Abstract Colorectal cancer (CRC) is among the most common cancers in Finland. Prognostic factors are important for predicting disease outcome and adjusting optimal treatment. The currently used prognostic methods for CRC have their limitations and consequently several biomarkers have been studied to find potential prognostic markers, but none have been adapted for routine use so far. In the present study, the relationships between the components of the immune system and other factors modulating tumor growth were assessed and their suitability to be used for use as prognostic tools in CRC were studied. The study material consisted of blood samples and surgical specimens collected from 148 CRC patients operated on in Oulu University Hospital and blood samples of 86 healthy controls. Concentrations of endostatin and 27 cytokines were measured from preoperative serum samples and control samples. Immunohistochemical methods were used for collagen XVIII and inflammatory cell analyses. The levels of several cytokines were altered in CRC patients compared to the controls. The serum cytokine profile achieved an excellent accuracy in discriminating CRC patients from healthy controls. Advanced CRCs were associated with elevated cytokine levels and a metastasized disease was linked to an orientation towards Th2 cytokine milieu. The presence of systemic inflammation, depicted by a modified Glasgow Prognostic Score (mGPS), correlated to CRC progression. The serum endostatin levels were elevated in CRC and correlated with invasion through muscular layer and systemic inflammation, but not with densities of local inflammatory cells. Collagen XVIII was expressed in tumor stroma and in the muscle layer of bowel wall. The serum cytokines and tumor infiltrating immune cells showed relatively weak associations. In conclusion, CRC is associated with significant alterations in serum cytokine milieu, which underlines the relevance of studying several cytokines and their relative alterations. The serum cytokine profile is a promising tool for discriminating CRC patients from healthy controls, but its clinical value needs to be validated. The elevated endostatin levels may result from invasion-related cleavage of collagen XVIII in the bowel wall, but further studies are needed to determine the value of endostatin in CRC prognosis. / Tiivistelmä Paksu- ja peräsuolisyöpä (kolorektaalisyöpä) on yleisimpiä syöpämuotoja Suomessa. Sen ennustetta kuvaavat mittarit ovat tärkeitä taudin etenemisen ennustamisessa ja hoidon suunnittelussa. Käytössä olevat kolorektaalisyövän ennusteen arvioinnin menetelmät eivät ole riittäviä. Uusia merkkiaineita onkin kehitetty ja testattu, mutta rutiinikäyttöön soveltuvia menetelmiä ei ole vielä löydetty. Tässä tutkimuksessa selvitettiin immuunijärjestelmän ja muiden kasvaimen kasvua säätelevien tekijöiden keskinäisiä yhteyksiä ja niiden merkitystä kolorektaalisyövän ennusteen arvioinnissa. Tutkimusmateriaali koostui Oulun yliopistollisessa sairaalassa leikattujen kolorektaalisyöpäpotilaiden (n = 148) leikkaus- ja verinäytteistä ja terveiden verrokkihenkilöiden (n = 86) verinäytteistä. Endostatiinin ja 27 sytokiinin pitoisuudet mitattiin seeruminäytteistä. Kollageeni XVIII:n ja tulehdussolujen analysoimiseen käytettiin immunohistokemiallisia menetelmiä. Useiden sytokiinien pitoisuudet olivat korkeammat potilailla kuin verrokeilla, mutta osassa sytokiineista pitoisuudet olivat alentuneet. Seerumin sytokiiniprofiili erotteli luotettavasti potilaat verrokeista. Pidemmälle edenneeseen tautiin liittyi sytokiinien korkeampia pitoisuuksia ja etäpesäkkeitä muodostanut tauti oli yhteydessä Th1-tyypin sytokiinien esiintymiseen. Systeeminen tulehdusreaktio oli yhteydessä syövän etenemiseen. Endostatiinipitoisuudet olivat kohonneet potilailla ja olivat yhteydessä kasvaimen invaasioon suolen seinämän lihaskerroksen läpi. Endostatiinipitoisuudet korreloivat myös systeemisen tulehdusreaktion kanssa, mutta eivät liittyneet paikallisten tulehdussolujen määrään. Kollageeni XVIII ilmentyi kasvaimen stroomassa ja suolen seinämän lihaskerroksessa. Sytokiineilla ja kasvaimen paikallisilla tulehdussoluilla todettiin olevan vain vähän keskinäisiä yhteyksiä. Kolorektaalisyöpään liittyy useita erisuuntaisia muutoksia seerumin sytokiinipitoisuuksissa, joten on olennaista tutkia eri sytokiinien suhteellisia muutoksia. Seerumin sytokiiniprofiili on lupaava potilaita ja verrokeita erotteleva mittari, jolla voi olla diagnostista arvoa. Kohonneet endostatiinipitoisuudet potilailla voivat johtua kasvaimen invaasioon liittyvästä kollageeni XVIII:n hajoamisesta suolen seinämässä, mutta lisätutkimuksia tarvitaan endostatiinin ennustetta kuvaavan arvon määrittämiseksi.

