Endotoxin in Aerosol Particles from Metalworking Fluids Measured with a Sioutas Cascade Impactor.

OBJECTIVES: The aim of this study was to characterize personal occupational exposure to endotoxin in size-separated airborne particles of MWF aerosol, using a Sioutas cascade impactor (SCI). METHODS: Exposure to inhalable fractions of MWF aerosol and endotoxin was measured by personal sampling of 52 individuals over an 8-h work shift using a PAS-6 sampler in parallel with a SCI (<0.25, 0.25-0.5, 0.5-1.0, 1.0-2.5, and 2.5-10 µm). Aerosol mass concentration was measured for each worker with a real-time instrument (DataRAM) during a full shift. Samples of MWF were collected from the machines and central tanks during the work shift. RESULTS: A total of 117 measurements of inhalable MWF aerosols were made among 52 workers. The geometric mean of inhalable MWF aerosol was 0.16 mg m-3 air. The geometric mean of endotoxin concentration on the inhalable sampler was 0.15 EU m-3. Airborne endotoxin was found on all size fractions from the impactor, with the major part seen in the fraction (2.5-10 µm). There was a correlation between the inhalable fraction of endotoxin measured by the PAS-6 sampler and on the SCI sampler (2.5-10 µm), estimated to be 0.51 for all samples (P < 0.0001). The concentration of endotoxin varied between the MWFs, as did the proportion of Gram-negative bacteria among the culturable bacteria (>80% in one MWF and <1.5% in the other three). CONCLUSIONS: The personal exposure to inhalable fractions of endotoxin contained in the MWF aerosol were low, where most of the endotoxin were found in fraction (2.5-10 µm), measured by SCI. There are differences between factories and MWF systems regarding the distribution of endotoxin and so results from one context should not be generalized to other plants and systems. Compressed air was used for less than 10 min shift-1. The mixed-effect model showed that working with open machines and grinding as cutting task were important determinants of exposure to inhalable aerosol. It is important to keep occupational exposure to aerosols low with the help of good ventilation systems, enclosed machines, and organization of work.

Dust Exposures in Swedish Soft Tissue Paper Mills.

OBJECTIVES: Paper dust has previously been linked to adverse health effects. However, a comprehensive dataset of paper dust exposures does not appear to have been published previously. Our study was intended to address this need by describing a large dataset of measurements made in Swedish soft tissue paper mills. METHODS: We compiled personal and area total dust exposure measurements collected from a range of operations by our research staff at four soft tissue paper mills in Sweden. We also compiled measurements made by the occupational health staff at each mill and the Swedish Work Environment Authority. We analyzed these measurements to describe patterns and trends in exposures and used mixed-effects regression models to identify measurement characteristics that predicted exposure levels. RESULTS: We compiled 1578 measurements from 1971 to 2009, of which 1026 (65%) were personal samples. Statistically significant differences were found between measurements made by research, mill, and Swedish Work Environment Authority staff, as well as between personal and area measurements. The measurement data suggest that, beginning in the 1980s, exposures declined at three of the four mills, but that overexposures were still common at the end of the period. Papermaking and converting operations had the highest observed dust exposures. One mill had significantly lower exposures than the others. Type of measurement (personal versus area) and source of measurement (research staff, company, or regulatory agency) were not significant predictors of measured total dust exposure after controlling for mill, operation, and time. CONCLUSIONS: Our analysis of measured paper dust exposures may be useful for historical and contemporary exposure assessment in our own and other epidemiological studies. We have identified specific characteristics (i.e. papermaking operations and mill) and time trends that are important data features to consider, and documented continuing overexposure situations. Our results highlight the ongoing need for application of exposure controls to reduce paper dust exposures in the soft tissue paper industry.

Intra-individual variation of particles in exhaled air and of the contents of Surfactant protein A and albumin.

