Standardized whole-blood transcriptional profiling enables the deconvolution of complex induced immune responses

Figura como autor también el Milieu Intérieur ConsortiumSystems approaches for the study of immune signaling pathways have been traditionally based on purified cells or cultured lines. However, in vivo responses involve the coordinated action of multiple cell types, which interact to establish an inflammatory microenvironment. We employed standardized whole-blood stimulation systems to test the hypothesis that responses to Toll-like receptor ligands or whole microbes can be defined by the transcriptional signatures of key cytokines. We found 44 genes, identified using Support Vector Machine learning, that captured the diversity of complex innate immune responses with improved segregation between distinct stimuli. Furthermore, we used donor variability to identify shared inter-cellular pathways and trace cytokine loops involved in gene expression. This provides strategies for dimension reduction of large datasets and deconvolution of innate immune responses applicable for characterizing immunomodulatory molecules. Moreover, we provide an interactive R-Shiny application with healthy donor reference values for induced inflammatory genes

Standardized Whole-Blood Transcriptional Profiling Enables the Deconvolution of Complex Induced Immune Responses

SummarySystems approaches for the study of immune signaling pathways have been traditionally based on purified cells or cultured lines. However, in vivo responses involve the coordinated action of multiple cell types, which interact to establish an inflammatory microenvironment. We employed standardized whole-blood stimulation systems to test the hypothesis that responses to Toll-like receptor ligands or whole microbes can be defined by the transcriptional signatures of key cytokines. We found 44 genes, identified using Support Vector Machine learning, that captured the diversity of complex innate immune responses with improved segregation between distinct stimuli. Furthermore, we used donor variability to identify shared inter-cellular pathways and trace cytokine loops involved in gene expression. This provides strategies for dimension reduction of large datasets and deconvolution of innate immune responses applicable for characterizing immunomodulatory molecules. Moreover, we provide an interactive R-Shiny application with healthy donor reference values for induced inflammatory genes

Interaction of TLR-3 stimulated retinal pigment epithelium and retinal microglia in view of the age-related macular degeneration

Die altersabhängige Makuladegeneration (AMD) ist Hauptursache für schwere Sehstörungen in der westlichen Welt. Im Fokus dieser Arbeit steht die noch unklare Interaktion TLR-3 aktivierten retinalen Pigmentepithels (RPE) und retinaler Mikroglia,die für die Pathogenese der AMD und viraler Infektionen mitverantwortlich gemacht wird.Das Expressionsmuster ausgewählter Zytokine und Enzyme wurde mittels real-time RT-PCR offengelegt.Das RPE als Teil der Blut-Retina-Schranke fungiert als Schnittstelle zwischen der Aderhaut und der immunprivilegierten Retina.Die retinale Mikroglia ist die gewebeständige Abwehrzelle.Der TLR-3 ist Teil des angeborenen Immunsystems.Mit viraler doppelsträngiger RNA (dsRNA) als Ligand dient er der Virusabwehr. Wir etablierten eine Zellkultur retinaler Mikroglia. Zum Mikroglianachweis dienten Immunfärbungen mit Iba-1.Unsere Experimente zeigen, dass durch polyinosinic polycitidylic acid (Poly I:C – synthetische dsRNA) TLR-3 stimulierte retinale Mikroglia mit einer konzentrationsabhängigen Induktion der proinflammatorischen Zytokine IL-1β, IL-6,TNF-α und der Enzyme COX-2,iNOS reagiert. Die Interaktion TLR-3 stimulierten RPEs und retinaler Mikroglia ergab eine Expressionssteigerung von IL-1β, IL-6 und COX-2 gegenüber der mit identischer Poly I:C-Konzentration behandelten Kontrollgruppe retinaler Mikroglia. Wir detektierten eine Reduktion der Expression für iNOS, sowie ein erhöhtes Expressionsniveau für das antiinflammatorische Zytokin IL-10. Wir beobachteten eine Induktion von COX-2 und IL-1β bei gleichzeitiger Reduktion der Expression von iNOS– entsprechend einer Entzündungsreaktion mit gleichzeitiger Limitation der Neurotoxizität. Eine Aktivierung des RPEs könnte einen feed-forward Mechanismus vermittelt durch lösliche Fakoren, wie Zytokine auslösen,die retinale Mikroglia aktivieren und die beobachtete Immunantwort induzieren, deren Modulation ein Schritt vorwärts in der Therapie der AMD, aber auch akuter Prozesse, wie viraler Infektionen darstellt.Age-related macular degeneration (AMD) is the main reason for severe visual impairment in the western world.The focus of this exploration was on the then to be defined interaction of TLR-3 stimulated retinal pigment epithelium (RPE) and retinal microglia which appears to be an immanent element of AMD and viral infections.A cluster of cytokines and enzymes was tested in real-time RT-PCR in order to map the reactions.The RPE works as an interface between the choroid and the immune privileged retina.Retinal microglia is the resident immune cell of the retina.The toll-like 3 receptor (TLR-3) is part of the innate immune system with viral double stranded RNA (dsRNA) as its ligand.We established a cell culture of retinal microglia.For cell identification immunocytochemical staining with Iba-1 was conducted.Our experiments show that retinal microglia that has been stimulated with polyinosinic polycitidylic acid(Poly I:C-synthetic dsRNA)via TLR-3 reacts with a concentration dependent induction of the proinflammatory cytokines IL-1β, IL-6, TNF-α and of the enzymes COX-2 and iNOS.With focus on the interaction of TLR-3 stimulated RPE and retinal microglia we found an increase in the expression of IL-1β, IL-6, COX-2 and in basal stimulation,of the antiinflammtory IL-10 in retinal microglia that has been treated with supernatant of TLR-3 stimulated RPE in comparison to the control group of retinal microglia that had solely been treated with equal concentrations of Poly I:C.We observed an induction of the expression of COX-2 and IL-1β accompanied by a reduction of iNOS correlating to an inflammatory reaction chaperoned by a limitation of neurotoxicity.An activation of the RPE might induce a feed-forward mechanism via soluble factors like cytokines from RPE to retinal microglia resulting in the observed immune response.A modulation of the immune reaction of RPE and retinal microglia appears to be a step ahead in the therapy of AMD but also of acute processes like viral infections