Teblad innehåller en mängd substanser som på olika sätt kan påverka biologiska processer i människan. I föreliggande arbete har en sammanställning gjorts utifrån vetenskapliga artiklar över teets antagna effekter på människans immunsystem .Främst har den mest studerade komponenten i te, epigallocatechin gallate, EGCG, undersökts. De variabler som studerats i immunförsvaret är neutrofiler, tumörnekrosfaktor-α, interferon-γ, interleukin-1β, interleukin-2, interleukin-5, interleukin-6, interleukin-8, interleukin-10 ,interleukin-12, interleukin-13, eikosanoidmetabolismen, lymfocyter, makrofager, dendritiskaceller och T-cellsdifferentiering. Sammafattningsvis föreföll te hämma följande: Neutrofilaktiviteten, TNF-α-bildningen, IFN-γ-aktiviteten, stimuli-inducerad IL-β-bildning, IL-8-aktivering och IL-10-bildning. EGCG visades öka dendritiska cellers apoptos och ändra dedenderitiska cellernas morfologi och minska deras förmüga att stimulera T-cellproliferationen. EGCG-behandling av celler ledde till ökad makrofagaktivitet, ökad produktion av T-regceller kontra andra T-cellslinjer. Tebehandling ökade bildningen av IL-12 ochIL-13. EGCGs effekt på IL-6 var tvetydig; enligt en del artiklar påverkade EGCG ej IL-6, medan det enligt andra artiklar antingen ökade eller minskade IL-6-bildningen. Trots att resultaten ej var entydiga kunde slutsatsen att EGCG (och därmed te) utövade immunsupprimerande effekt dras men att det sannolikt krävs betydligt högre koncentrationer än de som är fysiologiskt uppnåbara (1 μM i plasma och 3 mM i tarmen) genom reguljärt tedrickande för att få en avgörande effekt på människans immunsystem. EGCGs hämmnade effekt på immunsystemet kan dock vara av intresse i högre – farmakologiska – koncentrationer genom att EGCG skulle kunna nyttjas som komplement eller alternativ till för närvarande nyttjade immunförtryckande läkemedel som ciklosporin och glukokortikoider.

