Are Metastatic Central Lymph Nodes (D3 volume) in right-sided Colon Cancer a Sign of Systemic Disease? A sub-group Analysis of an Ongoing Multicenter Trial.

OBJECTIVE: Assess outcomes of patients with right-sided colon cancer with metastases in the D3 volume after personalized surgery. BACKGROUND: Patients with central lymph node metastasis (D3-PNG) are considered to have a systemic disease with a poor prognosis. A 3-dimensional definition of the dissection volume allows the removal of all central nodes. MATERIALS AND METHODS: D3-PNG includes consecutive patients from an ongoing clinical trial. Patients were stratified into residual disease negative (D3-RDN) and residual disease positive (D3-RDP) groups. D3-RDN was further stratified into 4 periods to identify a learning curve. A personalized D3 volume (defined through arterial origins and venous confluences) was removed " en bloc" through medial-to-lateral dissection, and the D3 volume of the specimen was analyzed separately. RESULTS: D3-PNG contained 42 (26 females, 63.1 SD 9.9 y) patients, D3-RDN:29 (17 females, 63.4 SD 10.1 y), and D3-RDP:13 (9 females, 62.2 SD 9.7 y). The mean overall survival (OS) days were D3-PNG:1230, D3-RDN:1610, and D3-RDP:460. The mean disease-free survival (DFS) was D3-PNG:1023, D3-RDN:1461, and D3-RDP:74 days. The probability of OS/DFS were D3-PNG:52.1%/50.2%, D3-RDN:72.9%/73.1%, D3-RDP: 7.7%/0%. There is a significant change in OS/DFS in the D3-RDN from 2011-2013 to 2020-2022 (both P =0.046) and from 2014-2016 to 2020-2022 ( P =0.028 and P =0.005, respectively). CONCLUSION: Our results indicate that surgery can achieve survival in most patients with central lymph node metastases by removing a personalized and anatomically defined D3 volume. The extent of mesenterectomy and the quality of surgery are paramount since a learning curve has demonstrated significantly improved survival over time despite the low number of patients. These results imply a place for the centralization of this patient group where feasible.

Reconstructing topography and extent of injury to the superior mesenteric artery plexus in right colectomy with extended D3 mesenterectomy: a composite multimodal 3-dimensional analysis.

BACKGROUND: Superior mesenteric artery plexus (SMAP) injury is reported to cause postoperative intractable diarrhea after pancreatic/colonic surgery with extended lymphadenectomy. This study aims to describe the SMAP microanatomy and extent of injury after right colectomy with extended D3 mesenterectomy for cancer. METHODS: Three groups (I) anatomical dissection, (II) postmortem histology, and (III) surgical specimen histology were included. Nerve count and area were compared between groups II and III and paravascular sheath thickness between groups I and II. 3D models were generated through 3D histology, nanoCT scanning, and finally through 3D printing. RESULTS: A total of 21 specimens were included as follows: Group (I): 5 (3 females, 80-93 years), the SMAP is a complex mesh surrounding the superior mesenteric artery (SMA), branching out, following peripheral arteries and intertwining between them, (II): 7 (5 females, 71-86 years), nerve count: 53 ± 12.42 (38-68), and area: 1.84 ± 0.50 mm2 (1.16-2.29), and (III): 9 (5 females, 55-69 years), nerve count: 31.6 ± 6.74 (range 23-43), and area: 0.889 ± 0.45 mm2 (range 0.479-1.668). SMAP transection injury is 59% of nerve count and 48% of nerve area at middle colic artery origin level. The median values of paravascular sheath thickness decreased caudally from 2.05 to 1.04 mm (anatomical dissection) and from 2.65 to 1.17 mm (postmortem histology). 3D histology models present nerve fibers exclusively within the paravascular sheath, and lymph nodes were observed only outside. NanoCT-derived models reveal oblique nerve fiber trajectories with inclinations between 35° and 55°. Two 3D-printed models of the SMAP were also achieved in a 1:2 scale. CONCLUSION: SMAP surrounds the SMA and branches within the paravascular sheath, while bowel lymph nodes and vessels lie outside. Extent of SMAP injury on histological slides (transection only) was 48% nerve area and 59% nerve count. The 35°-55° inclination range of SMAP nerves possibly imply an even larger injury when plexus excision is performed (lymphadenectomy). Reasons for later improvement of bowel function in these patients can lie in the interarterial nerve fibers between SMA branches.