Mechanismen der Aktivierung des NLRP3 Inflammasoms bei chronischer Nierenerkrankung und kardiovaskulären Erkrankungen

Eine sterile Inflammation spielt eine wichtige Rolle bei der Entstehung und Progression einer chronischen Nierenerkrankung als auch bei kardiovaskulären Erkrankungen. In Vorarbeiten konnte unsere Arbeitsgruppe zeigen, dass die Lipoproteine HDL und LDL auf unterschiedliche Weise das angeborene Immunsystem wie beispielsweise Toll-like Rezeptor-2 (TLR2) aktivieren und so eine systemische Inflammation induzieren. Das NLRP3 Inflammasom stellt einen weiteren wichtigen Bestandteil des angeborenen Immunsystems dar, welcher pro-IL-1 in dessen aktive Form prozessiert. Ziel dieser Arbeit war es zu untersuchen, welche Rolle die verschiedenen Lipoproteinklassen bei der Aktivierung des NLRP3 Inflammasoms spielen. In humanen Monozyten führte VLDL aber nicht HDL oder LDL zur Freisetzung von IL-1. Wir konnten nachweisen, dass die Hauptproteinkomponente in VLDL Apolipoprotein C3 hierfür verantwortlich ist. ApoC3 führt auf der Oberfläche humaner Monozyten zur Heterotrimerisierung von TLR2 und TLR4 zusammen mit dem Adapterprotein SCIMP. Dies aktiviert die Kinase Syk, führt zum Calcium-Einstrom in die Zelle über den TRPM2 Kanal und induziert die NADPH Oxidase-abhängige Produktion reaktiver Sauerstoffspezies (ROS). Diese aktivieren Caspase-8, was zur Zusammenlagerung des NLRP3 Inflammasomkomplexes aus seinen Komponenten NLRP3, ASC und Caspase-1 führt. Das aktive NLRP3 Inflammasom prozessiert in der Folge pro- IL-1 in matures IL-1, welches von den Monozyten sezerniert wird und eine systemische proinflammatorische Antwort auslöst. Wir konnten nachweisen, dass dieser Signalweg der alternativen NLRP3 Aktivierung auf das humane System beschränkt ist. Zur Überprüfung der pathophysiologischen Relevanz dieser experimentellen Befunde wurden humanisierte Mäuse generiert. D.h. in immuninkompentente NOD-SCID Mäuse wurden humane CD14+ Monozyten transplantiert. Im murinen Carotis-Schädigungsmodell führte die Injektion von ApoC3 in die humanisierten Mäuse zu einer deutlichen Hemmung der endothelialen Regeneration. Nach unilateraler Ureterligatur förderte ApoC3 die renale Schädigung. ApoC3 Plasmakonzentrationen sind bei Patienten mit chronischer Nierenerkrankung als auch bei Patienten nach akutem Myocardinfarkt im Vergleich zu gesunden Probanden signifikant erhöht und auch mit einer höheren Mortalität assoziiert. Somit konnte in dieser Arbeit erstmals gezeigt werden, dass Triglycerid-reiche Lipoproteine via ApoC3 das NLRP3 Inflammasom aktivieren. Gleichzeitig konnte die alternative NLRP3 Aktivierung als ein pathophysiologisch relevanter Signalweg aufgeklärt werden. Diese Ergebnisse liefern somit wichtige Erkenntnisse über die Regulation des NLRP3 Inflammasoms generell als auch über die Mechanismen einer kardiorenalen Schädigung. ApoC3 und das NLRP3 Inflammasom stellen somit ein neues therapeutisches Target dar.Sterile inflammation represents a hallmark in the initiation and progression of chronic kidney disease as well as cardoiovascular diseases. In our previous, our group could show that the lipoproteins HDL and LDL activate the innate immune system via several distinct pathways such as toll-like receptor-2 (TLR2), which leads to systemic inflammation. The NLRP3 inflammasome represents another important component of the innate immune system by processing pro-IL-1 into its biologically active form. The aim of the present thesis was to assess the effects of the different lipoprotein classes on the activation of the NLRP3 inflammasome. In human monocytes, VLDL but not HDL or LDL induced the release of IL-1. We found that the major protein constituent of VLDL ApoC3 was responsible for this effect. On the surface of human monocytes, ApoC3 triggers hetertrimerization of TLR2, TLR4, together with the adapter protein SCIMP. This activates Syk kinase, leading to calcium influx via TRPM2, which induces NADPH oxidase-dependent production of reactive oxygen species (ROS). ROS activate caspase-8, which facilitates the assembly of the functional NLRP3 inflammasome complex from its components NLRP3, ASC, and caspase-1. The active NLRP3 inflammasome then processes pro-IL-1 into mature IL-1, which is released from human monocytes and triggers a systemic pro-inflammatory response. We found that this pathway of alterative NLRP3 activation is restricted the human system. To prove the pathophysiological relevance of these experimental findings, humanized mice were generated. Therefore, immune incompetent NOD-SCID mice were transplanted with human CD14+ monocytes. In the murine carotid injury model in humanized mice, injection of ApoC3 substantially suppressed endothelial regeneration. After unilateral ureter ligation, ApoC3 promoted kidney injury. ApoC3 plasma levels were significantly higher in patients after acute myocardial infarction or chronic kidney disease as compared to healthy control subjects. Moreover, higher ApoC3 plasma concentrations were associated with higher mortality during follow-up. In summary, this work provides first evidence that triglyceride-rich lipoproteins via ApoC3 activate the NLRP3 inflammasome. Moreover, we could describe alternative NLRP3 activation as a pathophysiologically relevant pathway. These results provide novel insights in the regulation of NLRP3 inflammasome in general as well as in the mechanisms of cardiorenal injury. Therefore, ApoC3 and the NLRP3 inflammasome represent a novel therapeutic target

Serum amyloid A (SAA) : Proinflammatory functions and their regulation by serum lipoproteins