Dans le cadre des recherches menées au sein du laboratoire de Physiologie Animale le gène codant pour la nicotinamide phosphorybosyltransférase (NAmPT) à été identifié et cloné. Au cours de ce travail, nous avons étudié le rôle de cette enzyme du métabolisme du Nicotinamide Adénine Dinucléotide ainsi que celui des enzymes dépendantes du NAD (PARP et sirtuines) dans le contrôle de la réponse inflammatoire. / Doctorat en Sciences / info:eu-repo/semantics/nonPublished

SUMMARY<p><p>Type 1 diabetes mellitus (T1DM) is an autoimmune disease caused by progressive destruction of insulin-producing pancreatic beta-cells. Both viral infections and the cytokines interleukin-1beta (IL-1beta) and interferon-gamma (IFN-gamma) have been suggested as potential mediators of beta-cell death in early T1DM. Nitric oxide (NO) is a highly diffusible, short-lived free radical gas, which plays a significant role in several physiological processes in a diversity of tissues and organisms. Prolonged exposure of rodent or human pancreatic beta-cells to combinations of cytokines induces the expression of the inducible form of nitric oxide synthase (iNOS) and Fas, NO production, and cell death. It also induces the expression of potential "defense" genes, such as manganese superoxide dismutase (MnSOD) and heat shock protein (hsp) 70. Recent studies have shown that NO, in addition to having cytotoxic actions, may also regulate gene transcription. It remains unclear whether NO mediates cytokine-induced gene expression and subsequent beta-cell death. Previous studies using NO synthase blockers yielded conflicting results, which may be due to non-specific effects of these agents. <p>In the first part of our work, we examined the role of NO in beta-cell dysfunction and death by using an iNOS knockout mice (iNOS-/-, background C57BL/6x129SvEv). We evaluated the effects of cytokines on gene expression, as determined by reverse transcriptase-polymerase chain reaction (RT-PCR), and viability, as determined by nuclear dyes, of pancreatic islet cells or fluorescence-activated cell sorter (FACS)-purified beta-cells isolated from iNOS knockout mice or their respective controls (C57BL/6x129SvEv). The combination of cytokines used was interleukin-1beta (50 U/ml) + gamma-interferon (1000 U/ml) + tumor necrosis factor-alpha (1000 U/ml). The lack of cytokine-induced iNOS activity in the iNOS-/- islet cells was confirmed by RT-PCR and nitrite determination. Cytokines induced a > 3-fold increase in Fas and MnSOD mRNA expression in wild-type (wt) and iNOS-/- islets. On the other hand, hsp 70 was induced in wt but not in iNOS-/- islets. Prolonged (6-9 days) exposure of wt islets to cytokines lead to an 80-90% decrease in islet cell viability, whereas viability decreased by only 10-30% in iNOS-/- islet cells. To determine the mode of cytokine-induced cell death, FACS-purified beta-cells were exposed to the same cytokines. After 9 days, the apoptosis index was similarly increased in wt (39 +/- 3%) and iNOS-/- (33 +/- 4 %) beta-cells. On the other hand, cytokines increased necrosis in wt (20 +/- 4 %) but not in iNOS-/- (7 +/- 3 %) beta-cells. From these data, we conclude that: 1) NO is required for cytokine-induced hsp 70 mRNA expression, but not for Fas and MnSOD expression; 2) cytokines induce both apoptosis and necrosis in mouse beta-cells; 3) cytokine-induced apoptosis is mostly NO-independent, whereas necrosis requires NO formation.<p>In the second part of our work, we examined the role of the viral product double-stranded RNA (dsRNA) in beta-cell dysfunction and death. DsRNA is produced by many viruses during their replicative cycle. We investigated whether dsRNA (here utilized as synthetic poly IC (PIC)) modifies the effects of IL-1beta and IFN-gamma on gene expression and viability of rat pancreatic beta-cells and the role of NO in this process. FACS-purified rat beta-cells were exposed for 6-16 h (study of gene expression by RT-PCR) or 6-9 days (study of viability by nuclear dyes) to PIC and/or IL-1beta or IFN-gamma. PIC increased the expression of Fas and Mn superoxide dismutase mRNAs by 5-10-fold. IL-1beta and a combination of PIC + IFN-gamma& / Doctorat en sciences biomédicales / info:eu-repo/semantics/nonPublished