DNA Repair Protein Expression and Oxidative/Nitrosative Stress in Ulcerative Colitis and Sporadic Colorectal Cancer.

BACKGROUND/AIM: Chronic inflammation generates large quantities of reactive oxygen and nitrogen species that damage DNA. DNA repair is important for cellular viability and genome integrity. MATERIALS AND METHODS: Expression levels of the DNA repair proteins OGG1, XPA, MLH1, PARP1, and XRCC6, which function in base excision repair, nucleotide excision repair, mismatch repair, single-strand break repair and double-strand break repair, respectively, were assessed using immunohistochemistry in ulcerative colitis and sporadic colorectal cancer biopsies. Levels of oxidative/ nitrosative stress biomarkers were also assessed. RESULTS: Ulcerative colitis and colorectal cancer lesions expressed significantly higher levels of all DNA repair proteins and oxidative/ nitrosative stress biomarkers compared to normal colonic mucosa. Ulcerative colitis had the highest XPA and XRCC6 expression. CONCLUSION: Oxidative/nitrosative stress is prevalent in the colon of both diseases. Nucleotide excision repair and non-homologous end-joining double-strand break repair may be compromised in colorectal cancer, but not in ulcerative colitis.

Role of the Wnt signaling pathway in keratoacanthoma.

BACKGROUND: Keratoacanthoma (KA) has a unique life cycle of rapid growth and spontaneous regression that shows similarities to the hair follicle cycle, which involves an active Wnt signaling during physiological regeneration. We analyzed the expression of the Wnt signaling proteins ß-catenin, Lef1, Sox9, and Cyclin D1 in young and old human KAs to investigate a possible role for Wnt signaling in KAs. AIM: To investigate the role of the Wnt/ß-catenin signaling pathway in human KAs. METHODS AND RESULTS: Formalin-fixed, paraffin-embedded tissue samples of 67 KAs were analyzed for protein expression using immunohistochemistry. The majority of KAs were positive for Sox9 and Cyclin D1 but not for nuclear-localized ß-catenin or Lef-1. No significant differences in protein expressions were seen between young and old KAs. However, we found a significant association between Ki67 and Cyclin D1 proteins (P= .008). CONCLUSIONS: The Wnt signaling pathway does not appear to play a significant role in the biogenesis of human KA. Sox9 overexpression may be indicative of inhibition of Wnt signaling. Sox-9 and Cyclin D1 are proliferation markers that are most likely transactivated by alternate signaling pathways.

The relationship between IFNL4 genotype and the rate of fibrosis in hepatitis C patients.

Introduction: IFNL4 rs12979860 genotype CC is associated with increased ALT activity and liver stiffness in hepatitis C virus (HCV) genotype (G) 3 infection but not in G1. The primary aim of this study is to assess an interaction between IFNL4 genotype, viral genotype and the stage of liver fibrosis. Secondary aims are to study the potential interactions between IFNL4 genotype, viral genotype and viral load as well as ALT levels. Methods: We performed a cross sectional study of patients with untreated chronic hepatitis C. Inflammation and liver fibrosis were scored using METAVIR. DNA was extracted from serum samples and the rs12979860 was genotyped using a custom made Taqman assay. Results: About 304 consecutive patients with chronic Hepatitis C were included. 52% had G1 infection and 48% had G3. Among patients with G3, advanced fibrosis or cirrhosis (F3F4) was present in 35% of the patients with IFNL4 CC and 28% with CT/TT (p = 0.24). Among patients with G1, F3F4 was present in 20% of the patients with IFNL4 CC and 19% with CT/TT (p = 0.52). IFNL4 CC was associated with higher mean value of normalized (n)ALT both in HCV G1 and G3 infection. Conclusions: IFNL4 genotype was not a predictor of advanced liver fibrosis in G3 or G1 infected patients. IFNL4 CC predicted a higher mean value of ALT among both G1 and G3 infected patients.