INTRODUCTION: Particles in exhaled air (PEx) provide samples of respiratory tract lining fluid from small airways containing, for example, Surfactant protein A (SP-A) and albumin, potential biomarkers of small airway disease. We hypothesized that there are differences between morning, noon, and afternoon measurements and that the variability of repeated measurements is larger between days than within days. METHODS: PEx was obtained in sixteen healthy non-smoking adults on 11 occasions, within one day and between days. SP-A and albumin were quantified by ELISA. The coefficient of repeatability (CR), intraclass correlation coefficient (ICC), and coefficient of variation (CV) were used to assess the variation of repeated measurements. RESULTS: SP-A and albumin increased significantly from morning towards the noon and afternoon by 13% and 25% on average, respectively, whereas PEx number concentration and particle mean mass did not differ significantly between the morning, noon and afternoon. Between-day CRs were not larger than within-day CRs. CONCLUSIONS: Time of the day influences the contents of SP-A and albumin in exhaled particles. The variation of repeated measurements was rather high but was not influenced by the time intervals between measurements.

A semi-quantitative job exposure matrix for dust exposures in Swedish soft tissue paper mills.

BACKGROUND: Total paper dust exposure has been associated with respiratory problems among workers in the soft tissue paper industry. However, a comprehensive job exposure matrix (JEM) has not been developed for application to this industry. Our study was intended to address this need and to support further studies of mortality and morbidity in a cohort of Swedish workers from this industry. METHODS: We evaluated four participating soft tissue paper mills in Sweden. We combined information on process and equipment status from the mills with knowledge of the mills obtained through research efforts and paper dust measurements made at all four mills to develop a semi-quantitative JEM with seven dust exposure levels. The JEM was targeted at workers enrolled into a soft tissue paper mill cohort and working any time between 1960 and 2009. RESULTS: The JEM includes a total of 14 421 cells, with each cell corresponding to the exposure for a job title, department, or work location for a one-year period. Exposure levels in the JEM were estimated to decline at three of the four mills from 1971 to 2009, but overexposures (ie, exceedances of the relevant occupational exposure limits) remained common at the end of the period. CONCLUSIONS: The JEM results highlight the need for ongoing exposure control efforts in the soft tissue paper industry, and will inform ongoing epidemiological studies of the health effects of exposure to paper dust in Sweden. It is freely available for use by other researchers.

Development of a Job Exposure Matrix for Noise in the Swedish Soft Tissue Paper Industry.

Objectives: Noise exposure is a common occupational hazard, but has not been sufficiently characterized in paper mills. We developed a job-exposure matrix (JEM) for noise exposure for use in estimating exposures among Swedish soft tissue paper mill workers. Methods: We used a combination of area and personal dosimetry noise exposure measurements made at four soft tissue paper mills by industry and research staff between 1977 and 2013 to estimate noise exposures by department, location, and job title. We then utilized these estimates, in conjunction with information on process and facility changes and use of hearing protection collected via focus groups, to create a seven-category, semi-quantitative JEM for all departments, locations, and job titles spanning the years 1940-2010. Results: The results of the 1157 area and personal dosimetry noise measurements indicated that noise levels have generally declined in Swedish paper mills over time, though these changes have been neither uniform nor monotonic within or across the four mills. Focus group results indicated that use of hearing protection has generally increased over time. The noise JEM totals 1917 cells, with each cell representing a unique combination of operation, job title, and single year. We estimated that ~50% of workers at the four mills assessed were exposed at or above the Swedish 8-h average noise exposure limit of an 85 dBA at the conclusion of the study period in 2010. Conclusions: Our results highlight the continuing need for hearing loss prevention and noise control efforts at these and similar mills, and the completed JEM now represents a tool for use in epidemiological studies of noise-related health outcomes.

Comparison of Multiple Measures of Noise Exposure in Paper Mills.

BACKGROUND: Noise exposures are associated with a host of adverse health effects, yet these exposures remain inadequately characterized in many industrial operations, including paper mills. We assessed noise at four paper mills using three measures: (i) personal noise dosimetry, (ii) area noise measurements, and (iii) questionnaire items addressing several different aspects of perceived noise exposure. METHODS: We assessed exposures to noise characterized using the three measures and compared the relationships between them. We also estimated the validity of each of the three measures using a novel application of the Method of Triads, which does not appear to have been used previously in the occupational health literature. RESULTS: We collected 209 valid dosimetry measurements and collected perceived noise exposure survey items from 170 workers, along with 100 area measurements. We identified exposures in excess of 85 dBA at all mills. The dosimetry and area noise measurements assigned to individual subjects generally showed good agreement, but for some operations within mill, large differences between the two measures were observed, and a substantial fraction of paired measures differed by >5 dB. Perceived noise exposures varied greatly between the mills, particularly for an item related to difficulty speaking in noise. One perceived noise exposure item related to difficulty hearing due to noise showed strong and significant correlations with both dosimetry and area measurements. The Method of Triads analysis showed that dosimetry measures had the highest estimated validity coefficient (0.70), and that the best performing perceived exposure measure had validity that exceeded that of area measurements (0.48 versus 0.40, respectively). CONCLUSIONS: Workers in Swedish pulp mills have the potential for exposures to high levels of noise. Our results suggest that, while dosimetry remains the preferred approach to exposure assessment, perceived noise exposures can be used to evaluate potential exposures to noise in epidemiological studies.

