Instrumental variable method for the causal relationship between soluble receptor for advanced glycation end-products (sRAGE) and risk of pancreatic cancer

This thesis project is motivated by the potential problem of using observational data to draw inferences about a causal relationship in observational epidemiology research when controlled randomization is not applicable. Instrumental variable (IV) method is one of the statistical tools to overcome this problem. Mendelian randomization study uses genetic variants as IVs in genetic association study. In this thesis, the IV method, as well as standard logistic and linear regression models, is used to investigate the causal association between risk of pancreatic cancer and the circulating levels of soluble receptor for advanced glycation end-products (sRAGE). Higher levels of serum sRAGE were found to be associated with a lower risk of pancreatic cancer in a previous observational study (255 cases and 485 controls). However, such a novel association may be biased by unknown confounding factors. In a case-control study, we aimed to use the IV approach to confirm or refute this observation in a subset of study subjects for whom the genotyping data were available (178 cases and 177 controls). Two-stage IV method using generalized method of moments-structural mean models (GMM-SMM) was conducted and the relative risk (RR) was calculated. In the first stage analysis, we found that the single nucleotide polymorphism (SNP) rs2070600 of the receptor for advanced glycation end-products (AGER) gene meets all three general assumptions for a genetic IV in examining the causal association between sRAGE and risk of pancreatic cancer. The variant allele of SNP rs2070600 of the AGER gene was associated with lower levels of sRAGE, and it was neither associated with risk of pancreatic cancer, nor with the confounding factors. It was a potential strong IV (F statistic = 29.2). However, in the second stage analysis, the GMM-SMM model failed to converge due to non- concaveness probably because of the small sample size. Therefore, the IV analysis could not support the causality of the association between serum sRAGE levels and risk of pancreatic cancer. Nevertheless, these analyses suggest that rs2070600 was a potentially good genetic IV for testing the causality between the risk of pancreatic cancer and sRAGE levels. A larger sample size is required to conduct a credible IV analysis.^

Regulation of pyridoxal $5\prime$-phosphate biosynthesis in {\it Escherichia coli\/} K-12

Vitamin B$\sb6$ (or pyridoxal 5$\sp\prime$-phosphate, PLP) is an essential, ubiquitous coenzyme that affects many aspects of amino acid and cellular metabolism in all organisms. The goal of this thesis is to examine the regulation of PLP biosynthesis in Escherichia coli K-12. First, PdxH oxidase is a PLP biosynthetic enzyme, which uses molecular oxygen as an electron acceptor under aerobic assay conditions. To test if facultative anaerobic E. coli uses another enzyme to replace the function of PdxH oxidase anaerobically, suppressors of a pdxH null mutant were isolated anaerobically after 2-aminopurine or spontaneous mutagenesis. Only one specific bypass mutation in another PLP biosynthetic gene pdxJ was found, suggesting that PdxH oxidase is able to function anaerobically and PdxT utilizes D-1-deoxyxyulose as a substrate. Second, regulation of the serC (pdxF)-aroA operon, which is involved the biosynthesis of L-serine, PLP and aromatic compounds was examined. A serC (pdxF) single gene transcript and a serC (pdXf)-aroA cotranscript initiated at P$\sb{serC\ (pdxF)}$ upstream of serC (pdxF) were detected. The expression of the operon is activated by leucine responsive regulatory protein (LRP) and repressed by cAMP receptor protein-cAMP complex (CRP$\cdot$cAMP) at the transcriptional level. LRP activates the operon by directly binding to the upstream consensus box. Binding of CRP$\cdot$cAMP to the upstream CRP box diminishes the activation effect of LRP. However, deletion of the CRP box did not affect the repression of CRP$\cdot$cAMP, suggesting that CRP$\cdot$cAMP may repress the operon indirectly by stimulating the activity or level of an unidentified repressor. The overall effect of this regulation is to maximize the expression of the operon when the cells are growing in minimal-glucose medium. In addition, the binding and the transcription of P$\sb{serC\ (pdxF)}$ by RNA polymerase require a supercoiled circular DNA, indicating that DNA supercoiling affects the transcription of the operon. Third, regulation of another PLP biosynthetic gene gapB was also examined. gapB is activated by CRP$\cdot$cAMP and repressed by catabolic repressor activator protein (CRA). However, the activation of CRP$\cdot$cAMP is epistatic to the repression of CRA. Due to the CRA repression, gapB was expressed at a low level in all the media tested, suggesting that it may be the rate-limiting step of PLP biosynthesis. In summary, unlike genes in many biosynthetic pathways, PLP biosynthetic genes are regulated by global regulators that are important for carbon and amino acid metabolism, instead of the end product(s) of the pathway. ^

Ketamine-induced hepatoprotection: the role of heme oxygenase-1.