The aim of cancer immunotherapy is to eradicate tumours by inducing a tumour-specific immune response. This thesis focuses on how antibodies and peptides can improve antigen presentation and the subsequent tumour-specific T cell response. Tumour recognition by the immune system can be promoted through delivery of antigen in the form of a vaccine. One example is the development of a therapeutic peptide vaccine containing both CD4+ and CD8+ T cell epitopes. So far, peptide vaccinations have shown limited success in clinical trials and further improvements are needed, such as choice of adjuvant and T cell epitopes, as well as targeted delivery of peptides and adjuvants to the same DC. In paper I, we describe the development of a peptide-peptide conjugate (with a tumour T cell epitope) that, via immune complex formation and FcγR binding, enhance antigen uptake and activation of DCs. The conjugate consists of three tetanus toxin-derived linear B cell epitopes (MTTE) that were identified based on specific IgG antibodies in human serum. Three MTTE peptide sequences were conjugated to a synthetic long peptide (SLP) that consists of a T cell epitope derived from the desired target tumour. In paper II, the conjugate was evaluated in a modified Chandler loop model containing human blood, mimicking blood in circulation. The conjugate was internalised by human monocytes in an antibody-dependent manner. A conjugate containing the model CMV-derived T cell epitope pp65NLV generated recall T cell responses dependent on MTTE-specific antibodies and the covalent conjugation of the three MTTE with the SLP. In paper III, a CD40-specific antibody was characterised for local treatment of solid tumours. The antibody eradicated bladder tumours in mice and induced T cell-mediated immunological memory against the tumour. In paper IV, we characterised the Chandler loop model (used in paper II) for its potential use in predicting cytokine release syndrome (CRS) in response to monoclonal antibodies (mAbs). Superagonistic antibodies (e.g., OKT3) induced rapid cytokine release whereas no cytokine release was induced by antibodies (e.g., cetuximab) associated with low incidence of CRS in the clinic. In conclusion, this thesis work demonstrates proof-of-concept of improved strategies for antibody- and peptides-based cancer immunotherapies and their potential use in multiple cancer indications.

Abstract It is a generally accepted dogma that sepsis disturbs skin function and wound healing, but surprisingly there is only remote pathophysiological evidence available behind that presumption. As the skin is the largest defensive barrier, the skin dysfunction in severe sepsis deserves more attention. In this study, the suction blister model was used to create experimental wounds. The study population included 44 patients with severe sepsis and 15 controls. The blister fluid was collected to analyse cytokine profile of the skin. The transepidermal water loss and blood flow from the wound were measured. A 4mm biopsy was taken under local anaesthesia on the first and the eighth day of the study from the healthy looking skin. Then 15 healing suction blisters were excised. Serum samples were also collected on the first day of the study. The barrier restoration was diminished, and the inflammation in the wound was more intense in severe sepsis than in the controls. The expression of the basement membrane components Laminin-332 and type IV collagen decreased during the septic disease, but increased over the next 3 months without achieving the level oft he controls. The expression of tight junction proteins remained nearly intact in the healing wound in severe sepsis