The immune response is operated by two integrated systems, the adaptive and innate immune responses. Innate immunity includes both cellular and soluble components. The cellular part consists of host cells at the front line of defence - macrophages, monocytes, dendritic cells, neutrophils, endothelial cells and mast cells - that express receptors capable of recognizing common pathogen constituents, hence called pattern-recognition receptors, PRRs. Several cooperating PRR families, for example Toll-like receptors (TLRs) and receptors with nucleotide-binding domain leucine-rich repeats (NLRs), have been identified. They recognize two different classes of structures, pathogen-associated molecular patterns (PAMPs) and non-microbial, danger-associated molecular patterns (DAMPs). The soluble component of the innate immune system includes an arsenal of acute-phase proteins, the expression of which is induced during the acute-phase response (APR), an immediate systemic reaction triggered by a local or systemic abnormal condition such as tissue injury, infection or trauma. In addition, the innate immune response is driven by numerous proinflammatory cytokines and mediators, most notably interleukin (IL-) 1β. The activity of IL-1β is tightly controlled; the induction of gene expression and the activation of pro-IL-1β require separate stimuli. IL-1β maturation takes place in cytosolic protein platforms called inflammasomes, of which NLRP3 is the most characterized. The major acute-phase proteins in human are C-reactive protein (CRP) and serum amyloid A (SAA). In response to inflammatory stimulus, SAA concentration in plasma can increase up to 1000-fold. SAA circulates in association with high-density lipoprotein (HDL) and is, thus, suggested to play a role in lipid metabolism and transport. In addition, SAA possesses strong cytokine-like and proinflammatory properties. A pathogenic role for SAA has most clearly been implicated in AA amyloidosis, a systemic protein misfolding disease that can complicate chronic inflammatory conditions. Current evidence indicates that SAA is also as an active mediator in cardiovascular diseases. The aim of the study was to elucidate the interaction between SAA and two types of innate immune system cells, human mast cells and macrophages, and the consequences of this interaction in the pathogenesis of AA amyloidosis and atherosclerosis, as well as the regulation of SAA in inflammation. It was demonstrated that SAA is a potent activator of mast cells and macrophages, as indicated by a dose-dependent production of key proinflammatory cytokines, IL-1β and tumor necrosis factor (TNF) -α, in both cell types. In mast cells, this activation led to the degradation of SAA by the mast cell-derived protease tryptase and to the formation of amyloid-like structures, suggesting a pathogenic role for mast cells in AA amyloidosis. The secretion of IL-1β was studied in more detail in human macrophages, in which SAA was found to be able to induce both the gene expression of IL1B, via TLR2 and TLR4, and the activation of the NLRP3 inflammasome, resulting in the secretion of mature IL-1β. The activation of NLRP3 involved the ATP-receptor P2X7 and cathepsin B activity. Native serum lipoproteins were shown to inhibit the activity of SAA and this inhibition was further enhanced by lipoprotein oxidation. Besides the expression of IL1B, oxidized LDL inhibited also the activation of the NLRP3 inflammasome. A decrease in the SAA-induced IL-1β production was observed also in vivo, suggesting that oxidized LDL, although possessing many pathological features, may represent a novel and significant regulator of SAA activity in inflamed tissues, including atherosclerotic lesions. All together, the findings of this study stress the significance of SAA in the pathogenesis of inflammatory diseases, such as atherosclerosis, and provide new insights into mechanisms leading to AA amyloidogenesis.Immuunivaste