Tuomo J. Karttunen

Stage-dependent alterations of the serum cytokine pattern in colorectal carcinoma

Background:Inflammation contributes to the pathogenesis of colorectal cancer (CRC), and cytokine levels are altered during colorectal carcinogenesis.Methods:The serum levels of 13 cytokines and their relation to clinical and pathological parameters, and systemic inflammatory response (mGPS, CRP and neutrophil–lymphocyte ratio), were analysed from a prospective series of 148 CRC patients and 86 healthy age- and sex-matched controls.Results:CRC patients had higher serum platelet-derived growth factor, interleukin (IL)-6, IL-7, and IL-8 levels and lower monocyte chemotactic protein-1 (MCP-1) levels than the controls. A logistic regression model for discriminating the patients from the controls – including the five most predictive cytokines (high IL-8, high IL-6, low MCP-1, low IL-1ra, and low IP-10) – yielded an area under curve value of 0.890 in receiver operating characteristics analysis. Serum cytokines showed distinct correlation with other markers of systemic inflammatory response, and advanced CRCs were associated with higher levels of IL-8, IL-1ra, and IL-6. A metastasised disease was accompanied by an orientation towards Th2 cytokine milieu.Conclusion:CRC is associated with extensive alterations in serum cytokine environment, highlighting the importance of studying relative cytokine level alterations. Serum cytokine profile shows promise in separating CRC patients from healthy controls but its clinical value is yet to be confirmed.

Children with untreated food allergy express a relative increment in the density of duodenal γδ+ T cells.

(2000). Children with Untreated Food Allergy Express a Relative Increment in the Density of Duodenal ?d+ T Cells. Scandinavian Journal of Gastroenterology: Vol. 35, No. 11, pp. 1137-1142.Background: We investigated whether children with food allergy (FA) express increased densities of intraepithelialg x87 T cells similarly to subjects with celiac disease. Methods: The duodenal specimens taken by gastroduodenoscopy from 20 children with untreated FA, 17 with treated FA, 12 with celiac disease (CD) and 12 controls were studied with monoclonal antibodies and a three-layer peroxidase staining method. Results:The subjects with untreated FA expressed equal densities of total intraepithelial CD3x87 anda/b x87 T cells, but significantly higher densities of g x87 cells than the subjects currently on an elimination diet for FA or the controls. Accordingly, their g x87/CD3x87 ratio was higher. On the other hand, the results differed clearly from CD, where all the three cell populations showed high densities. Another finding that discriminated the subjects with FA from the CD patients was endoscopic examination. Lymphonodular hyperplasia (LNH) of the duodenal bulb with a normal villous pattern was demonstrated in 14 (70%) of the 20 subjects with untreated FA and in 8 (47%) of the 17 with treated FA, but in none of the celiac patients or controls. Surprisingly, the biopsy samples from the subjects with FA showed quite normal histological findings. Conclusions:According to this preliminary observation, high densities of intraepithelialg x87 T cells and a significantly elevated g x87/CD3x87 ratio are associated with untreated FA. If seen LNH in a gastroduodenoscopy and/or increased densities of g x87 T cells in the biopsy specimen, the possibility of gastrointestinal FA should be reliably assessed by a food challenge.

Loss of heterozygosity at 18q21 is indicative of recurrence and therefore poor prognosis in a subset of colorectal cancers.

