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Lynch syndrome (LS) is the most common inherited cancer syndrome. It is inherited via a monoallelic germline variant in one of the DNA mismatch repair (MMR) genes. LS carriers have a broad 30-80% risk of developing various malignancies, and more precise, individual risk estimations would be of high clinical value, allowing tailored cancer prevention and surveillance. Due to MMR deficiency, LS cancers are characterized by the accumulation of frameshift mutations leading to highly immunogenic frameshift peptides (FSPs). Thus, immune surveillance is proposed to inhibit the outgrowth of MMR-deficient cell clones. Recent studies have shown that immunoediting during the evolution of MMR-deficient cancers leads to a counter-selection of highly immunogenic antigens. The immunogenicity of FSPs is dependent on the antigen presentation. One crucial factor determining antigen presentation is the HLA genotype. Hence, a LS carrier's HLA genotype plays an important role in the presentation of FSP antigens to the immune system, and may influence the likelihood of progression from pre-cancerous lesions to cancer. To address the challenge of clarifying this possibility including diverse populations with different HLA types, we have established the INDICATE initiative (Individual cancer risk by HLA type, http://indicate-lynch.org/), an international network aiming at a systematic evaluation of the HLA genotype as a possible cancer risk modifier in LS. Here we summarize the current knowledge on the role of HLA type in cancer risk and outline future research directions to delineate possible association in the scenario of LS with genetically defined risk population and highly immunogenic tumors. peerReviewed

The NRF2 pathway is frequently activated in various cancer types, yet a comprehensive analysis of its effects across different malignancies is currently lacking. We developed a NRF2 activity metric and utilized it to conduct a pan-cancer analysis of oncogenic NRF2 signaling. We identified an immunoevasive phenotype where high NRF2 activity is associated with low interferon-gamma (IFNγ), HLA-I expression and T cell and macrophage infiltration in squamous malignancies of the lung, head and neck area, cervix and esophagus. Squamous NRF2 overactive tumors comprise a molecular phenotype with SOX2/TP63 amplification, TP53 mutation and CDKN2A loss. These immune cold NRF2 hyperactive diseases are associated with upregulation of immunomodulatory NAMPT, WNT5A, SPP1, SLC7A11, SLC2A1 and PD-L1. Based on our functional genomics analyses, these genes represent candidate NRF2 targets, suggesting direct modulation of the tumor immune milieu. Single-cell mRNA data shows that cancer cells of this subtype exhibit decreased expression of IFNγ responsive ligands, and increased expression of immunosuppressive ligands NAMPT, SPP1 and WNT5A that mediate signaling in intercellular crosstalk. In addition, we discovered that the negative relationship of NRF2 and immune cells are explained by stromal populations of lung squamous cell carcinoma, and this effect spans multiple squamous malignancies based on our molecular subtyping and deconvolution data. peerReviewed

Background Immune response against cancer has prognostic impact but its role in gastric cancer is poorly known. The aim of the study was to assess the prognostic significance of immune cell score (CD3+, CD8+), tumour immune escape (PD-L1, PD-1) and immune tolerance (Clever-1). Methods After exclusion of Epstein-Barr virus positive (n = 4) and microsatellite instable (n = 6) tumours, the study included 122 patients with GC undergoing D2 gastrectomy. CD3+ and CD8+ based ICS, PD-L1, PD-1 and Clever-1 expressions were evaluated. Differences in survival were examined using Cox regression adjusted for confounders. The primary outcome was 5-year survival. Results The 5-year overall survival rate was 43.4%. High ICS was associated with improved overall survival (adjusted HR 0.48 (95% CI 0.26–0.87)) compared to low ICS. In the high ICS group, patients with PD-L1 expression (5-year survival 69.2 vs. 53.1%, p = 0.317), high PD-1 (5-year survival 70.6 vs. 55.3% p = 0.312) and high Clever-1 (5-year survival 72.0% vs. 45.5% (p = 0.070) had poor prognosis. Conclusions High ICS was associated with improved survival. In the high ICS group, patients with high PD-L1, PD-1 and Clever-1 had poor prognosis highlighting the importance of immune escape and immune tolerance in GC.

Like many bacterial species, Borrelia burgdorferi, the pleomorphic bacterium that causes Lyme borreliosis, produces outer membrane vesicles (OMVs). Borrelial OMVs (BbOMVs) have been identified as containing virulence factors, such as outer surface proteins (Osps) A, B, and C, as well as DNA. However, the pathogenicity of BbOMVs in disease development is still unclear. In this study, we characterized purified BbOMVs by analyzing their size and immunolabeling for known antigenic markers: OspA, OspC, p39, and peptidoglycan. In addition, BbOMVs were cocultured with human non-immune cells for cytotoxicity analysis. The results demonstrated that, on average, the vesicles were small, ranging between 11 and 108 nm in diameter. In addition, both OspA and OspC, as well as Lyme arthritis markers p39 and peptidoglycan, were detected from BbOMVs. Furthermore, BbOMVs were cocultured with non-immune cells, which did not result in cell death. Combined, these results suggested that BbOMVs could participate in the induction of infection by functioning as a decoy for the host immune system. Furthermore, BbOMVs might serve as a means for persistent antigens to remain in the host for prolonged periods of time. peerReviewed

Liikunta vahvistaa tutkitusti rokotusten tehoa. Liikuntaharrastus pienentää myös riskiä sairastua vakavaan koronatautiin. nonPeerReviewed

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Lyme borreliosis is a multisystemic disease caused by the pleomorphic bacteria of the Borrelia burgdorferi sensu lato complex. The exact mechanisms for the infection to progress into a prolonged sequelae of the disease are currently unknown, although immune evasion and persistence of the bacteria in the host are thought to be major contributors. The current study investigated B. burgdorferi infection processes in two human cell lines, both non-immune and non-phagocytic, to further understand the mechanisms of infection of this bacterium. By utilizing light, confocal, helium ion, and transmission electron microscopy, borrelial infection of chondrosarcoma (SW1353) and dermal fibroblast (BJ) cells were examined from an early 30-min time point to a late 9-days post-infection. Host cell invasion, viability of both the host and B. burgdorferi, as well as, co-localization with lysosomes and the presence of different borrelial pleomorphic forms were analyzed. The results demonstrated differences of infection between the cell lines starting from early entry as B. burgdorferi invaded BJ cells in coiled forms with less pronounced host cell extensions, whereas in SW1353 cells, micropodial interactions with spirochetes were always seen. Moreover, infection of BJ cells increased in a dose dependent manner throughout the examined 9 days, while the percentage of infection, although dose dependent, decreased in SW1353 cells after reaching a peak at 48 h. Furthermore, blebs, round body and damaged B. burgdorferi forms, were mostly observed from the infected SW1353 cells, while spirochetes dominated in BJ cells. Both infected host cell lines grew and remained viable after 9 day post-infection. Although damaged forms were noticed in both cell lines, co-localization with lysosomes was low in both cell lines, especially in BJ cells. The invasion of non-phagocytic cells and the lack of cytopathic effects onto the host cells by B. burgdorferi indicated one mechanism of immune evasion for the bacteria. The differences in attachment, pleomorphic form expressions, and the lack of lysosomal involvement between the infected host cells likely explain the ability of a bacterium to adapt to different environments, as well as, a strategy for persistence inside a host. peerReviewed