Real-life chromoendoscopy for dysplasia surveillance in ulcerative colitis.

AIM: To evaluate the use of chromoendoscopy for surveillance of ulcerative colitis in a real-life community hospital setting. METHODS: Patients with extensive ulcerative colitis, having disease duration of more than 8 years and who presented between the years of 1999 to 2013, were offered enrolment in this single cohort prospective study. All participants underwent standard bowel preparation with sodium phosphate and chromoendoscopy. Two expert endoscopists, novice to chromoendoscopy, evaluated each segment of the colon with standard-definition colonoscopes after spray application of 0.4% indigo carmine. All observed lesions were recorded and evaluated before being removed and/or biopsied. In addition, nontargeted biopsies were taken from each segment of the colon. The dysplasia detection rate and dysplasia detection yield were ascertained. RESULTS: A total of 21 neoplastic lesions (2 carcinomas, 4 of high-grade dysplasia and 15 of low-grade dysplasia) and 27 nondysplastic lesions were detected in 16 of the total 67 patients (70% male; median disease duration: 17 years; median age at diagnosis: 25 years; 92% aminosalicylate-treated). The dysplasia detection rate was 10.5% (7/67 patients). The dysplasia detection yield was 20.8% (10/48) for targeted biopsies and 3.5% (11/318) for nontargeted biopsies. The sensitivity and specificity for the macroscopic evaluation of neoplasia using chromoendoscopy were 48% [95% confidence interval (CI): 26%-70%] and 96% (95%CI: 93%-98%), respectively. The positive predictive and negative predictive values were 42% (95%CI: 27%-59%) and 97% (95%CI: 95%-98%), respectively. A total of 19/21 dysplastic lesions were detected in mucosa with histologic inflammation. CONCLUSION: Chromoendoscopy seems to be of value for dysplasia surveillance of ulcerative colitis in a community hospital setting. The yield of non-targeted biopsies is negligible.

Nondysplastic Ulcerative Colitis Has High Levels of the Homologous Recombination Repair Protein NUCKS1 and Low Levels of the DNA Damage Marker Gamma-H2AX.

Background: The colon and rectum are continuously exposed to oxidative stress that generates reactive oxygen species, which are a major cause of DNA double-strand breaks (DSB). Furthermore, chronic inflammatory diseases such as ulcerative colitis (UC) are characterized by an excess of reactive nitrogen species that can also lead to DNA double-strand breakage and genomic instability. We investigated the expression of the nuclear casein kinase and cyclin-dependent kinase substrate 1 (NUCKS1) protein in UC and sporadic colorectal cancer (CRC) due to its involvement in both DNA double-strand break repair and inflammatory signaling. Methods: NUCKS1 expression and expression of the DNA double-strand break marker gamma-H2AX (γH2AX) were assessed in formalin-fixed, paraffin-embedded UC and CRC patient biopsies using peroxidase immunohistochemistry. Expression levels for both proteins were evaluated together with previously published expression-level data for hTERT and TP53 proteins in the same material. Results: Nondysplastic UC lesions had 10-fold lower γH2AX expression and approximately 4-fold higher NUCKS1 expression compared with sporadic CRC, indicating minimal DNA DSB damage and heightened DNA DSB repair in these lesions, respectively. NUCKS1 expression in UC tended to decrease with increasing grades of dysplasia, whereas γH2AX, hTERT, and TP53 expression tended to increase with increasing grades of dysplasia. The highest γH2AX expression was seen in sporadic CRC, indicating considerable DNA DSB damage, whereas the highest NUCKS1 expression and hTERT expression were seen in nondysplastic UC. Conclusions: Overall, our data suggest that NUCKS1 may be involved in DNA DSB repair and/or inflammatory signaling in UC, but a more thorough investigation of both pathways in UC is warranted.