Food intake, tumor growth, and weight loss in EP2 receptor subtype knockout mice bearing PGE2-producing tumors.

Previous studies in our laboratory have demonstrated that prostaglandin (PG) E2 is involved in anorexia/cachexia development in MCG 101 tumor-bearing mice. In the present study, we investigate the role of PGE receptor subtype EP2 in the development of anorexia after MCG 101 implantation in wild-type (EP2 (+/+)) or EP2-receptor knockout (EP2(-/-)) mice. Our results showed that host absence of EP2 receptors attenuated tumor growth and development of anorexia in tumor-bearing EP2 knockout mice compared to tumor-bearing wild-type animals. Microarray profiling of the hypothalamus revealed a relative twofold change in expression of around 35 genes including mRNA transcripts coding for Phospholipase A2 and Prostaglandin D2 synthase (Ptgds) in EP2 receptor knockout mice compared to wild-type mice. Prostaglandin D2 synthase levels were increased significantly in EP2 receptor knockouts, suggesting that improved food intake may depend on altered balance of prostaglandin production in hypothalamus since PGE2 and PGD2 display opposing effects in feeding control.

Preoperative low dose NSAID treatment influences the genes for stemness, growth, invasion and metastasis in colorectal cancer.

Preclinical data, and an increasing list of clinical investigations, show anti-inflammatory agents to favourably influence the biology of colorectal tumor. We have earlier reported on re-expression of activated immune cells after three days preoperative treatment of patients with colorectal carcinoma, randomized to receive oral NSAID (indomethacin or celebrex). Antisecretory prophylaxis (esomeprasol) was provided to all patients and served as sham treatment. Concomittant to MHC locus activation, Prominin1/CD133, a marker associated with stemness and poor prognosis in several solid tumors, was downregulated. The aim of the present study was to evaluate expression of additional regulators belonging to the stem cell niche, OCT4, SOX2 and BMP7, as well as some microRNAs, reported to act as tumor suppressors or oncomiRs. Peroperative tumor biopsies were analyzed by microarrays, quantitative real-time PCR and immunohistochemistry (IHC). The stem cell master regulator SOX2 was increased by NSAIDs (p<0.01), as well as the tumor suppressor miR-630 (p<0.01), while BMP7, a marker for poor prognosis in CRC, was downregulated by NSAID (indomethacin, p<0.02). The upregulation of SOX2, but not of its heterodimer binding partner OCT4, could imply a negative feed-back loop, with a switch­off for stemness preservation of tumor cells. This is supported by the overall evaluation of gene expression profiles with subsequent events, indicating less aggressive tumors following NSAID treatment.

Rear seat child safety in near-side impacts: a modeling study of common sitting positions.