voidaan jakaa kahteen integroituneeseen järjestelmään, luontaiseen ja hankittuun immuunivasteeseen. Luontaiseen immuunijärjestelmään kuuluu sekä solu- että liukoisia komponentteja. Näistä ensimmäiseen lukeutuvat immuunipuolustuksen etulinjassa olevat solut - makrofagit, monosyytit, dendriittisolut, neutrofiilit, endoteelisolut ja syöttösolut - sekä näiden erityiset reseptorit, nk. hahmontunnistusreseptorit (engl. pattern-recognition receptors, PRR). Useita yhteistyössä toimivia PRR-perheitä tunnetaan, esimerkiksi TLR-reseptorit ja NLR-reseptorit. Nämä tunnistavat niin taudinaiheuttajaperäisiä kuin muihin elimistön hälytystiloihin tai vaurioihin liittyviä, ei-mikrobisia molekyylirakenteita. Luontaisen immuunijärjestelmän liukoinen osa käsittää lukuisia nk. akuutin vaiheen proteiineja, joiden tuotto indusoituu akuutin vaiheen reaktion aikana. Akuutin vaiheen reaktio on paikallisen tai systeemisen epänormaalin tilan, kuten kudosvaurion, infektion tai trauman, laukaisema välitön systeeminen reaktio, jota voidaan kutsua myös systeemiseksi tulehdukseksi. Luontaista immuunivastetta ohjaavat lisäksi lukuisat sytokiinit ja muut välittäjäaineet, erityisesti interleukiini-1β (IL-1β). Tulehdusta voimakkaasti edistävän IL-1β:n aktiivisuus on tiukan säätelyn alaisuudessa; IL1B-geenin tuotto ja IL-1β-proteiinin esiasteen aktivoituminen vaativat soluissa erilliset ärsykkeet. Näistä jälkimmäinen, esiasteen aktivoituminen, tapahtuu solunsisäisissä proteiinikomplekseissa, inflammasomeissa, joista NLRP3 on tunnetuin ja tutkituin. Merkittävimmät akuutin vaiheen proteiinit ihmisessä ovat C-reaktiivinen proteiini ja seerumin amyloidi A, SAA (engl. serum amyloid A). SAA:n pitoisuus veressä voi tulehduksellisen ärsykkeen vasteena nousta jopa 1000-kertaiseksi normaaliin verrattuna. SAA kulkee verenkierrossa HDL:n mukana ja sillä on tämän vuoksi arveltu olevan lipidien metaboliaan ja kuljetukseen liittyviä tehtäviä. SAA:lla on lisäksi sytokiinien kaltaisia ja tulehdusta edistäviä ominaisuuksia. SAA:n patogeenisyys on osoitettu selkeästi sekundäärisessä AA-tyypin amyloidoosissa, joka on pitkittyneisiin tulehdustiloihin liittyvä systeeminen, proteiinien väärinlaskostumisesta johtuva tauti. Nykytiedon valossa SAA näyttää toimivan myös aktiivisena välittäjämolekyylinä sydän- ja verisuonitaudeissa. Tämän tutkimuksen tavoitteena oli selvittää SAA:n sekä kahden luontaiseen immuunijärjestelmään kuuluvan solutyypin, syöttösolun ja makrofagin, välistä vuorovaikutusta sekä tämän seurauksia AA-amyloidoosin ja ateroskleroosin kannalta. Myös SAA:n aktiivisuuden säätelyä tulehduksen aikana tutkittiin. SAA aiheutti tulehdusta edistävien sytokiinien erityksen sekä syöttösoluista että makrofageista annosriippuvaisella tavalla viitaten siihen, että SAA toimii molempien solutyyppien tehokkaana aktivaattorina. Syöttösoluissa aktivaatio johti SAA:n itsensä pilkkoutumiseen syöttösolulähtöisen tryptaasi-proteaasin toimesta ja edelleen amyloidin kaltaisten rakenteiden muodostumiseen. Tämä viittaa siihen, että syöttösolut saattavat olla osallisina AA-amyloidoosin patogeneesissä. Sytokiinieritystä tutkittiin tarkemmin makrofageissa, joissa havaitsimme SAA:n lisäävän sekä IL1B-geenin tuottoa että NLRP3-inflammasomin aktivaatiota johtaen aktiivisen IL-1β-proteiinin eritykseen. Osoitimme myös, että seerumin lipoproteiinit, HDL ja LDL, estivät SAA:n aktiivisuutta ja että tämä estovaikutus tehostui lipoproteiinien hapettumisen myötä. SAA-välitteisen IL-1β-erityksen väheneminen todennettiin myös steriilin vatsakalvontulehduksen hiirimallissa. On siis mahdollista, että sinällään monia patologisia piirteitä omaava hapettunut LDL saattaa edustaa uutta ja merkittävää SAA:n aktiivisuuden säätelijää tulehtuneissa kudoksissa, mukaan lukien ateroskleroottiset leesiot. Yhteenvetona, tämän tutkimuksen tulokset korostavat SAA:n merkitystä kroonisten tulehdustautien, kuten ateroskleroosin, patogeneesissä sekä tuovat lisävaloa AA-amyloidoosin tautimekanismeihin