Multi-target stool DNA test in the surveillance of inflammatory bowel disease: a cross-sectional cohort study.

BACKGROUND AND AIM: Colonoscopic surveillance is recommended in patients with longstanding inflammatory bowel disease (IBD) as they are at increased risk of colorectal cancer (CRC). Non-invasive surveillance may improve compliance and access. Multi-target stool DNA (MT-sDNA) has been validated for screening of sporadic CRC but has not been assessed in IBD. Our aim was to assess the performance of a MT-sDNA test in a real-life surveillance setting of patients with longstanding IBD. MATERIAL AND METHODS: A total of 192 IBD patients enrolled from two prospective cohorts submitted an EDTA buffered stool sample and underwent chromo- or white light colonoscopy. Stools were assayed for methylated BMP3 & NDRG4, mutant KRAS and ß-actin by a laboratory blinded to clinical data. RESULTS: The multitarget-sDNA panel was positive in 2/2 CRC and 5/15 low-grade dysplasia (LGD) < 1 cm in diameter. Sensitivities were 100% (95% CI 16-100%) for CRC and 33% (95% CI 13-61%) for LGD lesions <1 cm, with specificities of 87% (95% CI 81-91%) and 93% (95% CI 88-96%), respectively. The estimated number of patients needed to screen to detect a single CRC was 96 (95% CI 93-99%) and was 28 (95% CI 22-34%) to detect any colorectal neoplasia (CRN). CONCLUSION: The MT-sDNA panel detected CRC in IBD. Sensitivity for sub-centimeter colorectal neoplasms in IBD patients appears similar to that observed in the general population. The test may be a valuable tool for detection of malignancy during structured surveillance of long-term IBD in a first line hospital setting.

Symptoms and Mucosal Changes Stable During Rapid Increase of Pediatric Celiac Disease in Norway.

OBJECTIVES: We aimed to study whether the incidence of pediatric celiac disease (CD) in South-Eastern Norway changed from 2000 to 2010. We also examined whether there was a change in symptoms and histopathological morphology in the duodenal biopsies during the same period. METHODS: In 3 hospitals in South-Eastern Norway, records from pediatric patients (0-14.9 years) diagnosed with CD during two 3-year periods (2000-2002 and 2008-2010) were reviewed. Only cases with a duodenal biopsy diagnosis of CD classified as Marsh grade 2 and 3a-c were included. Frequencies of symptoms, anthropometric data, and laboratory results were compared, in addition to re-examinations of histological sections from one of the hospitals. RESULTS: A total of 400 cases were diagnosed with a female to male ratio of 1.5:1. The incidence rate for 2000 to 2002 was 15.9 cases per 100,000 person-years (95% confidence interval 12.8-19.4), compared with 45.5 cases per 100,000 person-years during 2008 to 2010 (95% confidence interval 40.5-50.9), P < 0.001. The relative frequencies of symptoms and the distribution of histopathological changes were similar in the 2 periods, whereas weight z scores and hemoglobin levels were significantly lower in the first period. CONCLUSIONS: We found a 3-fold increase in the incidence rate for CD in the Norwegian pediatric population during the decade 2000 to 2010. Slightly higher weight and hemoglobin levels at diagnosis in the latter period may be due to improved CD awareness. Unaltered relative frequencies of symptoms and histopathological changes in the gut, however, suggest a true increase of CD in Norwegian children.

Induction of Apoptosis in Intestinal Toxicity to a Histone Deacetylase Inhibitor in a Phase I Study with Pelvic Radiotherapy.