OBJECTIVE: The purpose of this study was to evaluate and propose improvements to the injury mitigation systems, in near-side impacts, for 6 common sitting positions of young adolescents using a previously validated model. METHODS: The evaluation was made by using a model of a complete passenger car, including head and thorax-pelvis air bags, which was impacted laterally by a barrier in 2 load cases. The SID-IIs finite element model was used for the evaluations and was seated in 6 different positions in the rear outboard seat: the nominal anthropomorphic test device (ATD) position, 1 inboard position, 3 outboard positions, and 1 braking (forward) position. These positions have previously been identified as common sitting positions in awake and asleep children. The studied dependent variables were head injury criterion (HIC) 36, resultant head linear acceleration, resultant head rotational acceleration, chest viscous criterion, and chest deflection. RESULTS: The lowest head injury measures were seen in the braking positions and in the nominal ATD position, and the highest were seen in the inboard and outboard positions. The lowest chest injury measures were recorded in the inboard and nominal ATD positions, and the highest were recorded in the outboard and braking positions. The occupant in the outboard positions interfered with the air bags during their deployment. The occupant in inboard and braking positions tended to push the curtain air bag over the windowsill. CONCLUSIONS: Studies that investigate the injury mitigation effects in common sitting positions, beyond the nominal ATD position, are essential to highlight means to provide improved and robust safety for child occupants. This study was based on the SID-IIs 5th percentile female, which has very similar anthropometry to a 50th percentile 12-year-old. Therefore, the conclusions of this study are applicable to many 11-year-olds up to young adolescents, as well as to small females. The outboard and inboard positions of this study resulted in the highest head injury measures. Although all of the injury measures were only slightly higher than the nominal position, the trends suggest that, in near-side impacts, these positions should be discouraged. The extensively outboard positions resulted in unfavorable air bag positioning during deployment. The inboard position resulted in head strikes further forward of the nominal one; the curtain air bags need inflated cells at all locations of head strike.

Evaluation of the head kinematics of the Q3 model and a modified Q3 model by means of crash reconstruction.

OBJECTIVE: One objective of this study is to evaluate the head kinematics of the Q3 model. Another objective is to evaluate the effect on head kinematics of increased thoracic spine flexibility; more humanlike mass distribution; and more humanlike body geometry in the Q3 model. The evaluations were based on the head kinematics of children deduced from real crashes and on new data of mass distribution and updated body dimensions for 3-year-olds. METHODS: The head kinematics of the Q3 model was evaluated by comparing the Q3 model's head displacement response with the deduced response of 3-year-old children in real crashes. To do so, data from crashes were collected. The data were used to develop the mathematical vehicle and restraint system models (MADYMO, TASS, the Netherlands). Three crashes involving 3-year-old children in frontal impacts were reconstructed. The models were run 35 times each (one model per crash), each time with a different setting to each of the variables for which the exact value was not known. Examples of those variables include crash pulse, initial dummy position, and initial seat belt position. Two versions of the Q3 model were used: one that correlated with the Q3 ATD and one that was modified regarding anthropometry and thoracic flexibility. The basis for the updated anthropometry was new data regarding characteristic dimensions and mass distribution collected at a Swedish hospital. RESULTS: In the anthropometry study, 26 children were measured. The main differences between the average of the measured children and the Q3 model were found in the mass distribution of the torso and thighs: the Q3's pelvis was too heavy and the thorax, abdomen/lumbar spine, and thighs were too light. Another difference was identified in the buttock-knee length. Two of the 3 reconstructed crashes had confirmed head impacts. The Q3 model responded with head kinematics that reflected the deduced courses of events for the real children in one of 3 crashes (the one without head impact). The modified Q3 model reflected the real children in 2 of 3 crashes. CONCLUSIONS: In high-severity, straight frontal crashes, the Q3 model predicted non-head impact adequately. However, in oblique frontal crashes, the Q3 model did not sufficiently predict the head impacts. The modified Q3 model predicted the head impacts better than the Q3 model did. Greater flexibility of the thorax and redistributed mass made a positive difference regarding the head kinematics.

Downregulation of Prominin 1/CD133 expression in colorectal cancer by NSAIDs following short-term preoperative treatment.