SummaryAdjuvant therapies are increasingly used in colorectal cancers for the prevention of recurrence. These therapies have side-effects and should, thus, be used only if really beneficial. However, the development of recurrence cannot be predicted reliably at the moment of diagnosis, and targeting of adjuvant therapies is thus based only on the primary stage of the cancer. Loss of heterozygosity (LOH) in the long arm of chromosome 18 is suggested to be related to poor survival and possibly to the development of metastases. We studied the value of LOH at 18q21 as a marker of colorectal cancer prognosis, association with clinicopathological variables, tumour recurrence and survival of the patients. Of the 255 patients studied, 195 were informative as regards LOH status when analysed in primary colorectal cancer specimens using the polymerase chain reaction (PCR) and fragment analysis. LOH at 18q21 was significantly associated with the development of recurrence (P= 0.01) and indicated poor survival in patients of Dukes’ classes B and C, in which most recurrences (82%) occurred. An increased rate of tumour recurrence is the reason for poor survival among patients with LOH at 18q21 in primary cancer. These patients are a possible target group for recurrence-preventing adjuvant therapies.

Detailed analysis of inflammatory cell infiltration in colorectal cancer

Background:Higher-grade inflammatory infiltrate is a promising marker for better prognosis in colorectal cancer (CRC). However, the knowledge on the interrelationships between different inflammatory cells and classifications is fragmentary.Methods:We analysed the densities of eight types of inflammatory cells in a prospectively recruited group of 117 CRC patients and determined their interrelationships and contributions to Klintrup–Mäkinen (K–M) score of overall peritumoural inflammation. We characterised the inflammatory infiltrate in relation to stage and recurrences in 24-month follow-up.Results:There were high positive correlations between the inflammatory cell densities, with the exception of mast cells and CD1a+ immature dendritic cells. High K–M score associated with high peri- and intratumoural densities of CD3+, CD8+, CD68+, CD83+, and FoxP3+ cells and neutrophils. Advanced stage associated with low K–M score, as well as low CD3+, CD8+, CD83+, and FoxP3+ cell counts, of which low K–M score, low CD3+ T-cell count, and low FoxP3+ T-cell count were linked to higher recurrence rate.Conclusion:The density of CRC inflammatory infiltrate declines as stage advances. Especially, low K–M score and low T-cell counts predict higher recurrence rate. The high positive correlations between the individual inflammatory markers support the value of overall inflammatory reaction scoring.

Type I and III Collagens in Human Colon Cancer and Diverticulosis

BACKGROUNDnCollagens are major proteins in the extracellular matrix, providing tissues with tensile strength. They are also important for cell adhesion and the invasion of malignant tumours.nnnMETHODSnThirty-nine samples of human colon (24 diverticulosis, 6 malignant tumours, 9 controls) were collected during elective surgery. Immunoassays for different domains of type I and III collagens and procollagens were used in soluble tissue extracts and trypsin digests of tissue samples.nnnRESULTSnThe contents of cross-linked type I and III collagen telopeptides and total collagen were similar in diverticulosis and healthy tissue, whereas in malignant tissue maturely cross-linked type III collagen was scarce. Furthermore, some of the cross-linked type I telopeptide antigens were exceptionally small in size, indicating that the cross-linking of type I collagen in collagen fibres is impaired in cancer. The rate of type I collagen synthesis was clearly increased in malignancy, but not significantly in diverticulosis. However, type III collagen synthesis was increased in diverticulosis, but not in malignancy.nnnCONCLUSIONSnIn colon malignancy, the collagen cross-linking process was aberrant and the synthesis of type I collagen increased. In diverticulosis, the synthesis of type III collagen was increased, suggesting only moderately increased metabolic activity.Background: Collagens are major proteins in the extracellular matrix, providing tissues with tensile strength. They are also important for cell adhesion and the invasion of malignant tumours. Methods: Thirty-nine samples of human colon (24 diverticulosis, 6 malignant tumours, 9 controls) were collected during elective surgery. Immunoassays for different domains of type I and III collagens and procollagens were used in soluble tissue extracts and trypsin digests of tissue samples. Results: The contents of cross-linked type I and III collagen telopeptides and total collagen were similar in diverticulosis and healthy tissue, whereas in malignant tissue maturely cross-linked type III collagen was scarce. Furthermore, some of the cross-linked type I telopeptide antigens were exceptionally small in size, indicating that the cross-linking of type I collagen in collagen fibres is impaired in cancer. The rate of type I collagen synthesis was clearly increased in malignancy, but not significantly in diverticulosis. However, type III collagen synthesis was increased in diverticulosis, but not in malignancy. Conclusions: In colon malignancy, the collagen cross-linking process was aberrant and the synthesis of type I collagen increased. In diverticulosis, the synthesis of type III collagen was increased, suggesting only moderately increased metabolic activity.