PURPOSE: When integrating molecularly targeted compounds in radiotherapy, synergistic effects of the systemic agent and radiation may extend the limits of patient tolerance, increasing the demand for understanding the pathophysiological mechanisms of treatment toxicity. In this Pelvic Radiation and Vorinostat (PRAVO) study, we investigated mechanisms of adverse effects in response to the histone deacetylase (HDAC) inhibitor vorinostat (suberoylanilide hydroxamic acid, SAHA) when administered as a potential radiosensitiser. MATERIALS AND METHODS: This phase I study for advanced gastrointestinal carcinoma was conducted in sequential patient cohorts exposed to escalating doses of vorinostat combined with standard-fractionated palliative radiotherapy to pelvic target volumes. Gene expression microarray analysis of the study patient peripheral blood mononuclear cells (PBMC) was followed by functional validation in cultured cell lines and mice treated with SAHA. RESULTS: PBMC transcriptional responses to vorinostat, including induction of apoptosis, were confined to the patient cohort reporting dose-limiting intestinal toxicities. At relevant SAHA concentrations, apoptotic features (annexin V staining and caspase 3/7 activation, but not poly-(ADP-ribose)-polymerase cleavage) were observed in cultured intestinal epithelial cells. Moreover, SAHA-treated mice displayed significant weight loss. CONCLUSION: The PRAVO study design implemented a strategy to explore treatment toxicity caused by an HDAC inhibitor when combined with radiotherapy and enabled the identification of apoptosis as a potential mechanism responsible for the dose-limiting effects of vorinostat. To the best of our knowledge, this is the first report deciphering mechanisms of normal tissue adverse effects in response to an HDAC inhibitor within a combined-modality treatment regimen.

Tp53/p53 status in keratoacanthomas.

BACKGROUND: Keratoacanthoma (KA) is a common keratinocytic skin neoplasm that typically develops rapidly and undergoes complete spontaneous regression. As the pro-apoptotic p53 protein may be involved in the lifecycle of KA, we studied the p53 status throughout the main stages of KA that include proliferation, maturation and regression in a large series of lesions. METHODS: One-hundred and twenty-four KAs were characterized with respect to age of the lesions both clinically and histopathologically, in addition to phenotypic characteristics such as cellular atypia, infiltration, inflammation and fibrosis. Tp53 mutations were detected by capillary electrophoresis, and p53 protein levels were assessed by immunohistochemistry. RESULTS: Tp53 mutations were detected in 49 cases (39.5%) and were associated with high p53 protein levels (p = 0.007) and histopathologic age of the lesions (p = 0.044). Significant association was also seen between high p53 protein levels and atypia (p = 0.036), whereas the association with infiltration showed borderline significance (p = 0.057). High p53 protein levels were significantly associated with gene mutations in transplanted, but not in non-transplanted patients. CONCLUSION: We show a high frequency of Tp53 mutations in KAs that is associated with increased p53 levels. The results indicate a role for the p53 protein in KA development.

TP53/p53 alterations and Aurora A expression in progressor and non-progressor colectomies from patients with longstanding ulcerative colitis.

Aneuploidy is a common feature in the colonic mucosa of patients suffering from the inflammatory bowel disease ulcerative colitis (UC) and often precedes the development of dysplasia and cancer. Aneuploidy is assumed to be caused by missegregation of chromosomes during mitosis, often due to a faulty spindle assembly checkpoint. p53 is a tumour suppressor protein known to regulate the spindle assembly checkpoint and is frequently mutated in aneuploid cells. Aurora A is a presumed oncoprotein, also involved in regulation of the spindle assembly checkpoint. In the present study, we examined the mutational frequency of TP53 and the protein levels of p53 in a set of 20 progressor and 10 non-progressor colectomies from patients suffering from longstanding UC. In addition, we re-examined previously published immunohistochemical data on Aurora A expression using the same material. Levels of Aurora A were re-examined with regard to DNA ploidy status and dysplasia within the progressors, as well as in relation to p53 accumulation and TP53 mutational status. We detected p53 accumulation only within the progressor colectomies, where it could be followed back 14 years prior to the colectomies, in pre-colectomy biopsies. TP53 mutations were detected in both progressors and non-progressors. Expression levels of Aurora A were similar in the progressors and non-progressors. Within the group of progressors however, low levels of Aurora A were associated with areas of DNA aneuploidy, as well as with increasing degrees of dysplasia. Our results indicate that alterations in p53 may be an early biomarker of a progressor colon, and that p53 is accumulated early in UC-related carcinogenesis. Furthermore, a decreased Aurora A expression is associated with the development of DNA aneuploidy, as well as with dysplasia in UC progressors.