Expression of Prominin 1/CD133 is associated with poor prognosis and chemoresistance in several types of solid tumors. The aim of the present study was, therefore, to evaluate Prominin 1/CD133 expression in colorectal carcinoma after short-term preoperative treatment by non-steroidal anti-inflammatory drugs (NSAIDs). Patients aimed at curative operation for colon cancer were randomized to receive NSAIDs (indomethacin 50 mg x2 or celecoxib 100 mg x2) three days preoperatively. Antisecretory prophylaxis (esomeprasol 40 mg x1) was provided to all patients and served as sham intake. CD133 expression in tumor tissue was also assessed in tumors from Dukes' B patients selected for either long or short postoperative survival. No patients received perioperative chemoradiotherapy. Tumor tissue was collected at surgery for quantification of mRNAs (Prom1 and Wnt4) by qPCR. Immunohistochemistry stained for CD133, Ep-CAM, CD34 and CD45. PGE2 content in tumor tissue was determined. Transcript of CD133 in tumor tissue was lower in patients treated with NSAIDs (0.28 ± 0.07 vs. 0.51 ± 0.08; p<0.03) as well as some other stem cell-related transcripts. In treated patients 36% of all tumors stained positive for CD133 compared to 71% in sham-treated control patients (p<0.05). Short survivors with Dukes' B tumors displayed 78% CD133 expression as compared to 33% of tumors in long-term survivors (p<0.002). Tumor tissue PGE(2) content was negatively related to patient survival. Our results show that short-term preoperative NSAID treatment downregulates colon cancer tissue expression of Prominin 1/CD133, a stem cell marker indicative of survival prognosis as confirmed.

Parameter study for child injury mitigation in near-side impacts through FE simulations.

OBJECTIVE: The objective of this study is to investigate the effects of crash-related car parameters on head and chest injury measures for 3- and 12-year-old children in near-side impacts. METHODS: The evaluation was made using a model of a complete passenger car that was impacted laterally by a barrier. The car model was validated in 2 crash conditions: the Insurance Institute for Highway Safety (IIHS) and the US New Car Assessment Program (NCAP) side impact tests. The Small Side Impact Dummy (SID-IIs) and the human body model 3 (HBM3) (Total HUman Model for Safety [THUMS] 3-year-old) finite element models were used for the parametric investigation (HBM3 on a booster). The car parameters were as follows: vehicle mass, side impact structure stiffness, a head air bag, a thorax-pelvis air bag, and a seat belt with pretensioner. The studied dependent variables were as follows: resultant head linear acceleration, resultant head rotational acceleration, chest viscous criterion, rib deflection, and relative velocity at head impact. The chest measurements were only considered for the SID-IIs. RESULTS: The head air bag had the greatest effect on the head measurements for both of the occupant models. On average, it reduced the peak head linear acceleration by 54 g for the HBM3 and 78 g for the SID-IIs. The seat belt had the second greatest effect on the head measurements; the peak head linear accelerations were reduced on average by 39 g (HBM3) and 44 g (SID-IIs). The high stiffness side structure increased the SID-IIs' head acceleration, whereas it had marginal effect on the HBM3. The vehicle mass had a marginal effect on SID-IIs' head accelerations, whereas the lower vehicle mass caused 18 g higher head acceleration for HBM3 and the greatest rotational acceleration. The thorax-pelvis air bag, vehicle mass, and seat belt pretensioner affected the chest measurements the most. The presence of a thorax-pelvis air bag, high vehicle mass, and a seat belt pretensioner all reduced the chest viscous criterion (VC) and peak rib deflection in the SID-IIs. CONCLUSIONS: The head and thorax-pelvis air bags have the potential to reduce injury measurements for both the SID-IIs and the HBM3, provided that the air bag properties are designed to consider these occupant sizes also. The seat belt pretensioner is also effective, provided that the lateral translation of the torso is managed by other features. The importance of lateral movement management is greater the smaller the occupant is. Light vehicles require interior restraint systems of higher performance than heavy vehicles do to achieve the same level of injury measures for a given side structure.

COX-2 gene expression in colon cancer tissue related to regulating factors and promoter methylation status.

BACKGROUND: Increased cyclooxygenase activity promotes progression of colorectal cancer, but the mechanisms behind COX-2 induction remain elusive. This study was therefore aimed to define external cell signaling and transcription factors relating to high COX-2 expression in colon cancer tissue. METHOD: Tumor and normal colon tissue were collected at primary curative operation in 48 unselected patients. COX-2 expression in tumor and normal colon tissue was quantified including microarray analyses on tumor mRNA accounting for high and low tumor COX-2 expression. Cross hybridization was performed between tumor and normal colon tissue. Methylation status of up-stream COX-2 promoter region was evaluated. RESULTS: Tumors with high COX-2 expression displayed large differences in gene expression compared to normal colon. Numerous genes with altered expression appeared in tumors of high COX-2 expression compared to tumors of low COX-2. COX-2 expression in normal colon was increased in patients with tumors of high COX-2 compared to normal colon from patients with tumors of low COX-2. IL1ß, IL6 and iNOS transcripts were up-regulated among external cell signaling factors; nine transcription factors (ATF3, C/EBP, c-Fos, Fos-B, JDP2, JunB, c-Maf, NF-κB, TCF4) showed increased expression and 5 (AP-2, CBP, Elk-1, p53, PEA3) were decreased in tumors with high COX-2. The promoter region of COX-2 gene did not show consistent methylation in tumor or normal colon tissue. CONCLUSIONS: Transcription and external cell signaling factors are altered as covariates to COX-2 expression in colon cancer tissue, but DNA methylation of the COX-2 promoter region was not a significant factor behind COX-2 expression in tumor and normal colon tissue.