Lipopolysaccharide (LPS) causes hepatic injury that is mediated, in part, by upregulation of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2). Ketamine has been shown to prevent these effects. Because upregulation of heme oxygenase-1 (HO-1) has hepatoprotective effects, as does carbon monoxide (CO), an end product of the HO-1 catalytic reaction, we examined the effects of HO-1 inhibition on ketamine-induced hepatoprotection and assessed whether CO could attenuate LPS-induced hepatic injury. One group of rats received ketamine (70 mg/kg ip) or saline concurrently with either the HO-1 inhibitor tin protoporphyrin IX (50 micromol/kg ip) or saline. Another group of rats received inhalational CO (250 ppm over 1 h) or room air. All rats were given LPS (20 mg/kg ip) or saline 1 h later and euthanized 5 h after LPS or saline. Liver was collected for iNOS, COX-2, and HO-1 (Western blot), NF-kappaB and PPAR-gamma analysis (EMSA), and iNOS and COX-2 mRNA analysis (RT-PCR). Serum was collected to measure alanine aminotransferase as an index of hepatocellular injury. HO-1 inhibition attenuated ketamine-induced hepatoprotection and downregulation of iNOS and COX-2 protein. CO prevented LPS-induced hepatic injury and upregulation of iNOS and COX-2 proteins. Although CO abolished the ability of LPS to diminish PPAR-gamma activity, it enhanced NF-kappaB activity. These data suggest that the hepatoprotective effects of ketamine are mediated primarily by HO-1 and its end product CO.

The Role of Cortisol in the Cycle of Violence

Child abuse and neglect are universal risk factors for delinquency, violence and aggression; this phenomenon is known as the cycle of violence. Despite a wide body of research demonstrating this phenomenon, the processes which mediate this relationship remain largely unknown. One potentially relevant result of abuse and neglect may be disruptions in the development of the body’s stress response, specifically the function of the Hypothalamic-Pituitary-Adrenal (HPA) axis. The HPA-axis, and its end-product, cortisol, may play a role in regulating aggressive behavior, but this function may be disrupted following abuse and neglect. Another risk factor for aggression, psychopathy, may mediate the cycle of violence or independently contribute to aggressive behavior. This study examined the relationship between child abuse and neglect, HPA-axis function, psychopathy and aggression. History of abuse was measured using a self-report questionnaire, the Childhood Trauma Questionnaire. Using a within-subject, placebo-controlled, counter-balanced dosing design, 67 adults were given an acute dose of 20mg cortisol as a challenge to the HPA-axis. Following dosing, measures of cortisol response were obtained through saliva samples, and state-aggressive behavior was measured by a laboratory task, the Point-Subtraction Aggression Paradigm (PSAP). Basal measures of cortisol were obtained prior to dosing. Psychopathy and a trait-measure of aggression were assessed through self-report questionnaires. PSAP data and trait-aggression scores were normalized and summed for an overall aggression score. Linear regression analyses indicated that a history of abuse and neglect robustly predicted aggression, supporting the cycle of violence hypothesis. Further, abuse and neglect predicted a diminished HPA-axis response to the cortisol challenge. Although a diminished HPA-axis response significantly predicted increased aggression, mediation analysis revealed that HPA-axis reactivity did not mediate a significant portion of the effect of abuse and neglect on aggression. However, HPA-axis reactivity did mediate part of the effect, indicating that HPA-axis function may be a factor in the cycle of violence. Psychopathy robustly predicted increased aggression. Although the results indicate that cortisol, psychopathy and HPA-axis function are involved in the cycle of violence, further research is required to better understand the complex interaction of these factors.