Reduced hTERT protein levels are associated with DNA aneuploidy in the colonic mucosa of patients suffering from longstanding ulcerative colitis.

Longstanding ulcerative colitis (UC) is a disease of chronic inflammation of the colon. It is associated with the development of colorectal cancer through a multistep process including increasing degrees of dysplasia and DNA-ploidy changes. However, not all UC patients will develop these characteristics even during lifelong disease, and patients may therefore be divided into progressors who develop dysplasia or cancer, and non-progressors who do not exhibit such changes. In the present study, the amount of hTERT, the catalytic subunit of the enzyme telomerase, was estimated by using peroxidase immunohistochemistry (IHC) in a set of progressor and non-progressor UC colectomies. The protein levels in the colonic mucosa of the progressors and non­progressors were compared, and further comparisons between different categories of dysplastic development and to DNA-ploidy status within the progressors were made. Levels of hTERT were elevated in the colonic mucosa of the progressors and non-progressors when compared to non-UC control samples, but no difference was observed between the hTERT levels in the mucosa of progressors and non-progressors. The levels of hTERT associated with levels of Ki67 to a significant degree within the non-progressors. hTERT expression in lesions with DNA-aneuploidy were decreased as compared to diploid lesions, when stratified for different classes of colonic morphology. Our results indicate an association between hTERT protein expression and aneuploidy in UC-progressor colons, and also a possible protective mechanism in the association between hTERT and Ki67, against development of malignant features within the mucosa of a UC-colon.

Telomere shortening correlates to dysplasia but not to DNA aneuploidy in longstanding ulcerative colitis.

BACKGROUND: Ulcerative colitis (UC) is a chronic, inflammatory bowel disease which may lead to dysplasia and adenocarcinoma in patients when long-lasting. Short telomeres have been reported in mucosal cells of UC patients. Telomeres are repetitive base sequences capping the ends of linear chromosomes, and protect them from erosion and subsequent wrongful recombination and end-to-end joining during cell division. Short telomeres are associated with the development of chromosomal instability and aneuploidy, the latter being risk factors for development of dysplasia and cancer. Specifically, the abrupt shortening of one or more telomeres to a critical length, rather than bulk shortening of telomeres, seems to be associated with chromosomal instability. METHODS: We investigated possible associations between dysplasia, aneuploidy and telomere status in a total of eight lesions from each of ten progressors and four nonprogressors suffering from longstanding UC. We have analyzed mean telomere length by qPCR, as well as the amount of ultra-short telomeres by the Universal STELA method. RESULTS: An increased amount of ultra-short telomeres, as well as general shortening of mean telomere length are significantly associated with dysplasia in longstanding UC. Furthermore, levels of ultra-short telomeres are also significantly increased in progressors (colons harbouring cancer/dysplasia and/or aneuploidy) compared to nonprogressors (without cancer/dysplasia/aneuploidy), whereas general shortening of telomeres did not show such associations. CONCLUSIONS: Our data suggest that ultra-short telomeres may be more tightly linked to colorectal carcinogenesis through development of dysplasia in UC than general telomere shortening. Telomere status was not seen to associate with DNA aneuploidy.

The mucosal and systemic immune responses elicited by a chitosan-adjuvanted intranasal influenza H5N1 vaccine.