compared to the controls. The expression of occludin on the leading edge of the migrating keratinocytes was more restricted and late in severe sepsis compared to the controls. The levels of the tumour necrosis factor (TNF), interleukin-10 (IL-10) and IL-6 in skin blister fluid were higher in the sepsis compared to controls. The blister fluid and serum cytokine response in the sepsis differed since the levels of epidermal growth factor, vascular endothelial growth factor, TNF and basic fibroblastic growth factor (bFGF) in the blister fluid did not correlate with the levels of serum. The septic patients with multiple organ failure had higher levels of several cytokines than patients without organ failure. Survivors had lower levels of IL-10 and bFGF in blister fluid than the non-survivors. This study offers novel findings for skin and wound healing in sepsis. Together, all the findings suggest that skin dysfunction in severe sepsis exists even when the most profound structures remain intact. Understanding these mechanisms of impaired wound healing can improve future treatments, such as the timing of surgery. / Tiivistelmä Sepsiksen ajatellaan heikentävän haavanparanemista, mutta tieteellistä näyttöä on niukasti. Iholla on keskeinen osa elimistön puolustuksessa ja tasapainon ylläpidossa, joten sen toiminnan häiriintyminen systeemisessä tulehduksessa ansaitsee suuremman huomion. Imurakkulahaavat tehtiin 44 septiselle potilaalle ja 15 kontrollille. Haavoista mitattiin veden haihtumista ja veren virtausta sekä otettiin imurakkulaneste näytteeksi sytokiinimäärityksiä varten. Tutkimuksen ensimmäisenä ja kahdeksantena päivänä otettiin 4mm biopsiat terveeltä iholta ja 15 potilaalta poistettiin näytteeksi paraneva imurakkulahaava. Seeruminäytteet otettiin tutkimuksen ensimmäisenä päivänä. Veden haihtuminen haavalta oli voimakkaampaa eli ihon barrierin palautuminen oli hidastunut septisillä potilailla verrattuna kontrolleihin. Haavassa havaittu tulehdus oli sepsiksessä voimakkaampi. Tyvikalvon komponenttien Laminiini-332:n ja tyypin IV kollageenin ilmentyminen oli vähäisempää sepsiksen aikana ja lisääntyi 3kk kohdalla, mutta ei kuitenkaan saavuttanut kontrollien tasoa. Tiivisliitosproteiinien ilmentyminen oli lähes muuttumatonta sepsiksessä kontrolleihin verrattuna. Okludiinin ilmentyminen sen sijaan paranevassa haavassa vaeltavien keratinosyyttien etureunassa oli rajoittuneempaa ja myöhäisempää sepsiksessä kuin kontrolleilla. Sytokiineistä tuumorinekroositekijä (TNF), interleukiini-10 (IL-10) ja IL-6 olivat koholla imurakkulanesteessä verrattuna kontrolleihin. Epidermaalinen kasvutekijä, verisuonten endoteelikasvutekijä, TNF ja perusfibroplastinen kasvutekijä (bFGF) pitoisuudet rakkulanesteessä erosivat seerumin pitoisuuksista eli ihon sytokiiniprofiili erosi systeemisestä sytokiiniprofiilista. Potilailla, joilla oli monielinvaurio, todettiin korkeampia sytokiinipitoisuuksia. Potilailla, jotka menehtyivät 30 vrk kuluessa, oli korkeammat pitoisuudet IL-10 ja bFGF rakkulanesteessä. Tämä tutkimus tarjoaa uutta tietoa ihosta ja haavanparanemisesta sepiksessä. Tulosten perusteella voidaan todeta, että ihon toimintahäiriö on sepsiksessä todellinen, vaikka kaikkein perustavimmat rakenteet säilyvät muuttumattomina. Toimintahäiriön mekanismien ymmärtäminen voisi auttaa septisen potilaan hoidossa, kuten kirurgisten toimenpiteiden ajoittamisessa paranemisen kannalta mahdollisimman otolliseen aikaan.

