GM-CSF (granulocyte-macrophage colony-stimulating factor): Modulation der durch UV-B-Strahlung induzierten Hautkarzinogenese

UV-B-Strahlung, die durch die fortschreitende Zerstörung der Ozonschicht zunimmt, ist hauptsächlich für das Entstehen von Basaliomen und Plattenepithelkarzinomen verantwort-lich, an denen jedes Jahr etwa 2-3 Millionen Menschen weltweit erkranken. UV-B indu-zierte Hautkarzinogenese ist ein komplexer Prozess, bei dem vor allem die mutagenen und immunsuppressiven Wirkungen der UV-B-Strahlung von Bedeutung sind. Die Rolle von GM-CSF in der Hautkarzinogenese ist dabei widersprüchlich. Aus diesem Grund wurde die Funktion von GM-CSF in vivo in der UV-B induzierten Hautkarzinogenese mittels zwei bereits etablierter Mauslinien untersucht: Erstens transgene Mäuse, die einen GM-CSF Antagonisten unter der Kontrolle des Keratin-10-Promotors in den suprabasalen Schichten der Epidermis exprimieren und zweitens solche, die unter dem Keratin-5-Promotor murines GM-CSF in der Basalschicht der Epidermis überexprimieren. Eine Gruppe von Tieren wurde chronisch, die andere akut bestrahlt. Die konstitutionelle Verfassung der Tiere mit erhöhter GM-CSF-Aktivität in der Haut war nach chronischer UV-B-Bestrahlung insgesamt sehr schlecht. Sie wiesen deshalb eine stark erhöhte Mortali-tät auf. Dies ist sowohl auf die hohe Inzidenz als auch dem frühen Auftreten der benignen und malignen Läsionen zurückzuführen. Eine verminderte GM-CSF Aktivität verzögerte dagegen die Karzinomentwicklung und erhöhte die Überlebensrate leicht. GM-CSF wirkt auf verschiedenen Ebenen tumorpromovierend: Erstens erhöht eine gesteigerte Mastzell-anzahl in der Haut der GM-CSF überexprimierenden Tiere per se die Suszeptibilität für Hautkarzinogenese. Zweitens stimuliert GM-CSF die Keratinozytenproliferation. Dadurch kommt es nach UV-B-Bestrahlung zu einer prolongierten epidermalen Hyperproliferation, die zur endogenen Tumorpromotion beiträgt, indem sie die Bildung von Neoplasien unter-stützt. Der Antagonist verzögert dagegen den Proliferationsbeginn, die Keratinozyten blei-ben demzufolge länger in der G1-Phase und der durch UV-B verursachte DNA-Schaden kann effizienter repariert werden. Drittens kann GM-CSF die LCs nicht als APCs aktivie-ren und eine Antitumorimmunität induzieren, da UV-B-Strahlung zur Apoptose von LCs bzw. zu deren Migration in Richtung Lymphknoten führt. Zusätzlich entwickeln GM-CSF überexprimierende Tiere in ihrer Haut nach UV-B-Bestrahlung ein Millieu von antago-nistisch wirkenden Zytokinen, wie TNF-a, TGF-b1 und IL-12p40 und GM-CSF, die proinflammatorische Prozesse und somit die Karzinomentwicklung begünstigen. Der Anta-gonist hemmt nach UV-B-Bestrahlung die Ausschüttung sowohl von immunsuppressiven Zytokinen, wie etwa TNF-a, als auch solchen, die die Th2-Entwicklung unterstützen, wie etwa IL-10 und IL-4. Dies wirkt sich negativ auf die Karzinomentwicklung aus.

In vivo and in vitro investigation of the tissue-specific activity of the human CYP3A4 and CYP3A5 promoters