CYTOPLASMIC MALATE DEHYDROGENASE IS THE AROMATIC ALPHA-KETO ACID REDUCTASE IN MAN (PHENYLKETONURIA, PKU, TYROSINE)

This dissertation presents evidence to support the hypothesis that cytoplasmic malate dehydrogenase (MDH-1) is the enzyme in humans which catalyzes the reduction of aromatic alpha-keto acids in the presence of NADH, and the enzyme which has been described in the literature as aromatic alpha-keto acid reductase (KAR; E.C. 1.1.1.96) is actually a secondary activity of cytoplasmic malate dehydrogenase.^ Purified MDH and purified KAR have the same molecular weight, subunit structure, heat-inactivation profile and tissue distribution. After starch gel electrophoresis, and using p-hydroxyphenylpyruvic acid (HPPA) as substrate, KAR activity co-migrates with MDH-1 in all species studied except some marine animals. Inhibition with malate, the end-product of malate dehydrogenase, substantially reduces or totally eliminates KAR activity. Purified cytoplasmic MDH from human erythrocytes has an alpha-keto acid reductase activity with identical mobility. All electrophoretic variants of MDH-1 seen in the fresh-water bony fish Xiphophorus, the amphibians Rana and humans exhibited identical variation for KAR, and the two traits co-segregated in the small group of offspring from one Rana heterozygote studied. Both enzymes show almost no electrophoretic variation among humans from many ethnic groups, and among several inbred strains of mice both MDH-s and KAR co-migrate with no variation. MDH-1 and KAR in mouse and Chinese hamster fibroblasts show identical mobility differences between species. Antisera raised against purified chicken cytoplasmic MDH totally inhibited both MDH-1 and KAR in chickens and humans. Mitochondrial MDH from tissue homogenates has no detectable KAR activity but purified MDH-2 does.^ The previous claim that the gene for KAR is on human chromosome 12 is disputed because both MDH-1 and LDH bands appear with slightly different mobility approximately midway between the human and hamster controls in somatic cell hybrid studies, and the meaning of this artifact is discussed. ^

A study in the molecular and cellular functions of GSK3beta in prostate adenocarcinoma

Kinases are part of a complex network of signaling pathways that enable a cell to respond to changes in environmental conditions in a regulated and coordinated way. For example, Glycogen Synthase Kinase 3 beta (GSK3β) modulates conformational changes, protein-protein interaction, protein degradation, and activation of unique domains in proteins that transduce signals from the extracellular milieu to the nucleus. ^ In this project, I investigated the expression and function that GSK3β exhibits in prostate cells. The capacity of GSK3β to regulate two transcription factors (JUN and CREB), which are known to be inversely utilized in prostate tumor cells, was measured. JUN/AP1 is constitutively activated in PC-3 cells; whereas, CREB/CRE activity is ∼20 fold less than the former. GSK3β overexpression obliterates JUN/AP1 activity. With respect to CREB GSK3β increases CREB/CRE activity. Cellular levels of active GSK3β can determine whether JUN or CREB is preferentially active in the PC-3s. Theoretically, in response to a particular cellular context or stimulus, a cell may coordinate JUN and CREB function by regulating GSK3β.^ A comparison of various prostate cell lines showed that active GSK3β is less expressed in normal prostate epithelial cells than in tumor cells. Differentially expressed active (GSK3β) may correlate with progression of prostate carcinoma. If a known marker associated with carcinoma of the prostate could be shown to be regulated by GSK3β then, further study of GSK3β may lead to a better understanding of both possible prevention of the disease and improved therapy for advanced stages. ^ The androgen receptor (AR) is an intriguing phosphoprotein whose regulation is potentially determined by a variety of kinases. One of these is (GSK3β) I found that (GSK3β) is a regulator of the androgen receptor in both the unliganded and liganded states. It can inhibit AR function as measured by reporter assays. Also, GSK3β associates with the AR at the DNA binding domain because deletion constructs expressing either the n-terminus or the c-terminus (both having the DBD in common) immunoprecipitated with GSK3β. Increased understanding of how GSK3β functions in prostate cancer would provide clues into how (1) certain signal pathways are coordinated and (2) the androgen receptor may be regulated. ^

An intervention mapping approach: Identification of human trafficking victims by health professionals