Abstract Prognostic features of colorectal cancer (CRC) are important for determining the optimal treatment for an individual patient. This study was carried out to evaluate the significance of the prognostic significance of inflammatory cell reaction and tumour budding at the invasive front of the tumour in CRC patients, to study the pattern of alterations in the serum cytokine levels of CRC patients compared to healthy controls, and to evaluate whether the patterns of the cytokine levels alter according to the stage of disease. Study material consists of a series of CRC patients operated in Oulu University Hospital (N=466, studies I-II, and N=148, study III). The intensities of inflammatory cell reaction and tumour budding were estimated and the association of these features with survival was analysed (I-II). Preoperative serum samples were collected from patients and age- and sex-matched controls, and concentrations of 13 cytokines and chemokines were analysed (III). Inflammatory cell reaction and tumour budding at the tumour invasive margin were independent prognostic markers in CRC. In patients with stage I-II disease, high-grade inflammation was associated with better 5-year survival (87.6%) than low-grade inflammation (47.0%). Tumour budding was present in 24.0% of cases and predicted worse 5-year survival (15.4% in contrast to 63.5%). Serum levels of PDGF, IL-6, IL-7, and IL-8 were higher, and levels of MCP-1 were lower in CRC patients compared to controls. A pattern of five most predictive cytokines reached an excellent capacity in discriminating patients from healthy controls and reached an AUC of 0.890 in the ROC analysis. High-stage CRC was associated with increased levels of IL-6, IL-8 and IL-1ra, and metastasised disease was accompanied by an orientation to Th2 cytokine milieu. According to this study, patients with CRC can be stratified into different prognostic groups by assessing inflammatory cell reaction and tumour budding at the invasive front of the tumour. Evaluation of these features may give additional information for making treatment decisions. Serum cytokine profile was shown to change during cancer progression and seems promising in separating CRC patients from healthy controls, but its clinical value is yet to be confirmed. / Tiivistelmä Kasvaimen ennusteeseen vaikuttavien tekijöiden tunteminen on tärkeää syöpäpotilaan yksilöllisen hoidon suunnittelussa. Tässä tutkimuksessa selvitettiin kasvaimen tulehdusreaktion ja kasvaimen reunan silmuilevan kasvutavan merkitystä kolorektaalisyöpäpotilaiden ennusteeseen. Lisäksi tutkimuksessa mitattiin tulehdusreaktion välittäjäaineiden, seerumin sytokiinien, pitoisuuksia potilailla ja verrokeilla sekä näiden pitoisuuksien vaihtelua suhteessa syövän levinneisyyteen. Tutkimusaineisto koostui Oulun yliopistollisessa sairaalassa leikatuista kolorektaalisyöpäpotilaista (N=466, tutkimukset I–II, N=148, tutkimus III). Kasvainnäytteistä tutkittiin kasvaimen tulehdusreaktion ja silmuilevan kasvutavan voimakkuutta ja arvioitiin näitä piirteitä suhteessa potilaiden elinaikatietoihin (tutkimukset I–II). Potilaiden ja verrokkihenkilöiden seeruminäytteistä mitattiin 13 sytokiinin pitoisuudet (tutkimus III). Kasvaimen tulehdusreaktio ja silmuileva kasvutapa osoittautuivat itsenäisiksi ennustetekijöiksi. Potilailla oli parempi 5-vuotisennuste (87.6 %), jos kasvaimen tulehdusreaktio oli voimakas verrattuna tapauksiin, joissa tulehdusreaktio oli heikko (47.0 %). 24 %:ssa kasvaimista kasvutapa oli silmuileva, ja näillä 5-vuotisennuste oli huonompi kuin ei-silmuilevissa (15.4 % vs. 63.5 %). Potilailla todettiin korkeammat seerumin PDGF, IL-6, IL-7, ja IL-8 -pitoisuudet ja matalammat MCP-1 -pitoisuudet kuin verrokeilla. Mittaamalla viiden ennustearvoltaan merkitsevimmän sytokiinin pitoisuudet voitiin potilaat luotettavasti erottaa verrokeista ROC-analyysin avulla, kun ROC-käyrän alle jäävä pinta-ala oli 0.890 %. Pidemmälle levinneissä taudeissa todettiin korkeampia IL-6, IL-8 ja IL-1ra -pitoisuuksia, ja etäpesäkkeisissä taudeissa sytokiinipitoisuudet muuttuivat Th2 -profiilin suuntaiseksi. Tutkimuksen mukaan kasvaimen tulehdusreaktion ja silmuilevan kasvutavan arvioinnilla kolorektaalisyöpäpotilaat voidaan jakaa ennusteellisiin ryhmiin, mitä on mahdollista hyödyntää hoidon suunnittelussa. Seerumin sytokiiniprofiilin osoitettiin muuttuvan taudin edetessä, ja sytokiinien pitoisuudet poikkeavat kolorektaalisyöpäpotilailla verrattuna terveiltä verrokeilta mitattuihin arvoihin.