The control of inflammation in airway epithelial cells

COPD is a severe chronic and complex airway disease that represents a major financial burden on the healthcare and economic system. Environmental risk factors such as cigarette smoke have been associated with the predisposition to COPD. Other factors such as exposure to viral pathogens can exacerbate airway inflammation and tissue destruction generated by recruited neutrophils, culminating in altered epithelial cell responses in COPD. This thesis investigated the physiological role of Pellino-1, an E3-ubiquitin ligase, and its regulation in human primary bronchial epithelial cells (HBEpCs) in response to viral infection. The viral mimic poly(I:C) increased Pellino-1 protein and gene expression in HBEpCs. In addition, Pellino-1 gene expression was significantly increased by RV-16 and RV-1B infection in primary bronchial epithelial cells from COPD patients. Pellino-1 knockdown in HBEpCs led to a reduction in NF-κB regulated cytokines CXCL8, IL-1α and β gene expression and release of CXCL8 in response to poly(I:C) while having no measurable effect on IFNβ mRNA expression. \ud Furthermore, the transient knockdown of Pellino-1 resulted in the decrease in IKKα/β phosphorylation. The role for Pellino-1 in the non-canonical NF-κB pathway was also investigated and while Pellino-1 knockdown did not alter the expression of the non-canonical NF-κB precursor protein NFKB1 following poly(I:C) stimulation, NFKB2 protein expression was suppressed. In contrast to Pellino-1 knockdown, the transient knockdown of NFKB2 resulted in significant increase in CXCL8 mRNA and protein expression and in turn did not regulate Pellino-1 mRNA expression to poly(I:C). These data suggest that following viral infection in airway epithelial cells, TLR3 activation culminates in the up-regulation of Pellino-1 leading to an increase in NFKB2 expression, resulting in the suppression of NF-κB specific gene transcription. In addition to activating non-canonical NF-κB, NFKB1 regulates the activation of ERK signalling via MEK1. Treatment of HBEpCs with MEK1 inhibitors, PD98059 and U0126, resulted in a significant reduction in Pellino-1 protein and gene expression which led to the suggestion of ERK as a potential Pellino-1 regulator. Proteomics analysis of primary epithelial cells obtained from COPD patient airways further identified a potential novel mechanism of action for Pellino-1 in the NF-κB signalling pathway, wherein it Pellino-1 may inhibit A20’s negative regulatory role or its adaptor proteins TNIP1 or TAX1BP. Taken in combination these data support Pellino-1 as a potential target to down-regulate neutrophilic inflammation whilst retaining antiviral immunity by selectively mediating the TLR3 TRIF-dependent NF-κB/MAPK pathway and not TLR3-mediated IRF3 and IFNβ activation

Mechanisms of pulmonary inflammation in ageing and chronic lung disease

Ph. D. Thesis.The human respiratory tract is exposed to copious antigen over the course of the lifespan by virtue of its free communication with the external environment through the process of ventilation. The host immune system must therefore distinguish innocuous inhaled antigen in the respiratory tract from antigen potentially associated with infection, produce an inflammatory response with minimal collateral host damage if required, and allow a return to homeostasis once infection is cleared. Inadequacies of these processes can result in a predisposition to respiratory illnesses. Respiratory diseases are leading causes of morbidity and mortality worldwide. It is notable that the burden of respiratory disease disproportionately falls upon older adults, and disorders of inflammation arising with ageing may contribute to this disparity. The work in this thesis describes a viable platform for the assessment of pulmonary inflammation that could be adapted to facilitate experimental medicine studies characterising inflammation in advanced age or early phase trials of immunomodulatory drugs that might alter the course and resolution of inflammation. This thesis also describes a method to identify candidate immunomodulatory drugs using connectivity maps and puts forward Del-1 as a target for drugs that enhance the resolution of inflammation. In considering the role of inflammation in chronic lung disease, this thesis also presents an exploration of the mechanisms of inflammation in chronic hypersensitivity pneumonitis, using a systems biology approach to implicate tissue-resident memory T lymphocytes in the pathogenesis of the disease. This opens up the possibility of IL-15 signalling as a potential target for treatment of the disease where few treatments are currently proven to be effective.NIHR Newcastle BRC, the MRC SHIELD AMR Research Consortium and the Medical Research Foundation National PhD Training Programme in Antimicrobial Resistance Researc

Type 2 diabetes in Sri Lanka : Genetic epidemiology and periodontal association.