The human cytochrome P450 3A4 (CYP3A4), the predominant but variably expressed cytochrome P450 in adult liver and small intestine is involved in the metabolism of over 50% of currently used drugs. Its paralog CYP3A5 plays a crucial role in the disposition of several drugs with low therapeutic index, including tacrolimus. Limited information is available for the CYP3A5 transcriptional regulation and its induction by xenobiotics remains controversial. In the first part of this study, we analysed the CYP3A5 transcriptional regulation and its induction by xenobiotics in vivo using transgenic mice. To this end, two transgenic strains were established by pronuclear injection of a plasmid, expressing firefly luciferase driven by a 6.2 kb of the human CYP3A5 promoter. A detailed analysis of both strains shows a tissue distribution largely reflecting that of CYP3A5 transcripts in humans. Thus, the highest luciferase activity was detected in the small intestine, followed by oesophagus, testis, lung, adrenal gland, ovary, prostate and kidney. However, no activity was observed in the liver. CYP3A5-luc transgenic mice were similarly induced in both sexes with either PCN or TCPOBOP in small intestine in a dose-dependent manner. Thus, the 6.2 kb upstream promoter of CYP3A5 mediates the broad tissue activity in transgenic mice. CYP3A5 promoter is inducible in the small intestine in vivo, which may contribute to the variable expression of CYP3A in this organ. rnThe hepato-intestinal level of the detoxifying oxidases CYP3A4 and CYP3A5 is adjusted to the xenobiotic exposure mainly via the xenosensor and transcriptional factor PXR. CYP3A5 is additionally expressed in several other organs lacking PXR, including kidney. In the second part of this study, we investigated the mechanism of the differential expression of CYP3A5 and CYP3A4 and its evolutionary origin using renal and intestinal cells, and comparative genomics. For this examination, we established a two-cell line models reflecting the expression relationships of CYP3A4 and CYP3A5 in the kidney and small intestine in vivo. Our data demonstrate that the CYP3A5 expression in renal cells was enabled by the loss of a suppressing Yin Yang 1 (YY1)-binding site from the CYP3A5 promoter. This allowed for a renal CYP3A5 expression in a PXR-independent manner. The YY1 element is retained in the CYP3A4 gene, leading to its suppression, perhaps via interference with the NF1 activity in renal cells. In intestinal cells, the inhibition of CYP3A4 expression by YY1 is abrogated by a combined activating effect of PXR and NF1 acting on their respective response elements located adjacent to the YY1-binding site on CYP3A4 proximal promoter. CYP3A4 expression is further facilitated by a point mutation attenuating the suppressing effect of YY1 binding site. The differential expression of CYP3A4 and CYP3A5 in these organs results from the loss of the YY1 binding element from the CYP3A5 promoter, acting in concert with the differential organ expression of PXR, and with the higher accumulation of PXR response elements in CYP3A4. rn

ELUCIDATING FUNCTIONAL ROLES FOR MYOGENIN IN ADULT SKELETAL MUSCLE METABOLISM, EXERCISE CAPACITY, AND REGENERATION

The four basic helix-loop-helix myogenic transcription factors, myogenin, Myf5, MRF4, and MyoD are critical for embryonic skeletal muscle development. Myogenin is necessary for the terminal differentiation of myoblasts into myofibers during embryogenesis, but little is known about the roles played by myogenin in adult skeletal muscle function and metabolism. Furthermore, while metabolism is a well-studied physiological process, how it is regulated at the transcriptional level remains poorly understood. In this study, my aim was to determine the function of myogenin in adult skeletal muscle metabolism, exercise capacity, and regeneration. To investigate this, I utilized a mouse strain harboring the Myogflox allele and a Cre recombinase transgene, enabling the efficient deletion of myogenin in the adult mouse. Myogflox/flox mice were stressed physically through involuntary treadmill running and by breeding them with a strain harboring the Duchenne’s muscular dystrophy (DMDmdx) allele. Surprisingly, Myog-deleted animals exhibited an enhanced capacity for exercise, running farther and faster than their wild-type counterparts. Increased lactate production and utilization of glucose as a fuel source indicated that Myog-deleted animals exhibited an increased glycolytic flux. Hypoglycemic Myog-deleted mice no longer possessed the ability to outrun their wild-type counterparts, implying the ability of these animals to further deplete their glucose reserves confers their enhanced exercise capacity. Moreover, Myog-deleted mice exhibited an enhanced response to long-term exercise training. The mice developed a greater proportion of type 1 oxidative muscle fibers, and displayed increased levels of succinate dehydrogenase activity, indicative of increased oxidative metabolism. Mdx:Myog-deleted mice exhibited a similar phenotype, outperforming their mdx counterparts, although lagging behind wild-type animals. The morphology of muscle tissue from mdx:Myog-deleted mice appears to mimic that of mdx animals, indicating that myogenin is dispensable for adult skeletal muscle regeneration. Through global gene expression profiling and quantitative (q)RT-PCR, I identified a unique set of putative myogenin-dependent genes involved in regulating metabolic processes. These data suggest myogenin’s functions during adulthood are distinctly different than those during embryogenesis, and myogenin acts as a high-level transcription factor regulating metabolic activity in adult skeletal muscle.

