. aureus orthopedic biofilm infection, accounting for >75% of the CD45+ population. Biofilm-associated MDSCs inhibited T cell proliferation and cytokine production, which correlated with a paucity of T cell infiltrates at the infection site. Analysis of FACS-purified MDSCs recovered from S. aureus biofilms revealed increased arginase-1, inducible NO synthase, and IL-10 expression, key mediators of MDSC suppressive activity. Targeted depletion of MDSCs and neutrophils using the mAb 1A8 (anti-Ly6G) improved bacterial clearance by enhancing the intrinsic proinflammatory attributes of infiltrating monocytes and macrophages. Furthermore, the ability of monocytes/macrophages to promote biofilm clearance in the absence of MDSC action was revealed with RB6-C85 (anti-Gr-1 or anti-Ly6G/Ly6C) administration, which resulted in significantly increased S. aureus burdens both locally and in the periphery, because effector Ly 6C monocytes and, by extension, mature macrophages were also depleted. Collectively, these results demonstrate that MDSCs are key contributors to the chronicity of S. aureus biofilm infection, as their immunosuppressive function prevents monocyte/macrophage proinflammatory activity, which facilitates biofilm persistence.

Characterization of intracellular inclusions in the urothelium of mice exposed to inorganic arsenic.

Inorganic arsenic (iAs) is a known human carcinogen at high exposures, increasing the incidences of urinary bladder, skin, and lung cancers. In most mammalian species, ingested iAs is excreted mainly through urine primarily as dimethylarsinic acid (DMA(V)). In wild-type (WT) mice, iAs, DMA(V), and dimethylarsinous acid (DMA(III)) exposures induce formation of intramitochondrial urothelial inclusions. Arsenite (iAs(III)) also induced intranuclear inclusions in arsenic (+3 oxidation state) methyltransferase knockout (As3mt KO) mice. The arsenic-induced formation of inclusions in the mouse urothelium was dose and time dependent. The inclusions do not occur in iAs-treated rats and do not appear to be related to arsenic-induced urothelial cytotoxicity. Similar inclusions in exfoliated urothelial cells from humans exposed to iAs have been incorrectly identified as micronuclei. We have characterized the urothelial inclusions using transmission electron microscopy (TEM), DNA-specific 4',6-diamidino-2-phenylindole (DAPI), and non-DNA-specific Giemsa staining and determined the arsenical content. The mouse inclusions stained with Giemsa but not with the DAPI stain. Analysis of urothelial mitochondrial- and nuclear-enriched fractions isolated from WT (C57BL/6) and As3mt KO mice exposed to arsenate (iAs(V)) for 4 weeks showed higher levels of iAs(V) in the treated groups. iAs(III) was the major arsenical present in the enriched nuclear fraction from iAs(V)-treated As3mt KO mice. In conclusion, the urothelial cell inclusions induced by arsenicals appear to serve as a detoxifying sequestration mechanism similar to other metals, and they do not represent micronuclei.

Podoplanin and clusterin are reliable markers of nonneoplastic synovium at various sites.

INTRODUCTION: Clusterin (CLU) has been noted to mark synovium adjacent to tenosynovial tumors, and studies suggest that podoplanin (PP) is upregulated in inflammatory arthritis. Characterization of synovial staining with CLU and PP in various nonneoplastic disease states has not been described. METHODS: A microarray was created from paraffin-embedded human synovium, including 19 normal/noninflammatory (10 weight-bearing joints, 8 non-weight-bearing joints), 9 rheumatoid arthritis, 10 synovial cysts, and 3 osteoarthritis and stained with PP (D2-40) and CLU. Staining intensity was graded semiquantitatively (0-3+). RESULTS: PP and CLU stained synovium in 88% and 95% cases, respectively. PP and CLU showed moderate to strong (3+) staining in 26% and 19% of noninflammatory and 44% and 0% of inflammatory synovia, respectively (P < .01). CONCLUSIONS: PP and CLU are reliable markers of human synovium and can confirm its presence in limited specimens. Although CLU was more sensitive, PP may be more useful in the setting of chronic inflammation.

Urothelial cell intracytoplasmic inclusions after treatment of promyelocytic leukemia with arsenic trioxide.

Intramitochondrial inclusions containing arsenite that occur within urothelial cells have been previously described in mice exposed to high concentrations of arsenic but not in rats. In epidemiology studies, similar urothelial cell inclusions have also been observed in the urine of humans exposed to high concentrations of arsenic in the drinking water; however, these inclusions were mistakenly identified as micronuclei. To further examine the urothelial cell inclusions that occur in inorganic arsenic-exposed humans, we evaluated two patients with a history of acute promyelocytic leukemia treated for disease relapse with a combination of all-trans retinoic acid and arsenic trioxide. Posttreatment examination of the patients' urine cytology specimens by light and electron microscopy demonstrated cytoplasmic inclusions in exfoliated superficial urothelial cells similar to those seen in mice. The inclusions were present in decreasing quantities at 3 and 7 months after completion of treatment. No comparable inclusions were detected in exfoliated urothelial cells in urine from six individuals not treated with arsenic trioxide. Based on the results of the examination by light and electron microscopy, we have determined that urothelial cell inclusions in the urine of humans previously identified as micronuclei are instead intracytoplasmic inclusions similar to those found in arsenic-treated mice.

Sequential protein and peptide immunoaffinity capture for mass spectrometry-based quantification of total human ß-nerve growth factor.