Immunological and epigenetic changes are interconnected and contribute to tumorigenesis. We determined the immunoprofiles and promoter methylation of inflammation-related genes for colitis-associated colorectal carcinomas (CA-CRC). The results were compared with Lynch syndrome (LS)-associated colorectal tumors, which are characterized by an active immune environment through inherited mismatch repair defects. CA-CRCs (n = 31) were immunohistochemically evaluated for immune cell scores (ICSs) and PDCD1 and CD274 expression. Seven inflammation-associated genes (CD274, NTSR1, PPARG, PTGS2, PYCARD, SOCS1, and SOCS2), the repair gene MGMT, and eight standard marker genes for the CpG Island Methylator Phenotype (CIMP) were investigated for promoter methylation in CA-CRCs, LS tumors (n = 29), and paired normal mucosae by multiplex ligation-dependent probe amplification. All but one CA-CRCs were microsatellite-stable and all LS tumors were microsatellite-unstable. Most CA-CRCs had a high ICS (55%) and a positive CD274 expression in immune cells (52%). NTSR1 revealed frequent tumor-specific hypermethylation in CA-CRC and LS. When compared to LS mucosae, normal mucosae from patients with CA-CRC showed significantly higher methylation of NTSR1 and most CIMP markers. In conclusion, CA-CRCs share a frequent ICShigh/CD274pos expression pattern with LS tumors. Elevated methylation in normal mucosa may indicate field cancerization as a feature of CA-CRC-associated tumorigenesis.

Simple Summary Metastasis is the main cause for cancer mortality. The most common metastatic sites of colorectal cancer (CRC) are the liver and lungs. Tumour-infiltrating lymphocytes are recognized as beneficial prognostic factors both in primary and metastatic CRC, but less is known about their reciprocal differences. The aim of our study was to evaluate immune microenvironment and its prognostic value in a series of mismatch proficient (pMMR) CRC with matched liver and lung metastases. The proportion of tumours with high immune cell infiltration together with PD-L1-positivity almost doubled in metastases compared to primary tumours. Our study confirmed the prognostic value of high ICS in least immune-infiltrated metastases in pMMR CRC patients. Major differences observed in immune contexture between primary tumours and metastases may have significance for treatment strategies for patients with advanced CRC. Purpose: To evaluate immune cell infiltration, the programmed death-1/programmed death ligand-1 (PD-1/PD-L1) expression and their prognostic value in a series of mismatch proficient (pMMR) CRC with matched liver and lung metastases. Methods: Formalin-fixed paraffin-embedded tissue sections stained for CD3, CD8, PD-L1 and PD-1 from 113 primary CRC tumours with 105 liver and 59 lung metastases were analyzed. The amount of CD3 and CD8 positive lymphocytes were combined as immune cell score (ICS). Comparative analyses on immune contexture were performed both between the primary tumour and matched metastases and between the metastatic sites. Results: In liver metastases, immune cell infiltration was increased in general compared to primary tumours but did not correlate case by case. On the contrary, ICS between lung metastases and primary tumours correlated well, but the expression of PD-1/PD-L1 was increased in lung metastases. The proportion of tumours with high ICS together with PD-L1-positivity almost doubled in metastases (39%) compared to primary tumours (20%). High ICS (compared to lowest) in patient's least immune-infiltrated metastasis was an independent prognostic marker for disease-specific (HR 9.14, 95%CI 2.81-29.68) and overall survival (HR 6.95, 95%CI 2.30-21.00). Conclusions: Our study confirms the prognostic value of high ICS in least immune-infiltrated metastases in pMMR CRC patients. Major differences observed in immune contexture between primary tumours and metastases may have significance for treatment strategies for patients with advanced CRC.

Hyaluronan (HA) accumulation has been associated with poor survival in various cancers, but the mechanisms for this phenomenon are still unclear. The aim of this study was to investigate the prognostic significance of stromal HA accumulation and its association with host immune response in pancreatic ductal adenocarcinoma (PDAC). The study material consisted of 101 radically treated patients for PDAC from a single geographical area. HA staining was evaluated using a HA-specific probe, and the patterns of CD3, CD8, CD73 and PD-L1 expression were evaluated using immunohistochemistry. HA staining intensity of tumour stromal areas was assessed digitally using QuPath. CD3- and CD8-based immune cell score (ICS) was determined. High-level stromal HA expression was significantly associated with poor disease-specific survival (p = 0.037) and overall survival (p = 0.013) In multivariate analysis, high-level stromal HA expression was an independent negative prognostic factor together with histopathological grade, TNM stage, CD73 positivity in tumour cells and low ICS. Moreover, high-level stromal HA expression was associated with low ICS (p = 0.017). In conclusion, stromal HA accumulation is associated with poor survival and low immune response in PDAC.

Aptamers are short, single-stranded DNA or RNA oligonucleotide molecules that function as synthetic analogs of antibodies and bind to a target molecule with high specificity. Aptamer affinity entirely depends on its tertiary structure and charge distribution. Therefore, length and structure optimization are essential for increasing aptamer specificity and affinity. Here we present a general optimization procedure for finding most populated atomistic structures of DNA aptamers. Based on the existed aptamer LC-18 for lung adenocarcinoma, a new truncated aptamer LC-18t was developed. A three-dimensional shape of LC-18t was reported based on small-angle X-ray scattering (SAXS) experiments and molecular modeling by fragment molecular orbital or molecular dynamic methods. Molecular simulations revealed an ensemble of possible aptamer conformations in solution that were in close agreement with measured SAXS data. The truncated aptamer LC-18t had stronger binding to cancerous cells in lung tumor tissues and shared the binding site with the original larger aptamer. The suggested approach reveals 3D shapes of aptamers and helps in designing better affinity probes. peerReviewed

The immune system can recognize and attack cancer cells, especially those with a high load of mutation-induced neoantigens. Such neoantigens are abundant in DNA mismatch repair (MMR)-deficient, microsatellite-unstable (MSI) cancers. MMR deficiency leads to insertion/deletion (indel) mutations at coding microsatellites (cMS) and to neoantigen-inducing translational frameshifts. Here, we develop a tool to quantify frameshift mutations in MSI colorectal and endometrial cancer. Our results show that frameshift mutation frequency is negatively correlated to the predicted immunogenicity of the resulting peptides, suggesting counterselection of cell clones with highly immunogenic frameshift peptides. This correlation is absent in tumors with Beta-2-microglobulin mutations, and HLA-A*02:01 status is related to cMS mutation patterns. Importantly, certain outlier mutations are common in MSI cancers despite being related to frameshift peptides with functionally confirmed immunogenicity, suggesting a possible driver role during MSI tumor evolution. Neoantigens resulting from shared mutations represent promising vaccine candidates for prevention of MSI cancers., DNA mismatch repair (MMR)-deficient cancers with microsatellite-instability are characterized by a high load of frameshift mutation-derived neoantigens. Here, by mapping the frameshift mutation landscape and predicting the immunogenicity of the resulting peptides, the authors show evidence of immunoediting in MMR-deficient colorectal and endometrial cancers.