BACKGROUND: Development of influenza vaccines that induce mucosal immunity has been highlighted by the World Health Organisation as a priority (Vaccine 2005;23:1529). Dose-sparing strategies and an efficient mass-vaccination regime will be paramount to reduce the morbidity and mortality of a future H5N1 pandemic. OBJECTIVES: This study has investigated the immune response and the dose-sparing potential of a chitosan-adjuvanted intranasal H5N1 (RG-14) subunit (SU) vaccine in a mouse model. METHODS: Groups of mice were intranasally immunised once or twice with a chitosan (5 mg/ml)-adjuvanted SU vaccine [7·5, 15 or 30 µg haemagglutinin (HA)] or with a non-adjuvanted SU vaccine (30 µg HA). For comparison, another group of mice were intranasally immunised with a whole H5N1 (RG-14) virus (WV) vaccine (15 µg HA), and the control group consisted of unimmunised mice. RESULTS: The chitosan-adjuvanted SU vaccine induced an immune response superior to that of the non-adjuvanted SU vaccine. Compared with the non-adjuvanted SU group, the chitosan-adjuvanted SU vaccine elicited higher numbers of influenza-specific antibody-secreting cells (ASCs), higher concentrations of local and systemic antibodies and correspondingly an improved haemagglutination inhibition (HI) and single radial haemolysis (SRH) response against both the homologous vaccine strain and drifted H5 strains. We measured a mixed T-helper 1/T-helper 2 cytokine response in the chitosan-adjuvanted SU groups, and these groups had an increased percentage of virus-specific CD4(+) T cells producing two Thelper 1 (Th1) cytokines simultaneously compared with the non-adjuvanted SU group. Overall, the WV vaccine induced higher antibody concentrations in sera and an HI and SRH response similar to that of the chitosan-adjuvanted SU vaccine. Furthermore, the WV vaccine formulation showed a stronger bias towards a T-helper 1 profile than the SU vaccine and elicited the highest frequencies of CD4(+) Th1 cells simultaneously secreting three different cytokines (INFγ(+) , IL2(+) and INFα(+) ). As expected, two immunisations gave a better immune response than one in all groups. The control group had very low or not detectable results in the performed immunoassays. CONCLUSION: The cross-clade serum reactivity, improved B- and T-cell responses and dose-sparing potential of chitosan show that a chitosan-adjuvanted intranasal influenza vaccine is a promising candidate vaccine for further preclinical development.

Improved lymph node harvest from resected colon cancer specimens did not cause upstaging from TNM stage II to III.

BACKGROUND: The number of lymph nodes retrieved and examined from a resected colon cancer specimen may be crucial for correct staging. We examined if efforts to increase the lymph node harvest to more than 12 lymph nodes per specimen would upstage some patients from TNM stage II to III. METHODS: Three hospitals compared results from 2000 with those of 2007 in 421 resected patients with stage II and III colon cancer. Hospital A endeavored to improve the surgical procedure while the pathologists enhanced the quality of lymph node sampling. Hospital B did not make any marked changes, while hospital C introduced the GEWF lymph node solvent (glacial acetic acid, ethanol, distilled water, and formaldehyde) in their pathology method. RESULTS: In 2000, 12 or more lymph nodes were harvested in 39.6, 45.0, and 21.1% of the specimens from the three hospitals, while the figures for 2007 were 85.7, 42.0, and 90.3%, respectively. The significant increase in lymph node harvest in two of the hospitals in 2007 compared to 2000 (p < 0.001) did not affect the share of patients with stage III in 2007 (38.7%) compared to 2000 (44.1%) (p = 0.260). The number of positive lymph nodes and the lymph node ratio (LNR) decreased from 2000 to 2007. A lymph node yield of 12 or more was not associated with an increased probability of positive lymph nodes in a multivariable logistic regression analysis. CONCLUSION: More radical surgery and dedicated pathologists and the use of the GEWF solvent significantly increased the lymph node yield but did not upstage patients from TNM stage II to III.

The endothelin axis in the metastatic process of colon carcinoma.

BACKGROUND: The endothelin axis has recently emerged as an important factor in tumour metastasis. The aim of this study was to investigate the endothelin axis and its downstream pathways related to metastasis in colon carcinoma. MATERIALS AND METHODS: mRNA expression of 36 genes associated with the endothelin axis in 18 non-metastatic and 20 metastatic colon carcinomas with individual-matched normal mucosa were evaluated using real-time reverse transcription polymerase chain reaction. RESULTS: Seventeen out of 36 genes, including endothelin A receptor, were significantly overexpressed in the tumour tissue compared to the individual-matched normal mucosa. Seven out of 36 genes, including endothelin B receptor, were significantly down-regulated in tumour tissue. Phosphatase and tensin homolog (PTEN) was significantly down-regulated in the metastatic patients compared to the non-metastatic patients. CONCLUSION: This study indicated that central genes in the endothelin axis are overexpressed when colon tissue becomes malignant. Down-regulation of PTEN may promote a progressive phenotype of colon carcinomas.