Effect of Booster Seat Design on Children's Choice of Seating Positions During Naturalistic Riding.

The purpose of this naturalistic study was to investigate the effect of booster seat design on the choice of children's seating positions during naturalistic riding. Data was collected through observations of children during in-vehicle riding by means of a film camera. The children were positioned in high back boosters in the rear seat while a parent drove the car. The study included two different booster designs: one with large head and torso side supports, and one with small head side supports and no torso side supports. Six children between three and six years of age participated in the study. Each child was observed in both boosters. The duration of the seating positions that each child assumed was quantified. The design with large side head supports resulted more often in seating positions without head and shoulder contact with the booster's back. There was shoulder-to-booster back contact during an average of 45% of riding time in the seat with the large head side supports compared to 75% in the seat with the small head supports. The children in the study were seated with the head in front of the front edge of the head side supports more than half the time, in both boosters. Laterally, the children were almost constantly positioned between the side supports of the booster in both seats. The observed seating positions probably reduce the desired protective effect by the side supports in side impact, and may increase the probability of head impact with the vehicle interior in frontal impact.

Mechanisms behind COX-1 and COX-2 inhibition of tumor growth in vivo.

Non-steroidal anti-inflammatory drugs (NSAIDs) attenuate tumor net growth in clinical and experimental cancer. Evaluations in cell culture experiments have implied involvement of growth factor and G-protein related signaling pathways to explain decreased proliferation, angiogenesis, increased cell adhesion and apoptosis. Sparse information is however available from studies on growing tumors in vivo. The aim of the present study was to map alterations in selected signal proteins in relation to heterogeneous tissue expression of COX-2 in tumors during COX inhibition. MCG 101 cells were exposed to indomethacin treatment both in vivo and in vitro to reduce PGE2 production. Tumor tissue specimens were taken for immunohistochemical analyses and qPCR determinations. Protein markers were selected to reflect cell proliferation and cell cycling, angiogenesis and metastasis in relationship to COX-2 staining in tumor tissue. Indomethacin did not change overall COX-2 staining in tumor tissue, but altered its distribution towards increased staining in cell nuclei/nucleoli and decreased COX-2 staining heterogeneity in tumor tissue. P53 staining was decreased, while PCNA and TGFß3 staining were increased by indomethacin in tumor areas with high presence of COX-2, which correlated to staining of BAX, TUNEL, Bcl-2, c-jun, p21, p27, p53 and NM23. Net tumor growth was predicted by EGF-R, p21 and p27 proteins in tumor tissue during indomethacin treatment (multivariate analysis). RNA transcript analyses showed decreased EGF-R and KRas expression in vivo, following indomethacin treatment, which also included KRas, PI3K, JAK1, STAT3 and c-jun, mRNAs in cultured tumor cells. In conclusion, our results extend earlier studies on cell culture experiments and demonstrate that EGF-R and downstream KRas pathways communicate effects of increased prostaglandin activity in tumor tissue in vivo.

Genes with relevance for early to late progression of colon carcinoma based on combined genomic and transcriptomic information from the same patients.