Toll-like receptor 5 (TLR5) expression is a novel predictive marker for recurrence and survival in squamous cell carcinoma of the tongue

Background:Toll-like receptor 5 (TLR5) is an immune receptor recognising bacterial flagellin. Activation of TLR5 results in cancer invasion and cytokine release. As certain bacteria have been linked to oral cancer, we wanted to study TLR5 expression in oral tongue squamous cell carcinoma (OTSCC).Methods:Samples from 119 patients with OTSCC were obtained, including 101 samples of adjacent normal lingual mucosa. The TLR5 histoscore (0–300) was assessed semiquantitatively by immunohistochemistry in a blinded manner.Results:Toll-like receptor 5 was expressed in 84 normal epithelia and 118 cancer samples. Expression of TLR5 was increased in cancer when compared with normal lingual epithelium (median histoscore 15 vs 135). In cancer, higher TLR5 was associated with age of >70 years at the time of diagnosis, female gender and disease recurrence. No association between TLR5 expression and tumour grade, stage or treatment was found. In multivariate analysis, TLR5 was an independent predictor of cancer mortality (hazard ratio (HR) 3.587, 95% confidence interval (CI) (1.632–7.882)) and disease recurrence (HR 4.455, 95% CI (2.168–9.158)).Conclusion:Toll-like receptor 5 has a previously undescribed role in the pathophysiology of OTSCC and might represent a link between bacteria and cancer. It could be a useful marker for predicting recurrence or survival of OTSCC patients.

Helicobacter pylori-Associated Gastritis Evolution of Histologic Changes over 10 Years

BACKGROUNDnHelicobacter pylori seems to be the commonest cause of chronic gastritis, but the natural course of H. pylori-associated gastritis is largely obscure.nnnMETHODSnWe present a histologic follow-up of 39 patients with H. pylori-positive gastritis. Gastroscopies with stepwise biopsies were performed in all the patients at an interval of 10 years.nnnRESULTSnOf the patients 87% (34/39) had a persistent infection and showed a significant decrease in the grades of antral gastritis, eosinophilic granulocytes, corpus eosinophilic granulocytes, and foveolar hyperplasia and a significant increase in the grade of corpus neutrophilic granulocytes. The quantities of H. pylori as estimated histologically did not change significantly during the follow-up period in patients with a persistent infection. In the five other patients (13%) the H. pylori infection had apparently disappeared spontaneously, and this was accompanied by decreases in the amount of inflammatory cells in the gastric mucosa.nnnCONCLUSIONSnH. pylori infection in the gastric mucosa is chronic and may be associated with both regressive and progressive histologic changes. Spontaneous healing of H. pylori infection is possible and is associated with partial resolution of the inflammatory changes in the gastric mucosa.

Immunoelectron microscopic localization of laminin, type IV collagen, and type III pN-collagen in reticular fibers of human lymph nodes.