Lymph node micrometastases and isolated tumor cells influence survival in stage I and II colon cancer.

PURPOSE: Lymph-node status is considered the most important prognostic factor in colorectal cancer. The aim of the present prospective study was to evaluate the influence of micrometastases and isolated tumor cells on recurrence and disease-free survival in colon cancer. METHODS: A total of 193 patients with colon cancer, operated on between 2000 and 2005, were enrolled in the study. All lymph nodes were examined by routine microscopy in hematoxylin and eosin-stained sections. If no metastases were identified in any node, all nodes were examined immunohistochemically with monoclonal antibody CAM 5.2. RESULTS: Ordinary metastases were found in 67 patients, leaving 126 patients in stage I/II. Immunohistochemistry showed that 5% (6/126) of these had micrometastases and 26% (33/126) had isolated tumor cells. A median of 5 years of follow-up revealed local or distant recurrence in 23% (9/39) of stage I/II patients with micrometastases or isolated tumor cells, compared with 7% (6/87) without micrometastases or isolated tumor cells (P = .010). Five-year disease-free survival for patients with and without micrometastases or isolated tumor cells was 75% and 93%, respectively (P = .012). When analyzed separately, patients with isolated tumor cells (excluding micrometastases) had also lower survival than node-negative patients (P = .012). CONCLUSION: The presence of micrometastases and isolated tumor cells was found to be a prognostic factor for recurrence and disease-free survival. This may have implications for future treatment of stage I/II colon cancer.

Evaluation of suitable reference genes for normalization of real-time reverse transcription PCR analysis in colon cancer.

BACKGROUND: Real-time reverse transcription PCR (qRT-PCR) is frequently used for gene expression quantification due to its methodological reproducibility and sensitivity. The gene expression is quantified by normalization to one or more reference genes which are presumed stably expressed throughout a given experiment. The aim of this study was to validate a standardized experimental setup to identifying reference genes for normalization of qRT-PCR in the metastatic and non-metastatic colon cancer. METHODS: In this study, expression of 16 commonly used reference genes was quantified in tumour tissue and individual-matched normal mucosa in 18 non-metastatic colon cancer patients and 20 colon cancer patients with distant metastases using TaqMan Low Density Array (TLDA). The expression stability was determined and compared by means of geNorm and NormFinder. RESULTS: Two pairs of genes, HPRT1/PPIA and IPO8/PPIA, were identified to be suitable to normalize gene expression data in metastatic and non-metastatic colon cancer patients, according to geNorm and NormFinder respectively. CONCLUSION: We propose a standardized approach of finding the most suitable reference gene(s) in every qRT-PCR experiment using TLDA.

The Prognostic Impact of Protein Expression of E-Cadherin-Catenin Complexes Differs between Rectal and Colon Carcinoma.

The E-cadherin-catenin complex provides cell-cell adhesion. In order for a carcinoma to metastasize, cancer cells must let go of their hold of neighboring cells in the primary tumor. The presence of components of the E-cadherin-catenin complex in 246 rectal adenocarcinomas was examined by immunohistochemistry and compared to their presence in 219 colon carcinomas. The expression data were correlated to clinical information from the patients' records. There were statistically significant differences in protein expression between the rectal and the colon carcinomas regarding membranous beta-catenin, gamma-catenin, p120-catenin, and E-cadherin, as well as nuclear beta-catenin. In the rectal carcinomas, there was a significant inverse association between the expression of p120-catenin in cell membranes of the primary tumors and the occurrence of local recurrence, while membranous protein expression of beta-catenin was inversely related to distant metastases.