Analysis of perinatal gene expression: hormone response elements mediate activation of a lacZ reporter gene in liver of transgenic mice.

The transcription of genes encoding gluconeogenic enzymes is tightly regulated during the perinatal period. These genes are induced by glucagon (cAMP) and glucocorticoids and repressed by insulin. To address the role of cAMP and glucocorticoids in the physiological activation of genes encoding gluconeogenic enzymes in the perinatal period, transgenic mice have been generated with chimeric constructs containing the reporter gene lacZ under the control of hormone response elements. The activity of the transgene is restricted to the liver by the presence of the enhancers from the alpha-fetoprotein gene and its transcription is driven by a promoter that contains a TATA box linked to either cAMP response elements (CREs) or glucocorticoid response elements (GREs). We demonstrate cAMP and glucocorticoid regulation, liver-specific expression, and perinatal activation of the reporter gene. These data indicate that the CRE and GRE are, independently, necessary and sufficient to mediate perinatal gene activation. Perinatal activation was not impaired when a CRE reporter transgene was assayed in mice that contain a targeted mutation of the CRE-binding protein (CREB) gene, providing further evidence for functional redundancy among the members of the CREB/ATF gene family.

Revue des traditions populaires.

Includes music (unaccompanied melodies)

Musée des monumens français, ou Description historique et chronologique des statues en marbre et en bronze, bas-reliefs et tombeaux des hommes et des femmes célèbres, pour servir à l'histoire de France et à celle de l'art; ornée de gravures; et augm. d'une dissertation sur les costumes de chaque siècle;

"Notes historiques sur les exhumations faites on 1793, dans l'Abbaye de Saint-Denis": v. 2, p. [cvi]-cxxxii.

Comparison of selective and non selective cyclo-oxygenase 2 inhibitors in experimental colitis exacerbation: role of leukotriene B4 and superoxide dismutase

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Ruolo dell'Infliximab nella terapia della colite ulcerosa moderato-severa

Background: Almost 10-15% of patients with active Ulcerative Colitis are refractory to conventional therapy. Infliximab is a treatment of proven efficacy in this group of patients. Aims: To evaluate the role of Inliximab in inducing and maintaining remission in patients with chronically active moderate-severe Ulcerative Colitis. Materials and methods: 53 patients were enrolled, 47 patients entered the study and were treated with a dose of 5 mg/kg. The remission was evaluated through endoscopy and clinical criteria. (Mayo Score). The primary endpoint were clinical and endoscopic remission in moderate-severe Ulcerative Colitis refractory to standard therapy, the secondary out point was the maintenance of remission in the long period. Results: 47 patients started the study, 43 completed the study, 4 dropped out for worsening disease or adverse events; 27 patients were treated with 3 infusions, 9 patients with 4 infusions, 7 patients with > o = 5 infusions. 34 /47 patients (72.3%) were responders 12 (25.5%) improved their symptoms, 22 ( 46.8%) were in remission after the treatment. Among the responders, 21/34 (61.8%) stopped the steroid therapy after 3 infusions, the others reduced the dose or maintained just topic therapy. 13/47 patients (27.7%) were non responders (p <0.001). After 3 months all 22 patients which had reached remission maintained low Mayo Score; 10/12 (83.3%) patients with clinical response maintained their low score, 2 relapsed . Conclusions: Infliximab is a valid therapy for the treatment of Ulcerative Colitis and can avoid surgery in selected patients.

Ruolo della Interleuchina-6 nella colite ulcerosa long-standing: regolazione della p53 ed induzione delle cellule mesenchimali epiteliali