BACKGROUND: Genetic and epigenetic alterations in colorectal cancer are numerous. However, it is difficult to judge whether such changes are primary or secondary to the appearance and progression of tumors. Therefore, the aim of the present study was to identify altered DNA regions with significant covariation to transcription alterations along colon cancer progression. METHODS: Tumor and normal colon tissue were obtained at primary operations from 24 patients selected by chance. DNA, RNA and microRNAs were extracted from the same biopsy material in all individuals and analyzed by oligo-nucleotide array-based comparative genomic hybridization (CGH), mRNA- and microRNA oligo-arrays. Statistical analyses were performed to assess statistical interactions (correlations, co-variations) between DNA copy number changes and significant alterations in gene and microRNA expression using appropriate parametric and non-parametric statistics. RESULTS: Main DNA alterations were located on chromosome 7, 8, 13 and 20. Tumor DNA copy number gain increased with tumor progression, significantly related to increased gene expression. Copy number loss was not observed in Dukes A tumors. There was no significant relationship between expressed genes and tumor progression across Dukes A-D tumors; and no relationship between tumor stage and the number of microRNAs with significantly altered expression. Interaction analyses identified overall 41 genes, which discriminated early Dukes A plus B tumors from late Dukes C plus D tumor; 28 of these genes remained with correlations between genomic and transcriptomic alterations in Dukes C plus D tumors and 17 in Dukes D. One microRNA (microR-663) showed interactions with DNA alterations in all Dukes A-D tumors. CONCLUSIONS: Our modeling confirms that colon cancer progression is related to genomic instability and altered gene expression. However, early invasive tumor growth seemed rather related to transcriptomic alterations, where changes in microRNA may be an early phenomenon, and less to DNA copy number changes.

Receptor and enzyme expression for prostanoid metabolism in colorectal cancer related to tumor tissue PGE2.

Prostaglandins support progression of colorectal cancer by several mechanisms. This conclusion is based on epidemiological and drug intervention long-term studies or retrieved from animal and cell culture experiments. The aim of the present study was to map receptor and enzyme expression for prostanoid metabolism in the presence of high or low PGE2 content within colon cancer tissue at primary tumor operation and after short-term preoperative provision of non-steroidal anti-inflammatory drug (NSAID). Twenty-three unselected patients with colon cancer were randomly selected to receive indomethacin (NSAID) or sham treatment for 3 days before surgery. Normal colon and tumor tissue were collected at operation for RNA extraction. Tissue PGE2 levels were measured by radioimmunoassay. Gene expression was quantified by microarray and real-time PCR. COX-1 expression increased proportionally to COX-2 expression in colon cancer tissue from untreated patients. Indomethacin reduced PGE2 content in normal and tumor tissue with subsequently decreased IP, HPGD and PPARgamma receptor expression in both tumor and normal colon tissue, while subtype EP1-4 receptors were not significantly influenced by indomethacin treatment. MPGES-1 expression was not related to overall PGE2 content in tumor and colon tissue, but decreased significantly in normal tissue during indomethacin exposure. Reduction of tumor tissue PGE2 was related to significant alteration in expression of several hundred genes indicating decreased cell cycling and increased apoptosis during indomethacin treatment, probably related to upregulation of acute phase reactants in tumor tissue. Increased prostanoid activity in colon cancer tissue is related to cross-talk between tumor and stroma cells.

Preoperative treatment with a non-steroidal anti-inflammatory drug (NSAID) increases tumor tissue infiltration of seemingly activated immune cells in colorectal cancer.

This study evaluates HLA gene expression and tumor infiltration by B-cells, macrophages, dendritic cells, T-helper and cytotoxic T-lymphocytes in response to short-term preoperative treatment with cyclooxygenase inhibitors. Patients with colorectal carcinoma were randomized to receive oral NSAID (indomethacin or celebrex) for three days preoperatively; controls received esomeprazol. Peroperative tumor biopsies and normal colon tissue were analyzed by microarray, quantitative PCR and immunohistochemistry. Efficacy of short-term systemic NSAID treatment was confirmed by measurement of PGE2 production in blood monocytes from healthy volunteers. NSAID treatment upregulated genes at the MHC locus on chromosome 6p21 in tumor tissue, but not in normal colon tissue, from the same patient. 23 of the 100 most upregulated genes belonged to MHC class II. HLA-DM, -DO (peptide loading), HLA-DP, -DQ, -DR (antigen presentation), granzyme B, H, perforin and FCGR3A (CD16) (cytotoxicity) displayed increased expression, as did CD8, a marker of cytotoxic T-lymphocytes, while HLA-A and -C expression were not increased by NSAID treatment. MHC II protein (HLA-DP, -DQ, -DR) levels and infiltration by CD4+ T-helper cells of tumor stroma increased upon NSAID treatment, while CD8+ cytotoxic T-lymphocytes increased in both tumor stroma and epithelium. Molecules associated with immunosuppressive T regulatory cells (FOXP3, IL-10) were significantly decreased in indomethacin-exposed tumors. Standard oral administration of NSAID three days preoperatively was enough to increase tumor infiltration by seemingly activated immune cells. These findings agree with previous information that high prostanoid activities in colorectal cancer increase the risk for reduced disease-specific survival following tumor resection.