Prevalence rates of type 2 diabetes and impaired fasting glyceamia (IFG) in Sri Lanka are high and an increasing number of people are succumbing to disease. Identifying people at risk of developing complications is a healthcare priority of the county to prevent morbidity and mortality. Type 2 diabetes is familiarly aggregated and maternal influence for disease transmission has been observed in European countries but not in South India. The severity of periodontitis is reported to be high when diabetes control decreases but data relating to periodontitis as a complication of diabetes is not present in this population. Periodontal associated genotype (PAG), presence of allele 2 of both {ILIA (+4845) & IL1B (+3954)}) is positively associated with periodontal disease in some populations but ruled out in other populations. Heightened levels of TNF-? level have been observed in both patients with type 2 diabetes and periodontitis.This study aimed to determine familial aggregation of type 2 diabetes and parental influence of disease transmission in the Sri Lankan population. The project also aimed to investigate periodontal status in diabetic patients and the effect of glycaemic control on periodontal status. The final aim of the project was to investigate the role of cytokine polymorphisms PAG and TNFA (-308) in association with periodontitis in people with type 2 diabetes.Family history data for one thousand patients was collected and analyzed. Subjects with established diabetes were recruited to the periodontal study (n=285) with an age and sex matched control population (n=72). All subjects underwent both periodontal and general health examination. Their periodontal parameters and metabolic parameters were measured including blood pressure, TG and LDL values. Patients were genotyped by PCR/RFLP method.The results of the study indicated that 59.4% of the diabetic subjects had at least one affected first degree relative in the family. It was also observed that 15.6% of mothers transfer the disease compared to (12.5%) of fathers (p 4mm, PD >5mm and maximum loss of attachment (LOA) scores than the non diabetic population (p0.05). Subjects with periodontitis were significantly older than those with gingivitis or periodontally healthy (p0.05). The percentage of PAG (12.5%) is low compared to Caucasians but higher compared to Chinese population. TNFA (-308) allele distribution had no significant effect on periodontal status. TNFA (-308) genotypes were significantly correlated with HDL.The results of the present study indicate that type 2 diabetes is familiarly aggregated in the Sri Lankan diabetic population and maternal excess is observed in the transmission pattern. There is a trend towards periodontital status to be higher in the diabetic population. Severity of periodontitis is higher in people with diabetes than those without and there is an indication that periodontitis may induce hyperglycaemia. PAG distribution shows population variation and the TNFA (-308) polymorphism may be associated with diabetes

Mechanisms involved in ulceration of the stomach and small bowel

Available from British Library Document Supply Centre-DSC:DXN017049 / BLDSC - British Library Document Supply CentreSIGLEGBUnited Kingdo

Impact of cationic host defence peptide LL-37 on human neutrophil death and inflammatory responses

Cathelicidins are cationic host defence peptides (CHDP) with essential roles in the innate defence system. These peptides have antimicrobial potential and the capacity to modulate innate immunity and inflammatory processes. Neutrophils (PMN) are the main reservoir of cathelicidins and play key roles in first line defence against infection. The appropriate regulation of PMN function, death, and clearance is critical to innate immunity, and the efferocytosis of apoptotic PMN, in contrast to necrotic cells, is proposed to promote the resolution of inflammation. In this thesis I demonstrate that the human cathelicidin LL-37 rapidly induced secondary necrosis of apoptotic human PMN and identify the essential C-terminal region of LL-37 required for this activity. In addition to the induction of secondary necrosis, higher concentrations of LL-37 also promoted PMN granule contents release. LL-37-induced secondary necrosis did not affect PMN ingestion by human monocyte-derived macrophages and, in contrast to expectation, was not proinflammatory. Interestingly, the anti-inflammatory effects of apoptotic PMN on activated macrophages were retained and even potentiated where LL-37-mediated secondary necrosis induced anti-inflammatory granule content release. Consistent with the results of in vitro studies, in vivo murine sterile peritonitis model revealed the same phenomenon: LL-37-induced secondary necrosis diminished inflammatory responses with decreased PMN influx. I also present data on LL-37- mediated modulation of innate immune effector cell cytokines responses to inflammatory signals. I propose that during acute inflammation LL-37 can modulate innate immune responses through its activity on cytokine production, and that LL-37-mediated secondary necrosis of apoptotic PMN has anti-inflammatory effects, but may also mediate host damage by promoting the release of potentially harmful intracellular contents under chronic or dysregulated conditions