Nerve growth factor (NGF) is a neurotrophin that is implicated in the modulation of pain perception. Tanezumab, a humanized monoclonal antibody (mAb) specific for NGF, is highly potent in sequestering NGF and has demonstrated efficacy for treatment of chronic pain in clinical trials. We describe a novel, sensitive immunoaffinity liquid chromatography-tandem mass spectrometry (LC-MS/MS) assay for quantitative determination of human serum NGF levels at baseline and after tanezumab treatment. The assay combines magnetic bead-based NGF immunoaffinity enrichment using a non-neutralizing polyclonal antibody followed by digestion and quantitation of a NGF-derived tryptic peptide via high-flow peptide immunoaffinity enrichment and nanoflow LC-MS/MS. Following validation, the assay was employed to measure total NGF concentrations in samples from clinical studies. The assay had a <10% interassay relative error and <15% interassay coefficient of variation across a range from 7.03 to 450 pg/mL human NGF. Generally, human basal serum NGF concentrations were between 20 and 30 pg/mL which, upon treatment with tanezumab, elevated in a dose-dependent manner into the high pg/mL to low ng/mL range. This is the first report of clinical trial implementation of a MS-based assay that uses sequential protein and peptide immunoaffinity capture for protein target quantitation. The use of robotic sample preparation and a robust chromatography configuration enabled this technology to advance into the routine clinical analysis and now provides a bioanalytical platform for the development of similar assays for other protein targets.

Diuron-induced rat bladder epithelial cytotoxicity.

Diuron, a substituted urea herbicide, is carcinogenic to the rat urinary bladder at high dietary levels (2500 ppm). To further elucidate the mode of action, this study aimed to determine the time course and sequence of bladder cytotoxic and proliferative changes induced by diuron treatment of male Wistar rats. Rats were randomized into two groups (control and 2500 ppm diuron) and treated for 28 days. Ten rats from each group were terminated on each of study days 1, 3, 7, or 28. Scanning electron micro scopy (SEM) showed urothelial cell swelling beginning on day 1, and by day 28, showed extensive necrosis, exfoliation and piling up of cells suggestive of hyperplasia. No difference in the bromo deoxyuridine labeling index was detected. In a second experiment, rats were randomized into control and diuron-treated groups and treated for 7 days or 8 weeks. After 7 days, transmission electron microscopy showed cell degenerative changes and distention of the cytoplasm, organelles, and nuclei characteristic of cytolysis. This resulted in protrusion of the superficial cells into the lumen, corresponding to the cell swelling observed previously by SEM. After 8 weeks, bladders in the diuron-treated group showed an increased incidence of simple hyperplasia by light microscopy (6/10, p < 0.05) compared with controls (0/10) and a significantly different SEM classification. In summary, our results support the hypothesis that urothelial cytotoxicity followed by regenerative cell proliferation are the sequential key events that occur with high-dose diuron exposure in rats.

Pericytoma with t(7;12) and ACTB-GLI1 fusion arising in bone.

Cytogenetic analysis of a primary bone neoplasm with pericytic features in a 67-year-old man revealed a t(7;12)(p22;q13) among other karyotypic abnormalities. Subsequent molecular studies confirmed the presence of an associated ACTB-GLI1 fusion transcript. An identical 7;12 translocation is known to characterize a discrete group of soft tissue tumors belonging to the myopericytic category termed pericytoma with t(7;12). To the best of our knowledge, this is the first case of pericytoma with t(7;12) arising in bone. Cytogenetic and molecular analyses were useful, if not essential, in classifying this rare diagnostic entity.

Rab11 is a useful tool for the diagnosis of microvillous inclusion disease.

Microvillous inclusion disease (MVID) is a congenital condition presenting with intractable diarrhea. Biopsies demonstrate abnormal apical PAS and CD10 staining in surface enterocytes correlating with the presence of characteristic cytoplasmic inclusions. MVID has been linked to mutations in myosin Vb, important in apical membrane recycling. Rab11 associates with myosin Vb in vesicle membranes and is also integral in recycling plasma membrane components. The authors performed Rab11 immunostaining on biopsies from 7 MVID cases, 10 normal small intestines, and 10 with chronic enteritis. In MVID cases, Rab11 showed diffuse apical cytoplasmic staining of surface enterocytes in a pattern similar to PAS and CD10, which was absent in all the 20 control cases. Ultrastructural examination confirmed localization to the external surface of MVID cytoplasmic inclusions. Rab11 staining may be a useful adjunct in MVID diagnosis and the results support that myosin Vb dysfunction is important in the pathogenesis of MVID.

Simultaneous and high-throughput quantitation of urinary tetranor PGDM and tetranor PGEM by online SPE-LC-MS/MS as inflammatory biomarkers.

Quantitation of urinary tetranor PGDM or tetranor PGEM (tPGDM and tPGEM) in the past was performed separately using off-line SPE LC-MS/MS methods. The manual SPE procedure is generally time-consuming and cost-ineffective. In addition, simultaneous quantitation of tPGDM and tPGEM is favorable yet very challenging because of the similar chemical structures and identical MRM transitions. This work describes the development and validation of a high-throughput online SPE-LC-MS/MS method, allowing simultaneous and high-throughput measurement of tPGDM and tPGEM in human urine. The reportable range of the assay was 0.2-40 ng/ml for tPGDM and 0.5-100 ng/ml for tPGEM. Intra- and inter-assay precision and accuracy determined using quality control samples were all within acceptable ranges (% CV and % Bias < 15%). Tetranor PGDM was stable under all tested conditions while tPGEM was stable at 4 °C and after three F/T cycles but not stable at room temperature for 24 h (recovery below 80%). The assay was applied to measure urinary tPGDM and tPGEM among healthy volunteers, smokers and COPD patients. Significantly higher urinary levels of both tPGDM and tPGEM were observed in COPD patients than those of non-smoking healthy volunteers. These results demonstrated that the high-throughput online SPE-LC-MS/MS assay provides sensitive, reproducible and accurate measurement of urinary tPGDM and tPGEM as biomarkers for assessing inflammatory diseases such as COPD.