There is insufficient evidence to support screening of various tick-borne diseases (TBD) related microbes alongside Borrelia in patients suffering from TBD. To evaluate the involvement of multiple microbial immune responses in patients experiencing TBD we utilized enzyme-linked immunosorbent assay. Four hundred and thirty-two human serum samples organized into seven categories followed Centers for Disease Control and Prevention two-tier Lyme disease (LD) diagnosis guidelines and Infectious Disease Society of America guidelines for post-treatment Lyme disease syndrome. All patient categories were tested for their immunoglobulin M (IgM) and G (IgG) responses against 20 microbes associated with TBD. Our findings recognize that microbial infections in patients suffering from TBDs do not follow the one microbe, one disease Germ Theory as 65% of the TBD patients produce immune responses to various microbes. We have established a causal association between TBD patients and TBD associated co-infections and essential opportunistic microbes following Bradford Hill’s criteria. This study indicated an 85% probability that a randomly selected TBD patient will respond to Borrelia and other related TBD microbes rather than to Borrelia alone. A paradigm shift is required in current healthcare policies to diagnose TBD so that patients can get tested and treated even for opportunistic infections.

Illnesses caused by a variety of micro- and macro- organisms can negatively affect individuals’ fitness, leading to the expectation that immunity is under positive selection. However, immune responses are costly and individuals must trade-off their immune response with other fitness components (e.g. survival or reproductive success) meaning that individuals with intermediate response may have the greatest overall fitness. Such a process might be particularly acute in species with strong sexual selection because the condition-dependence of male secondary sexual-traits might lead to striking phenotypic differences amongst males of different immune response levels. We tested whether there is selection on immune response by survival and reproduction in yearling and adult male black grouse (Lyrurus tetrix) following an immune challenge with a novel antigen and tested the hypothesis that sexual signals and body mass are honest signals of the immune response. We show that yearling males with highest immune response to these challenges had higher survival, but the reverse was true for adults. Adults with higher responses had highest mass loss and adult males with intermediate immune response had highest mating success. Tail length was related to baseline response in adults and more weakly in yearlings. Our findings reveal the complex fitness consequences of mounting an immune response across age classes. Such major differences in the direction and magnitude of selection in multiple fitness components is an alternative route underpinning the stabilising selection of immune responses with an intermediate immune response being optimal. peerReviewed

Exercise and exercise-induced weight loss have a beneficial effect on overall health, including positive effects on molecular pathways associated with immune function, especially in overweight individuals. The main aim of our study was to assess how energy deprivation (i.e., "semi-starvation") leading to substantial fat mass loss affects the immune system and immunosuppression in previously normal weight individuals. Thus, to address this hypothesis, we applied a high-throughput systems biology approach to better characterize potential key pathways associated with immune system modulation during intensive weight loss and subsequent weight regain. We examined 42 healthy female physique athletes (age 27.5 +/- 4.0 years, body mass index 23.4 +/- 1.7 kg/m(2)) volunteered into either a diet group (n = 25) or a control group (n = 17). For the diet group, the energy intake was reduced and exercise levels were increased to induce loss of fat mass that was subsequently regained during a recovery period. The control group was instructed to maintain their typical lifestyle, exercise levels, and energy intake at a constant level. For quantification of systems biology markers, fasting blood samples were drawn at three time points: baseline (PRE), at the end of the weight loss period (MID 21.1 +/- 3.1 weeks after PRE), and at the end of the weight regain period (POST 18.4 +/- 2.9 weeks after MID). In contrast to the control group, the diet group showed significant (false discovery rate

Highlights • Immune parameters are slightly changed upon exposure to plastic fibres or crumb rubber. • Chlorpyrifos caused significant changes in Porcellio scaber immune parameters. • Microplastics decreased the bioavailability of chlorpyrifos for P. scaber. • Mixtures of plastic fibres and chlorpyrifos resulted in greater response in haemocyte count. Microplastics and agrochemicals are common pollutants in terrestrial ecosystems. Their interaction during coexistence in soils may influence their fate and adverse effects on terrestrial organisms. The aim of this study was to investigate how the exposure to two types of microplastics; polyester fibres, and crumb rubber; induce changes in immune parameters of Porcellio scaber and if the co-exposure of microplastics affects the response induced by the organophosphate pesticide chlorpyrifos. A number of immune parameters, such as total haemocyte count, differential haemocyte count, and phenoloxidase-like activity were assessed. In addition, the acetylcholinesterase (AChE) activity in the haemolymph was evaluated as a measure of the bioavailability of chlorpyrifos. After three weeks of exposure, the most noticeable changes in the measured immune parameters and also a significantly reduced AChE activity were seen in chlorpyrifos-exposed animals. Both types of microplastic at environmentally relevant concentrations caused only slight changes in immune parameters which were not dependent on the type of microplastic, although the two types differed significantly in terms of the chemical complexity of the additives. Mixtures of chlorpyrifos and microplastics induced changes that differed from individual exposures. For example, alterations in some measured parameters suggested a reduced bioavailability of chlorpyrifos (AChE activity, haemocyte viability) caused by both types of microplastics exposure, but the increase of haemocyte count was promoted by the presence of fibres implying their joint action. In conclusion, this study suggests that immune processes in P. scaber are slightly changed upon exposure to both types of microplastics and microplastics can significantly modulate the effects of other co-exposed chemicals. Further research is needed on the short-term and long-term joint effects of microplastics and agrochemicals on the immunity of soil invertebrates.

Global climate change affects natural populations of many species by increasing the average temperature and the frequency of extreme weather events (e.g. summer heat waves). The ability of organisms to cope with these environmental changes can, however, depend on their genetic properties. For instance, genetic load owing to inbreeding could alter organisms’ responses to climate change-mediated environmental changes but such effects are often overlooked. We investigated the effects of an experimental heat wave (25°C versus 15°C) on life history (reproduction, size) and constitutive immune defence traits (phenoloxidase-like and antibacterial activity of haemolymph) in relation to inbreeding by manipulating the mating type (outcrossing, self-fertilization) in two populations of a hermaphroditic freshwater snail, Lymnaea stagnalis. High temperature increased reproduction and size of snails but impaired their immune function. In one of the two study populations, inbreeding reduced reproductive output of snails indicating inbreeding depression. Furthermore, this effect did not depend on the temperature snails were exposed to. Our results suggest that L. stagnalis snails can be negatively affected by inbreeding but it may not alter their responses to heat waves. ISSN:1932-6203

Recent studies of honeybees and bumblebees have examined combinatory effects of different stressors, as insect pollinators are naturally exposed to multiple stressors. At the same time the potential influences of simultaneously occurring agricultural agents on insect pollinator health remain largely unknown. Due to different farming methods, and the drift of applied agents and manure, pollinators are most probably exposed to insecticides but also bacteria from organic fertilizers at the same time. We orally exposed honeybee workers to sub-lethal doses of the insecticide thiacloprid and two strains of the bacterium Enterococcus faecalis, which can occur in manure from farming animals. Our results show that under laboratory conditions the bees simultaneously exposed to the a bacterium and the pesticide thiacloprid thiacloprid had significant higher survival rates 11 days post exposure than the controls, which surprisingly showed the lowest survival. Bees that were exposed to diet containing thiacloprid showed decreased food intake. General antibacterial activity is increased by the insecticide and the bacteria, resulting in a higher immune response observed in treated individuals compared to control individuals. We thus propose that caloric restriction through behavioural and physiological adaptations may have mediated an improved survival and stress resistance in our tests. However, the decreased food consumption could in long-term also result in possible negative effects at colony level. Our study does not show an additive negative impact of sub-lethal insecticide and bacteria doses, when tested under laboratory conditions. In contrast, we report seemingly beneficial effects of simultaneous exposure of bees to agricultural agents, which might demonstrate a surprising biological capacity for coping with stressors, possibly through hormetic regulation.