Prostanoid receptor expression in colorectal cancer related to tumor stage, differentiation and progression.

INTRODUCTION: Alterations in eicosanoid metabolism is well established in a variety of malignant tumors, particularly colorectal carcinoma. Recent studies in our laboratory have emphasized a role for EP subtype receptors in progression of colorectal cancer and disease specific mortality. Therefore, the aim of the present study was to extend our knowledge to include additional receptor expression (DP1, DP2, FP, IP, TP) for prostanoids (PGD2, TXA2, PGF2alpha, PGI2) in relationship to tumor stage, differentiation and progression of colorectal cancer. MATERIAL AND METHODS: Total RNA from 62 tumors and adjacent normal colon tissue (n = 48) was extracted. Quantification of receptor expression was performed by realtime PCR and related to the expression of an appropriate housekeeping gene (GAPDH). Tumors were assessed according to Dukes A-D (stage I-IV). RESULTS: DP1, DP2, FP and IP receptor subtypes displayed significantly reduced overall expression in tumor tissue compared to normal colon tissue, while the TP receptor subtype showed significantly higher expression in tumor tissue. Overall expression of the prostanoid receptors in tumor tissue was not related to clinical indexes as tumor stage and tumor cell differentiation evaluated by multivariate analyses. Cultured colorectal cancer cell lines with low (HT-29) and high (HCA-7) intrinsic PGE2 production at confluent state did not express DP1 and IP receptor subtypes, but displayed low expression of DP2, FP and TP receptor subtypes. CONCLUSION: The results in the present study indicate imbalanced expression of prostanoid receptors in colorectal cancer compared to normal colon tissue without clear cut relationship to disease progression. Therefore, future studies should be performed on defined cells within the tumor tissue compartment determining whether any prostanoid receptor(s) is useful as a molecular target in treatment or prevention of colorectal cancer.

EP1-4 subtype, COX and PPAR gamma receptor expression in colorectal cancer in prediction of disease-specific mortality.

The importance of prostaglandins in tumor growth and progression is well recognized, including antineoplastic activities by cyclooxygenase (COX) inhibitors. Variation in treatment response to COX inhibition has questioned differences in expression of cell surface and nuclear membrane receptors among tumors with different disease progression. The purpose of this study was to evaluate whether EP(1-4) subtype, PPAR gamma receptor and COX-1/COX-2 expression in colorectal cancer are related to tumor-specific mortality. Reverse transcription-polymerase chain reaction and immunohistochemistry were used to demonstrate expression and protein appearance in tumor tissue compared with normal colon tissue. EP(1) and EP(2) subtype receptor protein was highly present in tumor cells, EP(3) occurred occasionally and EP(4) was not visible. PPAR gamma, EP(2) and EP(4) mRNA were significantly higher in normal colon tissue compared with tumor tissue, without any distinct relationship to Dukes A-D tumor stage. Multivariate analyses indicated that increased tumor tissue EP(2) and COX-2 expression predicted poor survival (p<0.001). COX-1 expression was significantly higher than COX-2 expression in normal colon tissue. Average COX-2 mRNA was not increased in tumor tissue compared with normal colon. However, most tumor cells stained positive for COX-2 protein, which was low or undetectable in normal mucosa cells. COX-1 protein was preferentially visible in stroma. EP(1-4) subtype receptor mRNAs were generally positively correlated to both COX-1 and COX-2 in tumor tissue, but not in normal colon. Our results imply that both prostaglandin production (COX-2) and signaling via EP(1-4) subtype receptors, particularly EP(2), predict disease-specific mortality in colorectal cancer.