Doctor of Philosophy / Department of Anatomy and Physiology / Michael J. Kenney / Important bidirectional interactions exist between the central nervous system and the immune system. Neural-immune interactions provide a regulatory system in the body and disturbances in these interactions may lead to disease. Although the sympathetic nervous system is thought to play a key role in mediating neural-immune interactions, central neural mechanisms mediating sympathetic-immune interactions and the effect of centrally-induced alterations in sympathetic nerve discharge on immune function is not known. We tested the hypothesis that central activation of sympathetic neural circuits alters splenic cytokine gene expression. In a separate study, we tested the hypothesis that hypothermia-induced changes in visceral sympathetic nerve discharge (SND) would be attenuated in middle-aged and aged compared with young rats. Previous studies have demonstrated that skin sympathoexcitatory responses to skin cooling are attenuated in aged compared with young subjects, suggesting that advancing age influences sympathetic nerve responsiveness to cooling. The effect of age on sympathetic nerves innervating other targets organs during acute cooling remains unknown. Central activation of splenic SND was produced using three different experimental interventions: increased core body temperature produced by acute heating, intracerebroventricular injection of angiotensin II (ANGII), and decreased core body temperature produced by acute cooling. Changes in gene expression profiles were analyzed using inflammatory cytokine-specific gene-array and further validated using real-time RT-PCR analysis. The following observations were made. 1)Splenic SNDincreased in response to each experimental intervention except in acute cooled young rats where there was a decrease in splenic SND. 2) Splenic cytokine gene expression of pro-inflammatory cytokines (e.g., IL-1β, IL-6, IL-2) and chemokines (GRO1, CXCL2, CCCL2 and, CXCL10) was increased in response to each experimental intervention. 3) Expression of splenic cytokine genes was reduced after splenic-denervation except in acute cooled rats. 4) Progressive hypothermia reduced splenic, renal, and adrenal SND in rats and was generally attenuated in middle-aged and aged rats. These results demonstrate the functional significance of changes in sympathetic nerve activity on splenic immune cell activation and the effect of age on SND responses to core body cooling.

IL-12 family cytokines IL-23 and IL-27 play an important role linking innate and adaptive immunity, and regulating T-cell responses. The production of IL-12, a structurally similar cytokine, is decreased in chronic HIV infection; therefore IL-23 and IL-27 may also be influenced by HIV infection. I hypothesized that HIV inhibits LPS-induced IL-23 and IL-27 production in human MDMs by suppressing the activation of signalling pathways regulating their expression. In vitro HIV-infection of MDMs did not have any effect on basal secretion of IL-23 or IL-27; however, HIV inhibited LPS-induced production of IL-12/23 p40 and IL-23 p19, and IL-27 EBI3 and IL-27 p28 mRNA expression, and IL-23, IL-12/23 p40 and IL-27 secretion. In order to evaluate the molecular mechanisms by which HIV inhibits IL-23 and IL-27 in LPS-stimulated MDMs, the signalling pathways regulating their expression were evaluated. The PI3K, p38 MAPK, and JNK MAPK pathways were found to positively regulate LPS-induced IL-27 secretion. Interestingly, in vitro HIV infection inhibited LPS-induced p38 and JNK MAPK activation in MDMs. In summary, I have shown that HIV inhibits IL-23 and IL-27 production in LPS-stimulated MDMs and that HIV may inhibit LPS-induced IL-27 production through the inhibition of p38 and JNK MAPK activation. It is currently unknown whether PKCs regulate LPS-induced IL-23 or IL-27 in human monocytes/macrophages. I demonstrated that classical PKCs differentially regulate LPS-induced IL-23 and IL-27 secretion within THP-1 cells, primary monocytes, and MDMs. Classical PKCs were found to positively regulate LPS-induced IL-12/23 p40 and IL-27 p28 mRNA expression and IL-12/23 p40, IL-23, and IL-27 secretion in primary human monocytes. Similarly, the classical PKCs were found to positively regulate IL-27 p28 mRNA expression and IL-27 secretion in THP-1 cells. However, classical PKCs did not regulate LPS-induced IL-27 production in MDMs, or LPS-induced IL-23 production in THP-1 cells. Overall, this demonstrates that classical PKCs differentially regulate LPS-induced IL-23 and IL-27 production in different myeloid cells.

No description available.