L’infiammazione cronica è un fattore di rischio di insorgenza del cancro, e la citochina infiammatoria IL-6 gioca un ruolo importante nella tumorigenesi. In questo studio abbiamo dimostrato che L’IL-6 down-regola l'espressione e l'attività di p53. In linee cellulari umane, IL-6 stimola la trascrizione dell’rRNA mediante espressione della proteina c-myc a livello post-trascrizionale in un meccanismo p38MAPK-dipendente. L'up-regolazione della biogenesi ribosomiale riduce l'espressione di p53 attraverso l'attivazione della via della proteina ribosomale-MDM2. La down-regolazione di p53 produce l’acquisizione di modifiche fenotipiche e funzionali caratteristiche della epitelio mesenchimale di transizione, un processo associato a trasformazione maligna e progressione tumorale. I nostri dati mostrano che questi cambiamenti avvengono anche nelle cellule epiteliali del colon di pazienti affetti da colite ulcerosa, un esempio rappresentativo di una infiammazione cronica soggetta a trasformazione neoplastica, che scompaiono dopo trattamento con farmaci antinfiammatori. Questi risultati svelano un nuovo effetto oncogenico indotto dall’IL-6 che può contribuire notevolmente ad aumentare il rischio di sviluppare il cancro non solo in pazienti con infiammazioni croniche, ma anche in quei pazienti con condizioni patologiche caratterizzate da elevato livello di IL-6 nel plasma, quali l'obesità e e il diabete mellito di tipo 2.

The Salmonella pathogenicity island (SPI)-2 and SPI-1 type III secretion systems allow Salmonella serovar typhimurium to trigger colitis via MyD88-dependent and MyD88-independent mechanisms.

Salmonella typhimurium can colonize the gut, invade intestinal tissues, and cause enterocolitis. In vitro studies suggest different mechanisms leading to mucosal inflammation, including 1) direct modulation of proinflammatory signaling by bacterial type III effector proteins and 2) disruption or penetration of the intestinal epithelium so that penetrating bacteria or bacterial products can trigger innate immunity (i.e., TLR signaling). We studied these mechanisms in vivo using streptomycin-pretreated wild-type and knockout mice including MyD88(-/-) animals lacking an adaptor molecule required for signaling via most TLRs. The Salmonella SPI-1 and the SPI-2 type III secretion systems (TTSS) contributed to inflammation. Mutants that retain only a functional SPI-1 (M556; sseD::aphT) or a SPI-2 TTSS (SB161; DeltainvG) caused attenuated colitis, which reflected distinct aspects of the colitis caused by wild-type S. typhimurium: M556 caused diffuse cecal inflammation that did not require MyD88 signaling. In contrast, SB161 induced focal mucosal inflammation requiring MyD88. M556 but not SB161 was found in intestinal epithelial cells. In the lamina propria, M556 and SB161 appeared to reside in different leukocyte cell populations as indicated by differential CD11c staining. Only the SPI-2-dependent inflammatory pathway required aroA-dependent intracellular growth. Thus, S. typhimurium can use two independent mechanisms to elicit colitis in vivo: SPI-1-dependent and MyD88-independent signaling to epithelial cells and SPI-2-dependent intracellular proliferation in the lamina propria triggering MyD88-dependent innate immune responses.

Capsule endoscopy in inflammatory bowel disease type unclassified and indeterminate colitis serologically negative

BACKGROUND: The value of capsule endoscopy in the setting of inflammatory bowel disease type unclassified (IBDU) and indeterminate colitis (IC) remains obscure. The aim was to evaluate the clinical impact of capsule endoscopy on IBDU/IC patients with negative serology. METHODS: Eighteen patients with long-standing IBDU (n = 14) and IC (n = 4) were enrolled to undergo a capsule endoscopy and then followed prospectively. Lesions considered diagnostic of Crohn's disease (CD) were 4 or more erosions/ulcers and/or a stricture. The median follow-up time after capsule endoscopy was 32 ± 11 months (23-54 months). RESULTS: Total enteroscopy was possible in all patients. In 2 patients the examination was normal (Group 1). In 9 patients subtle findings were observed (Group 2): focal villi denudation (n = 1) and fewer than 4 erosions/ulcers (n = 8). In 7 patients, 4 or more erosions/ulcers were detected (Group 3), leading to a diagnosis of CD. However, their treatment was not reassessed on the basis of the capsule findings. Until now, a definitive diagnosis has been achieved in 2 additional patients: 1 from Group 1 (ulcerative colitis) and another patient from Group 2 (CD), who began infliximab infusions. Nine patients remained indeterminate at follow-up. CONCLUSIONS: Although capsule endoscopy enabled the diagnosis of CD in 7 patients, in none of them was the clinical management changed. Moreover, a change in therapy due to a diagnosis of CD was made for only 1 patient, who presented nonspecific findings. Our results suggest that capsule findings are not helpful in the work-up of these patients