The purpose of the present study was to examine the immediate and prolonged immune response in circulating cytokine and adipocytokine concentrations after two different resistance exercise bouts: hypertrophic (HYP1, 5 × 10, 80% of 1RM) and maximal explosive (POW1, 10 × 5, 60% of 1RM) resistance exercise bouts and how 12 weeks of resistance training (RT) modifies these responses (HYP2, POW2). Eight men completed the study. RE-induced interleukin-6 (IL-6), interleukin-1β (IL-1β), interleukin-1 receptor antagonist (IL-1ra), monocyte-chemoattractant protein-1 (MCP-1), leptin, resistin, and adiponectin were measured before (PRE) and immediately (POST0), 24 (POST24) and 48 (POST48) hours after RE bouts before and after RT. In the untrained state, IL-6 increased immediately after RE in HYP1 (p = 0.002) and in POW1 (p = 0.003) whereas no changes were observed after RT. Similar results were observed in IL-1β, whereas conversely, IL-1ra increased only after RT in HYP2 and POW2 (p < 0.05). Resistin increased before RT in HYP1 and in POW1 (p = 0.011 and p = 0.003, respectively), but after RT, significant responses were not observed. Interestingly, in HYP2, MCP-1 increased significantly at POST24 (p = 0.009) and at POST48 (p = 0.032) only following RT. The present study shows that RT modifies RE-induced cytokine responses towards an anti-inflammatory direction. peerReviewed

The aim of this study was to investigate immune response and its prognostic significance in colon carcinomas using the previously described Immunoscore (IS). A population-based series of 779 colorectal cancers, operated on between 2000 and 2010, were classified according to tumour, node, metastasis (TNM) status, mismatch repair (MMR), and BRAF mutation status. Rectal cancer cases (n = 203) were excluded as a high proportion of these patients received preoperative neoadjuvant chemoradiotherapy. Tissue microarray (TMA) samples collected from the tumour centre and invasive front were immunostained for CD3 and CD8. Lymphocytes were then digitally calculated to categorize IS from grade 0 to 4. Samples adequate for IS were available from 510 tumours. IS was significantly associated with AJCC/UICC stage, T stage, lymph node and distant metastases, perineural and lymphovascular invasion, MMR status, and BRAF mutation status. For IS0, IS1, IS2, IS3 and IS4, respectively, the 5-year disease-free survival (DFS) rates were 59, 68, 78, 83 and 94% (p

The applicability of immune responses in transplanted Baltic blue mussels (Mytilus trossulus) as biomarkers of immunotoxic effects was studied at differently contaminated locations in the Gulf of Bothnia (northern Baltic Sea). Here, we present a detailed report on the immune responses measured as complementary part of transplantation study by Turja et al. (2014).Various immunological endpoints such as total and differential cell count, morphological alterations,phagocytic activity, and caspase 3/7 activity of mussel haemocytes as well as haemolytic activity of the haemolymph were used. Mussels collected at a reference site at a Finnish coastal site (Hanko, H) were transplanted at the Swedish coast near industrial and urban regions of the cities Sundsvall (S1, S2) and Gavle (G1, G2), respectively. Based on the measured immunological responses, multivariate statistical analysis (PCA biplot) showed a clear separation of the most polluted site S1, indicating immunotoxic impacts of the mixture of contaminants present at this location. Based on these observations and results from Turja et al. (2014), we suggest the implementation of immunotoxic biomarkers for the evaluation of ecosystem health. However, these should be accompanied by complementary endpoints of biological effects encompassing i.e., physiological, antioxidant and bioenergetic markers.

HPV-infektiota, joka on yleisin sukupuoliteitse ja muissa limakalvokontakteissa tarttuva oireeton tauti, esiintyy sekä naisilla että miehillä. Korkean riskin HPV tyyppien infektiot, joista HPV16 ja 18 ovat yleisimmät, voivat pitkittyessään aiheuttaa esiasteiden kautta syöpää mm. kohdunkaulan, peräaukon, suun ja nenänielun ja sukupuolielinten alueilla. HPV aiheuttaa maailmanlaajuisesti 10 % naisten ja 5 % miesten kaikista syövistä. Vaikka organisoidulla papa-seulonnalla kohdunkaulasyöpä on Suomessa saatu vähenemään jopa 80 %, HPV-infektiot ovat silti lisääntyneet nuorilla. Maailmalla on havaittu HPV-infektioihin liittyvien peräaukonsyöpien ja suun- ja nenänielunsyöpien esiintymisen nopea lisääntyminen viime vuosikymmenen aikana. Tämän takia infektion esiintymisen vähentäminen rokotuksen avulla on ensisijaisen tärkeää. Väitöstutkimuksen tarkoituksena oli selvittää kaksivaikutteisen HPV16/18-rokotteen synnyttämää immuunivastetta. Väitöstyössä todettiin, että rokote aikaansaa korkeita vasta-ainepitoisuuksia veressä sekä tytöillä että pojilla. Nuorilla 10–14-vuotiailla rokotetuilla tytöillä vasta-ainetasot ovat merkitsevästi korkeammat kuin 15–25-vuotiaiden rokotettujen, saati luonnollisesti HPV16/18-infektion sairastaneiden naisten vasta-ainetasot. Jokaisella tutkittavalla vasta-ainetasot pysyivät korkeina neljän vuoden seurannassa. Tutkimuksessa todettiin, että rokotetuilla nuorilla naisilla, joilla oli korkeat veren vasta-ainepitoisuudet, vasta-aineet siirtyivät aina limakalvoille, kun taas matalammat veren vasta-ainepitoisuudet eivät tuottaneet mitattavissa olevia vasta-aineita limakalvoilla. Vasta-aineiden esiintyminen