We studied the ultrastructural distribution of laminin, type IV collagen, and the amino terminal pro-peptide of type III collagen (type III pN-collagen) in normal human lymph nodes. After fixation with freshly prepared 4% paraformaldehyde mixed with 0.1% glutaraldehyde, cryoultramicrotomy proved to preserve the antigenicity of these proteins better than embedding in Lowicryl K4M. Sections were treated with rabbit antibodies against the 7S domain of human type IV collagen, the fragment P1 of human laminin, and the amino terminal pro-peptide of human type III pro-collagen, followed by anti-rabbit IgG conjugated to 10-nm colloidal gold. Laminin and type IV collagen were seen in the basement membrane structures of the blood vessels and in the walls of sinuses. The amorphous material between the collagenous fibers in locations corresponding to reticular fibers also contained laminin and type IV collagen. The amino terminal pro-peptide of type III pro-collagen was present in the collagenous fibers in reticular fibers and in the walls of blood vessels and sinuses. Therefore, a significant number of the type III collagen molecules in these fibers must have retained their amino terminal pro-peptide. These results indicate that the basement membrane proteins laminin and type IV collagen are genuine components of reticular fibers, as suggested earlier by immunohistochemical studies at the light microscopic level.

Intestinal immune activation in juvenile idiopathic arthritis and connective tissue disease

Objectives: To examine the prevalence of immune activation in gastrointestinal (GI) mucosa in children with juvenile idiopathic arthritis (JIA) or connective tissue disease (CTD). Study design: We studied 27 children (15 girls, mean age 9.8±4.8 years) with JIA/CTD and GI symptoms, including nine with oligoarthritis, nine with polyarthritis, two with systemic arthritis, three with enthesitis‐related arthritis, and four with various CTDs. The control group consists of 54 children (31 girls, mean age 11.3±6.3 years) with GI symptoms but shown to have no significant GI or rheumatoid disorder. The subjects were examined by gastroduodenoscopy (22 patients, 50 controls) and colonoscopy (23 patients, 16 controls). Intraepithelial CD3+, α/β+, and γ/δ+ lymphocytes were counted from duodenal and ileal biopsies. Results: Five patients with JIA/CTD (19%) had ulcerative colitis. Lymphoid nodular hyperplasia (LNH) was more common in the patients [74% (20/27)] than in the controls [16% (8/50), p = 0.001], as well in the duodenal bulb [29% (7/24) vs. 10% (5/50)], terminal ileum [74% (14/19) vs. 38% (5/13)], and the colon [50% (11/22) vs. 14% (2/14)]. In the duodenum, CD3, α/β+, and γ/δ+ lymphocytes counts were higher in JIA/CTD (p<0.05). In the ileum, γ/δ+ cell numbers had increased in JIA/CTD (p<0.05). Either LNH, increased γ/δ+ count, or both were more common in JIA/CTD [89% (24/27)] than in the controls [13% (7/54), p<0.0001]. Conclusions: The majority of children suffering from JIA or CTD with GI symptoms show abnormalities consistent with activation of the intestinal immune system. The aetiology of this reaction remains unknown, but similar features are seen in delayed‐type food allergy.

High Endothelial Venules of the Lymph Nodes Express Fas Ligand

Fas (CD95, APO-1) is widely expressed on lymphatic cells, and by interacting with its natural ligand (Fas-L), Fas induces apoptosis through a complex caspase cascade. In this study we sought to survey Fas-L expression in vascular and sinusoidal structures of human reactive lymph nodes. Immunohistochemical Fas-L expression was present in all paracortical high endothelial venules (HEVs), in cells lining the marginal sinus wall, and in a few lymphocytes, but only occasionally in non-HEV vascular endothelium. In the paracortical zone over 60% of all vessels and all paracortical HEVs showed Fas-L expression, whereas in the medullary zone less than 10% of the blood vessels were stained with Fas-L. Normal vessels outside lymph nodes mostly showed no Fas-L expression. We show that in human reactive lymph nodes Fas-L expression is predominantly present in HEVs. Because the circulating lymphocytes gain entry to nodal parenchyma by transendothelial migration through HEVs, the suggested physiological importance of Fas-L expression in these vessels lies in the regulation of lymphocyte access to lymph node parencyhyma by possibly inducing Fas/Fas-L mediated apoptosis of activated Fas-expressing lymphoid cells. The Fas-L expressing cells in the marginal sinus might have a similar function for cells accessing the node in afferent lymph.