limakalvoilla antaa tärkeän suojan HPV-infektiota vastaan ja tartuttavuus vähenee. Myös 10–18-vuotiailla pojilla ja nuorilla miehillä vasta-aineet olivat korkeammat kuin 15–25-vuotiailla nuorilla naisilla. Rokotettujen nuorten miesten virtsanäytteissä HPV:n esiintyminen väheni jonkin verran neljä vuotta rokottamisen jälkeen, mikä myös antaa viitteen tartuttavuuden vähenemisestä Työssä rokotettiin 10–14-vuotiaita tyttöjä ja 15–25-vuotiata nuoria naisia HPV16/18-rokotteella (0,1 ja 6 kk) ja tutkittiin rokotteen aikaansaamia HPV16- ja HPV18-vasta-ainetasoja veressä 7, 24, 36 ja 48 kuukautta rokottamisesta. Rokotettujen 10–18-vuotiaden poikien ja nuorten miesten vasta-aineita veressä tutkittiin 7 kuukautta rokottamisesta. Veren ja kohdunkaulan limakalvolle siirtyneiden rokotteen vasta-aineiden tasoja vertailtiin nuorilla naisilla 24, 36 ja 48 kuukautta rokottamisesta. HPV:n esiintymistä vertailtiin HPV16/18-rokotettujen ja rokottamattomien 18–19-vuotiaiden nuorten miesten virtsassa neljä vuotta rokottamisen jälkeen. Tutkimustyö tehtiin Tampereen Yliopiston, Terveystieteen laitoksella rokotevalmistaja GlaxoSmithKlinen rahoittamana tutkimuksena vuosina 2004–2013. Vuonna 2007 myyntiluvan saanut HPV16/18-rokote otettiin Suomessa 12–13-vuotiaiden tyttöjen rokotusohjelmaan tavoitteena vähentää HPV-infektiota ja siten HPV:stä johtuvan kohdunkaulasyövän ja esiasteiden esiintyminen. Nuorten parempi vasta-ainetuotanto puhuu sen puolesta, että rokote kannattaa antaa teini-iässä ennen aktiivisen seksuaalielämän alkamista. HPV-infektio on kuitenkin myös miesten infektio, eikä HPV:n esiintyminen tai tarttuminen vähene, jos vain tytöt rokotetaan. HPV16/18-rokote aikaansaa erinomaisen immuunivasteen myös nuorilla miehillä, jotka toimivat tartuttajina mutta saattavat myös itse sairastua HPV-infektioon. Lauma-immuniteetin aikaansaamiseksi ja HPV:n esiintymisen, tautitaakan ja tartuttavuuden vähentämiseksi tulisi rokottaa sekä tytöt että pojat. The human papillomavirus (HPV) infection is the most common sexually transmitted disease in the world. Most sexually active individuals of both sexes will acquire an HPV infection at some point in their lives. Although most HPV infections clear within two years, approximately 10% persist, possibly resulting in the development of precancerous lesions and cancer. The most common high-risk (hr) HPV types 16 and 18 cause up to 70% of cervical cancers, and all hrHPVs are responsible for 5% and 10% of all cancers worldwide in males and females, respectively. HrHPVs cause up to 100% of cancers in the cervix, 90% in the anus, 35-50% in the oropharynx, 50% in the vulva, 64-91% in the vagina and 40-45% in the penis. Decreasing the transmission of prevalent undetected, asymptomatic, and long-term HPV infections by vaccination is therefore of utmost interest. The licensed vaccines - the bivalent vaccine comprising HPV types 16 and 18 and the quadrivalent vaccine comprising HPV types 6, 11, 16 and 18 are safe and 90-100% efficacious against persistent infections and precancerous cervical lesions associated with HPV types 16 and 18. Both vaccines have also shown 97-100% efficacy against cervical intraepithelial neoplasia (CIN) 2+, CIN3, and adenocarcinoma in situ (AIS) associated with the vaccine types. Both vaccines protect against anogenital infections with the non-vaccine HPV type 31, and the bivalent vaccine additionally against the non-vaccine types 33, 45 and 51. Some vaccine efficacy against oropharyngeal infection has been shown for the bivalent vaccine and against anal HPV infection for both vaccines. The purpose of this thesis is to evaluate the antibody responses following HPV 16/18-vaccination in adolescent girls and boys and young women and men. The antibody responses up to four years following the HPV 16/18 vaccination were evaluated in adolescent girls and young women. Also, antibody levels of HPV 16/18 in serum samples and cervical secretions were compared in young women. The prevalence of HPV deoxyribonucleic acid (DNA) was compared in HPV 16/18-vaccinated and non-vaccinated young men. The University of Tampere, School of Health Sciences, conducted these prospective clinical studies in collaboration with GlaxoSmithKline at several study sites in Finland and abroad during 2004-2013. The immune responses in adolescent girls and boys aged 10-14, and young women and men aged 15 to 25, were excellent, with approximately 100- and 40-fold (HPV 16 and 18) higher antibody titers as compared to those resulting from natural HPV infections. The antibody titers were higher in younger age groups of both genders than in young women. Sustainability of the high antibody levels following vaccination with the HPV 16/18 vaccine was demonstrated in adolescent girls and young women up to four years. The level of mucosal antibodies correlated with the serum antibody levels in young women four years post vaccination. HPV 16/18 antibodies, transudated through the epithelium to mucosal surfaces, provide protection at sites where infection occurs. Vaccination also reduced the prevalence of vaccine HPV types HPV 16 and 18 in males, ensuring reduced transmissibility. High antibody levels will most likely persist for a long time. As HPV infections are usually acquired within three years of sexual debut, the HPV vaccination should be administered prior to this to both girls and boys, to achieve the desired herd immunity with a reduction in the global HPV burden.

Epidemiological studies have shown that short or insufficient sleep is associated with increased risk for metabolic diseases and mortality. To elucidate mechanisms behind this connection, we aimed to identify genes and pathways affected by experimentally induced, partial sleep restriction and to verify their connection to insufficient sleep at population level. The experimental design simulated sleep restriction during a working week: sleep of healthy men (N = 9) was restricted to 4 h/night for five nights. The control subjects (N = 4) spent 8 h/night in bed. Leukocyte RNA expression was analyzed at baseline, after sleep restriction, and after recovery using whole genome microarrays complemented with pathway and transcription factor analysis. Expression levels of the ten most up-regulated and ten most down-regulated transcripts were correlated with subjective assessment of insufficient sleep in a population cohort (N = 472). Experimental sleep restriction altered the expression of 117 genes. Eight of the 25 most up-regulated transcripts were related to immune function. Accordingly, fifteen of the 25 most up-regulated Gene Ontology pathways were also related to immune function, including those for B cell activation, interleukin 8 production, and NF-κB signaling (P

Väitöskirjassa tutkitaan solujen välisessä viestinnässä tärkeän solun sisäisen JAK/STAT-tiedonsiirtoreitin säätelymekanismeja. Sytokiinit ovat elimistömme solujen tuottamia liukoisia välittäjäaineita, jotka säätelevät monia biologisia tapahtumia kuten immuunijärjestelmän solujen jakautumista, erilaistumista ja aktiivisuutta. Sytokiinien vaikutukset välittyvät kohdesolujen pinnalla olevien vastaanottimien eli reseptorien kautta. Sytokiinin sitoutuminen reseptoriinsa aikaansaa solun sisäisten JAK-signalointiproteiinien aktivaation. Aktivoidut JAK-proteiinit fosforyloivat STAT-transkriptiotekijöissä tietyn tyrosiini-aminohappotähteen, jonka seurauksena STAT-proteiinit pariutuvat ja siirtyvät solun tumaan, missä ne sitoutuvat niille spesifisten kohdegeenien säätelyalueille vaikuttaen kyseisten geenien luentaan. JAK/STAT-tiedonsiirtoreitin aktiivisuus on tarkoin säädelty reitin jokaisella tasolla. Säätelyn häiriytyminen ja reitin virheellinen toiminta on yhteydessä moniin sairauksiin, kuten erilaisiin autoimmuunitauteihin ja syöpään. Ymmärtääksemme tarkemmin näiden sairauksien molekyylitason mekanismeja ja kehittääksemme uusia hoitomuotoja näihin sairauksiin, on JAK/STAT-reitin säätelyjärjestelmien yksityiskohtainen tunteminen erittäin tärkeää. JAK/STAT-tiedonsiirtoreitti on säilynyt hyvin muuttumattomana eläinten evoluutiossa ja ihmisen reittiä vastaava tiedonsiirtokaskadi toimii myös banaanikärpäsen (Drosophila melanogaster) soluissa. Geneettisen muunneltavuutensa vuoksi banaanikärpänen on biolääketieteellisessä tutkimuksessa hyvin laajasti hyödynnetty malliorganismi. Väitöskirjatyössä on pyritty selvittämään banaanikärpästä malliorganismina käyttäen uusia JAK/STAT-signalointireitin aktiivisuuteen vaikuttavia molekyylejä. Sumolaatio on yleinen proteiinien translaationjälkeinen muokkaus, jossa kohdeproteiiniin liitetään SUMO-niminen pieni säätelyproteiini. Väitöskirjan ensimmäisessä osatyössä osoitettiin, että banaanikärpäsen ainoa STAT-transkriptiotekijä, Stat92E, on säädelty sumolaation välityksellä. SUMO:n löydettiin liittyvän Stat92E-proteiinissa lysiini 187 aminohappotähteeseen. Lisäksi sumolaation osoitettiin vaikuttavan negatiivisesti Stat92E-transkriptiotekijän aktiivisuuteen. Tutkimus osoittaa, että sumolaatio on STAT-transkriptiotekijöiden evolutiivisesti konservoitunut säätelymekanismi, sillä aiemmin myös ihmisen STAT1-transkriptiotekijän on osoitettu olevan säädelty sumolaation välityksellä. Väitöskirjan kaksi muuta osatyötä pohjautuvat banaanikärpäsen S2-solulinjassa tehtyyn RNA-häirintäseulaan, jonka tarkoituksena on ollut löytää uusia JAK/STAT-reitin aktiivisuuteen vaikuttavia geenituotteita. Seulassa löydetyistä geenituotteista kaksi, Eye transformer ja Not4, valittiin tarkempiin jatkotutkimuksiin. ET-proteiinin osoitettiin toimivan banaanikärpäsen JAK/STAT-reitin negatiivisena säätelijänä, vaikuttaen inhiboivasti Stat92E-proteiinin tyrosiinifosforylaatioon. Banaanikärpäsen Not4-proteiinin löydettiin puolestaan vaikuttavan positiivisesti JAK/STAT-reitin säätelemien geenien ilmentymiseen. Lisäksi Not4-proteiinia vastaavan ihmisproteiinin, CNOT4:n, poisto RNA-häirinnällä ihmisen HeLa-solulinjasta alensi STAT1- ja STAT6 -välitteisten kohdegeenien ilmentymistä. Jatkotutkimukset banaanikärpäsen soluissa osoittivat, että Not4 kykenee sitoutumaan Stat92E proteiiniin ja, että Not4 vaaditaan Stat92E-proteiinin sitoutumiseen tietyn TotM-kohdegeeninsä säätelyalueelle. Tässä tutkimuksessa on löydetty banaanikärpäsen Stat92E-transkriptiotekijän olevan negatiivisesti säädelty sumolaation kautta, osoittaen sumolaation olevan evoluutiossa säilynyt JAK/STAT-reitin säätelymekanismi. Lisäksi tutkimuksessa on karakterisoitu kaksi ennestään tuntematonta JAK/STAT-tiedonsiirtoreitin säätelijämolekyyliä; ET ja Not4/CNOT4. Nämä täysin uudet löydökset luovat hyvän pohjan jatkotutkimuksille immuunijärjestelmän ja sen häiriöiden ymmärtämiseksi. Cytokines are secreted proteins or glycoproteins that are responsible for mediating cell to cell signals and orchestrating numerous biological events including the activation, proliferation and differentation of the cells of the immune system. Cytokine signals are mediated through specific transmembrane receptors on the surface of their target cells. The binding of a hematopoietic cytokine to its receptor triggers the activation of receptor-associated Janus kinases (JAKs). This is followed by the activation of their downstream targets, Signal transducers and activators of transcription (STATs) through tyrosine phosphorylation. Activated STATs dimerize and translocate to the nucleus, where they modulate the transcription of their target genes. The activity of this signaling cascade is under the control of regulatory proteins, such as Protein inhibitors of activated STAT (PIAS), Suppressors of cytokine signaling (SOCS) and various protein tyrosine phosphatases (PTPs). Moreover, JAKs and STATs are regulated by different post-translational modifications, like in the case of STAT1 and STAT5 through the covalent conjugation of a small ubiquitin-like modifier (SUMO). The JAK/STAT pathway has been conserved throughout evolution. In the fruit fly, Drosophila melanogaster, the JAK/STAT cascade plays a crucial role in various developmental events as well as in the immune response, and it is activated by a set of secreted mediators called Unpaired (Upd). In this thesis Drosophila melanogaster was used as a model organism to study the regulation of the JAK/STAT pathway. The only known Drosophila STAT transcription factor, Stat92E, was shown to be modified with SUMO at a single lysine, K187. Mutating this SUMO conjugation site increased the Upd-induced transcriptional activity of Stat92E, suggesting that the sumoylation of Stat92E negatively regulates the Drosophila JAK/STAT pathway. This indicates that sumoylation is an evolutionary conserved regulatory mechanism of STAT-mediated signal transduction. RNA interference (RNAi) has been found to function exceptionally well in Drosophila cells, and also in vivo in adult flies. A genome-wide RNAi screen was conducted in Drosophila S2 cells to reveal novel JAK/STAT pathway associated genes. As a result six potential JAK/STAT pathway regulating genes were found, of which two; Eye transformer (ET) and Not4 were studied in more detail. ET encodes a gp130-related transmembrane protein that was found to inhibit Upd-induced Stat92E tyrosine phosphorylation, thus it functions as a negative regulator of the Drosophila JAK/STAT pathway. RNAi of Not4 was found to repress the Drosophila JAK/STAT pathway target gene expression in Drosophila cells, indicating that Not4 is needed for the JAK/STAT pathway to function properly. Furthermore, overexpression of Not4 resulted in enhanced Stat92E-mediated gene responses, confirming Not4 as a positive regulator of the Drosophila JAK/STAT pathway. We found that Not4 is able to interact with Stat92E, but does not affect Stat92E tyrosine phosphorylation. Finally, our experiments indicated that Not4 is needed for Stat92E to properly bind to its target DNA sequence at the promoter of the stress gene TurandotM (TotM). The Not4 mammalian homologue CNOT4 was also shown to participate in STAT1- and STAT6-mediated gene expression in human cells, indicating that Not4/CNOT4 is an evolutionary conserved regulator of JAK/STAT signaling.

Mutanen, Niina 2009. Valkosolu- ja verihiutalevasteet voimaharjoituksen ja voimaharjoittelun yhteydessä miehillä: erityisesti proteiinin ja ikääntymisen vaikutukset. Valmennus- ja testausopin kandidaatintutkielma. Liikuntabiologian laitos, Jyväskylän yliopisto, 39 s. Fyysisen kuormituksen immunologisia vaikutuksia on tutkittu paljon, mutta tutkimus on keskittynyt lähinnä kestävyystyyppiseen kuormitukseen. Tämän tutkimuksen tarkoituksena oli selvittää valkosolu- ja verihiutalevasteita voimaharjoituksen ja voimaharjoittelun yhteydessä miehillä. Erityisesti tutkittiin voimaharjoituksen yhteydessä nautitun heraproteiinin akuutteja vaikutuksia sekä ikääntymisen vaikutusta immuniteettireaktioihin.Tutkimuksessa käytettiin kolmen eri koeasetelman tuloksia. Ensimmäisessä koeasetelmassa koehenkilöinä oli kymmenen voimaharjoittelua harrastanutta nuorta miestä (23,9 ± 2,0 v). Jokainen koehenkilö kävi läpi kaksi koetilannetta: toisella kerralla he nauttivat 30 minuuttia ennen voimaharjoitusta herakaseinaattijuoman (25 g) ja toisella kerralla energiattoman plasebon. Voimaharjoitus oli raskas kontrastivoimaharjoitus jaloille. Molemmissa tilanteissa koehenkilöt nauttivat proteiinia ja hiilihydraattia sisältävän palautusjuoman välittömästi voimaharjoituksen jälkeen. Toiseen koeasetelmaan sisältyi kahdenlaisia akuuttitilanteita. Ensimmäinen koehenkilöryhmä koostui voimaharjoitteluun tottumattomista nuorista miehistä, jotka satunnaistettiin proteiini- (n=11, 25,0 ± 4,8 v), plasebo- (n=11, 27,6 ± 2,8 v) ja kontrolliryhmiin (n=8, 25.2 ± 3.4 v) ensimmäisen testikerran jälkeen. Toisessa tilanteessa koehenkilöinä oli harjoitelleita vanhoja miehiä, jotka jaettiin 21 viikon voimaharjoittelujakson jälkeen proteiini (n=9, 61,4 ± 4,3 v)- ja plaseboryhmiin (n=9, 62,l ± 4,2 v) osittaisella satunnaistamisella ja matsaamalla . Proteiiniryhmä nautti ennen ja jälkeen raskaan voimaharjoituksen heraproteiinijuoman (15 g) ja plaseboryhmä tilavuudeltaan saman määrän energiatonta plaseboa. Kolmannessa asetelmassa selvitettiin 21 viikkoa kestäneen voimaharjoittelujakson vaikutusta perustason valkosolu- ja verihiutalemääriin vanhoilla (n=18, 61,2 ± 4,1 v) ja nuorilla (n= 19, 26,4 ± 4,4 v) miehillä. Kontrolliryhmä koostui sekä nuorista että vanhoista miehistä (n=19, 42,5 ± 20,0 v). Voimaharjoittelujakson aikana koehenkilöt suorittivat kaksi ohjattua voimaharjoitusta viikossa. Kaikissa koeasetelmissa verinäytteistä analysoitiin verihiutaleet ja valkosolut, jotka eriteltiin tarkemmin kolmeen fraktioon. Tilastollisina menetelminä käytettiin keskiarvoa, keskihajontaa sekä ttestiä. Koeasetelmassa 1 lymfosyyttien lukumäärä nousi raskaan voimaharjoituksen seurauksena laskien kuormituksen jälkeen alemmaksi kuin ennen kuormitusta. Verihiutaleiden määrä nousi plasebotilanteessa pre J1- tilanteeseen (pre J1= proteiini- tai plasebojuoman nauttimista edeltävä verinäyte) verrattuna. Neutrofiilien määrä nousi kuormituksen aikana, lähti laskuun kuormituksen loputtua, mutta alkoi nousta jälleen 30 minuutin kohdalla. Verihiutaleiden määrä oli plaseboryhmällä koholla viisi minuuttia kuormituksen loputtua (p

Väitöstutkimuksessa selvitettiin eri laboratoriokokeiden ja niiden yhdistelmien käyttökelpoisuutta alkoholin suurkulutuksen tunnistamisessa sekä alkoholinkäytön aiheuttamia muutoksia immuunivasteessa. Tutkimuksessa kehitettyjen määritysmenetelmien ja erilaisten laboratoriokokeiden yhdistelmillä voitiin parantaa alkoholin suurkuluttajien tunnistamista. Tutkimuksessa havaittiin myös, että alkoholin aiheuttamille proteiinimuunnoksille spesifisten vasta-aineiden ja immuunivastetta säätelevien sytokiinien määrät muuttuvat verenkierrossa alkoholinkäytön seurauksena ja tämä saattaa vaikuttaa myös alkoholimaksasairauden muodostumiseen ja etenemiseen. Tutkimuksessa yhdistettiin kaksi alkoholinkäytön tunnistamisessa yleisesti käytettyä laboratoriokoetta, gammaglutamyylitransferaasi (GGT) ja desialotransferriini (CDT) ja tämän yhdistelmän todettiin toimivan alkoholistien tunnistamisessa paremmin kuin minkään yksittäisen laboratoriokokeen. Lisäksi tutkimuksessa kehitettiin menetelmä, jolla voidaan määrittää vasta-aineita asetaldehydiaddukteille, jotka muodostuvat, kun etanolin aineenvaihduntatuote asetaldehydi reagoi elimistön proteiinien kanssa. IgA-luokan vasta-aineiden määrän todettiin olevan suurempi alkoholisteilla kuin kohtuujuojilla tai täysraittiilla ja ero vasta-aineiden määrässä oli merkitsevä myös kohtuujuojien ja täysraittiiden välillä. Tutkimuksen perusteella tällaisten vasta-aineiden määritys voisi olla käyttökelpoinen myös alkoholin suurkulutuksen tunnistamisessa ja tunnistamiseen saataisiin edelleen lisää parannusta yhdistämällä tämän uuden menetelmän tulokset CDT:n kanssa (IgA-CDT). Luotettavien laboratoriokokeiden löytäminen olisi tärkeää alkoholin suurkuluttajien mahdollisimman aikaisen tunnistamisen kannalta. Nykyään liian moni alkoholin suurkuluttaja jää tunnistamatta terveydenhuollossa, mikä voi johtaa viivästyksiin hoidossa. Mitä aikaisemmin alkoholin suurkuluttaja pystytään ohjaamaan hoitoon, sitä parempi ennuste hänellä on toipumiseen. Alkoholin suurkulutus ja siitä aiheutuvat terveydelliset ongelmat kuormittavat merkittävästi terveydenhuoltoa. Vuonna 2005 alkoholisyyt nousivat työikäisten miesten suurimmaksi kuolinsyyksi ja naisillakin alkoholisyyt ovat toiseksi yleisin kuolinsyy heti rintasyövän jälkeen. Lääketieteellisten ja sosiaalisten ongelmien lisäksi alkoholin suurkulutus on muodostunut kansantaloudelliseksi rasitteeksi. On arvioitu, että alkoholin käyttö aiheutti yhteiskunnalle 640-850 miljoonan euron välittömät kustannukset vuonna 2005 ja ne tuskin laskevat lähivuosina. Excessive alcohol consumption and consequent medical disorders create a major burden for modern health care. In addition to medical and social problems, excessive drinking causes considerable strains on the national economy. In order to improve the diagnosis and treatment of patients suffering from ethanol-related health problems, reliable and accurate methods for recognizing excessive alcohol consumption in its early phase need to be developed. Currently, excessive drinkers tend to escape detection, which may lead to delays in intervention. The present study was set out to develop new approaches for detecting excessive drinking based on conventional and new laboratory tests and their combinations and to address the relationships between such markers and alcoholic liver disease. Conventional laboratory markers of excessive drinking (GGT, CDT, MCV, AST, ALT), a mathematically formulated combination of GGT and CDT, and autoimmune responses to proteins modified with acetaldehyde, the first metabolite of ethanol, were measured in alcoholics with or without liver disease, moderate drinkers and abstainers. Cytokine profiles of subjects were also studied in order to clarify the associations between alcohol intake and pathogenesis of alcoholic liver disease. The results show that even moderate drinking may increase levels of gamma-glutamyl transferase (GGT). When GGT was combined logarithmically with carbohydrate-deficient transferrin (CDT), the diagnostic performance of the combination GGT-CDT was found to markedly exceed that of the traditional markers, reaching a sensitivity of 90 % whereas the sensitivities of its parent components remained at 63 % (CDT) and 58 % (GGT). Alcohol consumption was also found to induce alterations in the immune system. An association between cytokine levels and alcohol use was observed, with most evident alterations in alcoholics with liver disease. Pro-inflammatory cytokines (IL-2, IL-6, IL-8, TNF-α) were found to increase in alcoholic liver disease whereas the levels of anti-inflammatory cytokines, particularly TGF-β1 showed a slight decrease. Acetaldehyde adducts are formed when acetaldehyde reacts with proteins and cellular constituents. Antibodies directed against acetaldehyde-modified proteins were found in the circulation of alcoholic patients. The highest anti-adduct IgA and IgG titres occurred in patients with alcoholic liver disease, while specific class IgM antibodies were most abundant in alcoholics without liver disease. The possible usefulness of anti-adduct IgA as a marker of excessive drinking was subsequently studied in alcoholics without liver disease, and the mean anti-adduct IgA levels were significantly higher than those in the moderate drinkers or abstainers (p In the light of these results, it may be suggested that GGT, CDT and IgAs against acetaldehyde-derived epitopes either alone or combined by means of mathematical equations can serve as useful markers of alcohol consumption. The elevation of GGT in moderate drinkers as compared with abstainers may affect its reference ranges, since they are usually calculated from general population in which the proportion of abstainers is decreasing. The results also indicate that the antibody responses and alterations in cytokine balance may contribute to the pathogenesis and progression of alcoholic liver disease in humans.

Väitöstutkimuksessa selvitettiin alkoholin suurkulutuksen aiheuttamia immuunivasteita ja niiden yhteyttä alkoholisairauksien syntyyn, erityisesti maksafibroosiin. Tutkimuksessa havaittiin, että alkoholin suurkuluttajilla esiintyy immuunivasteita, jotka kohdistuvat alkoholin aineenvaihduntatuotteisiin. Näillä immuunivasteilla saattaa olla sekä diagnostisia että patogeenisiä sovellutuksia alkoholin aiheuttamien sairauksien tutkimisessa. Tulokset viittaavat myös siihen, että maksasairailla alkoholisteilla sekä pro-inflammatoristen ja anti-inflammatoristen sytokiinien välinen tasapaino että kollageenin synteesin ja hajoamisen välinen tasapaino ovat häiriintyneet. Nämä tulokset tarjoavat uusia mahdollisuuksia tunnistaa ne ihmiset, joilla on suurentunut riski sairastua fibroosiin ja kirroosiin alkoholinkäytön seurauksena. Tutkimuksessa määritettiin perinteisten laboratoriokokeiden (MCV, GGT, CDT, AST, ALT) lisäksi autoim-muunivasteita asetaldehydin muokkaamia proteiineja vastaan, sytokiiniprofiileja, fibrogeneesin osoittimia (PIIINP, PINP ja hyaluronihappo) sekä kollageenin hajoamista kuvaavia markkereita (β-CrossLaps ja ICTP). Tutkimuksessa oli mukana maksasairaita alkoholisteja, alkoholisteja, joilla ei ole maksasairautta sekä alkoho-lin kohtuukäyttäjiä ja täysraittiita. Maksasairailla alkoholisteilla IgA-tasot sekä asetaldehydiaddukteja että kudostransglutaminaasia (tTG) (joka perinteisesti on totuttu liittämään keliakiaan) vastaan olivat merkittävästi korkeammat kuin terveillä kontrolleilla tai alkoholisteilla, joilla ei ole maksasairautta. Näiden kahden vasta-aineen kohoamisella havaittiin olevan myös merkittävä keskinäinen yhteys. Sidekudoksen hajoamista kuvaavat kokeet olivat maksasairailla alkoholisteilla matalammat kuin terveillä kontrolleilla tai alkoholisteilla, joilla ei ole maksasairautta. Sen sijaan sidekudoksen synteesiä kuvaavat kokeet olivat alkoholimaksasairailla merkittävästi korkeammat. Hajoamisen ja synteesin osoittimista lasketulla suhteella saavutettiin myös erinomainen herkkyys (94 %) ja tarkkuus (98 %) maksasairaiden alkoholistien tunnistamisessa. Suomessa vuonna 2005 alkoholisyyt nousivat työikäisten miesten yleisimmäksi kuolinsyyksi ja naisten toiseksi yleisimmäksi heti rintasyövän jälkeen. Maksakirroosin esiintyvyys maassamme on lisääntynyt lyhyessä ajassa yli 50 %:lla. Although excessive ethanol consumption is a significant cause of health problems in most Western countries, the primary mechanisms of ethanol-induced diseases have remained largely unknown. There has also been a lack of specific methods for assessing ethanol-induced health problems. This study sets out to explore the associations and diagnostic usefulness of immune responses induced by ethanol metabolites, biomarkers of ethanol con-sumption, cytokines and fibrogenesis in alcohol consumers. Conventional labora-tory markers of excessive ethanol consumption (MCV, GGT, CDT, AST, ALT), autoimmune responses to proteins modified by acetaldehyde (the first metabolite of ethanol), cytokine profiles and markers of fibrogenesis (carboxyterminal propeptides of type I and type III procollagens, PINP and PIIINP and hyaluronic acid), and markers of collagen degradation (aminoterminal telopeptides of type I collagen, β-CrossLaps and ICTP) were measured in alcoholics with or without liver disease, moderate drinkers and abstainers. Anti-acetaldehyde adduct IgA levels were significantly higher in the alcohol-ics than in the moderate drinkers or abstainers. The values in the moderate drink-ers were also higher than those in the abstainers. Anti-acetaldehyde adduct IgAs correlated significantly with the amount of ethanol consumed during the month preceding blood sampling (r = 0.67, p The patients with alcoholic liver disease (ALD) had significantly higher levels of IgAs, not only against acetaldehyde adducts but also against tissue transgluta-minase (tTG), than did the healthy controls or alcoholics without liver disease. There was also a significant co-occurrence between these two antibodies in the alcohol consumers. Anti-tTG IgA showed significant positive correlations with proinflammatory cytokines (IL-2, IL-6, IL-8, TNF-α) and negative correlations with anti-inflammatory cytokines (IL-1, TGF-β). Highly positive correlations oc-curred between anti-tTG IgA and markers of fibrogenesis (PIIINP, hyaluronic acid). Serum β-CrossLaps (β-CTx) in ALD patients were at a significantly lower level than in the healthy controls or the alcoholics without liver disease, while PINP and PIIINP, reflecting type I and type III collagen synthesis, respectively, were significantly increased. The alcoholics without liver disease showed values which were not significantly different from those of the healthy controls. Serum β-CTx correlated negatively with serum PIIINP and pro-inflammatory cytokines (IL-2, IL-6, IL-8, TNF-α), and positively with anti-inflammatory cytokines (IL-1, TGF-β), whereas serum PIIINP correlated positively with pro-inflammatory cyto-kines and negatively with anti-inflammatory cytokines. Calculation of the PIIINP/β-CTx ratio was found to yield an excellent sensitivity (94 %) and speci-ficity (98 %) in differentiating alcoholics with liver disease. The present data suggest that anti-acetaldehyde adduct IgAs may provide a new clinically useful marker of excessive ethanol consumption. Specific immune responses to ethanol metabolites may prove to have both diagnostic and patho-genic implications in ethanol-related diseases. The close association between anti-tTG and anti-acetaldehyde adduct IgAs may indicate that, under conditions in which the integrity of the intestinal barriers has been disrupted upon ethanol in-take and acetaldehyde exposure, there may be common mucosal responses to such antigens. The results also suggest that patients with alcoholic liver disease have a disturbed balance between pro-inflammatory and anti-inflammatory cytokines and between collagen synthesis and degradation, which may facilitate fibrogenesis in ALD. These findings also have implications for the development of non-invasive tools for discriminating individuals at risk for fibrogenesis.