Background. Today modern day slavery is known as human trafficking and is a growing pandemic that is a grave human rights violation. Estimates suggest that 12.3 million people are working under conditions of force, fraud or coercion. Working toward eradication is a worthy effort; it would free millions of humans from slavery, mostly women and children, as well as uphold basic human rights. One tactic to eradicating human trafficking is to increase identification of victims among those likely to encounter victims of human trafficking.^ Purpose. This study aims to develop an intervention that improves certain stakeholders' ability, in the health clinic setting, to appropriately identify and report victims of human trafficking to the National Human Trafficking Resource Center.^ Methods. The Intervention Mapping (IM) process was used by program planners to develop an intervention for health professionals. This methodology is a six step process that guides program planners to develop an intervention. Each step builds on the others through the execution of a needs assessment, and the development of matrices based on performance objectives and determinants of the targeted health behavior. The end product results in an ecological, theoretical, and evidence based intervention.^ Discussion. The IM process served as a useful protocol for program planners to take an ecological approach as well as incorporate theory and evidence into the intervention. Consultation with key informants, the planning group, adopters, implementers, and individuals responsible for institutionalization also contributed to the practicality and feasibility of the intervention. Program planners believe that this intervention fully meets recommendations set forth in the literature.^ Conclusions. The intervention mapping methodology enabled program planners to develop an intervention that is appropriate and acceptable to the implementer and the recipients.^

THE ASSOCIATION BETWEEN CANCER-RELATED-FATIGUE, RESPONSE TO FATIGUE TREATMENT, AND CYTOKINES AND THEIR RECEPTORS, IN PATIENTS WITH ADVANCED CANCER

Background: Inflammation is implicated in the development of cancer related fatigue (CRF). However there is limited literature on the mediators of inflammation (namely), cytokines and their receptors, associated with clinically significant fatigue and response to treatment. Methods: We reviewed 37 advanced cancer patients with fatigue (≥4/10), who participated in two Randomized Controlled Trials, of anti-inflammatory agents (Thalidomide and Dexamethasone) for CRF. Responders showed improvement in FACIT-F subscale at the end of study (Day 15). Baseline patient characteristics and symptoms were assessed by FACIT-F, ESAS; serum cytokines [IL-1β and receptor antagonist (IL-1RA), IL-6, IL-6R, TNF-α and sTNF-R1 and R2, IL-8, IL-10, IL-17] levels measured by Luminex. Data were analyzed using principal component analysis (PCA) [reporting cumulative variance (variance) for the first four components] to determine their association with fatigue and response to treatment. Results: Females were 54%. Mean (SD) was as follows for age, 61(14); baseline FACIT (F) scores, 21.4(8.6); ESAS Fatigue item, 6.5(1.9); and FACIT-F change, 6.4(9.7); ESAS (fatigue) change, -2 (2.41). Baseline median in pg/mL for IL-6, TNF-α, IL-1β were 31.9; 18.9; 0.55, respectively. Change in IL-6 negatively correlated with change in FACIT-F scores (p=0.02). Baseline CRF (FACIT-F score) was associated with IL-6, IL-6R and IL-17, Variance = 78% whereas IL-10, IL-1RA, TNF-α and IL-1β were associated with improvement of CRF, Variance=74%. Conversely, IL-6 and IL-8 were associated with no improvement or worsening of CRF, Variance= 93%. Conclusions: Change in IL-6 negatively correlated with change in FACIT-F scores. IL-6, IL-6R and IL-17 are associated with CRF while IL-6 and IL-8 were associated with no improvement of CRF. Further studies are warranted confirm our findings.

Time to renal disease and end-stage renal disease in PROFILE: a multiethnic lupus cohort.

BACKGROUND: Renal involvement is a serious manifestation of systemic lupus erythematosus (SLE); it may portend a poor prognosis as it may lead to end-stage renal disease (ESRD). The purpose of this study was to determine the factors predicting the development of renal involvement and its progression to ESRD in a multi-ethnic SLE cohort (PROFILE). METHODS AND FINDINGS: PROFILE includes SLE patients from five different United States institutions. We examined at baseline the socioeconomic-demographic, clinical, and genetic variables associated with the development of renal involvement and its progression to ESRD by univariable and multivariable Cox proportional hazards regression analyses. Analyses of onset of renal involvement included only patients with renal involvement after SLE diagnosis (n = 229). Analyses of ESRD included all patients, regardless of whether renal involvement occurred before, at, or after SLE diagnosis (34 of 438 patients). In addition, we performed a multivariable logistic regression analysis of the variables associated with the development of renal involvement at any time during the course of SLE.In the time-dependent multivariable analysis, patients developing renal involvement were more likely to have more American College of Rheumatology criteria for SLE, and to be younger, hypertensive, and of African-American or Hispanic (from Texas) ethnicity. Alternative regression models were consistent with these results. In addition to greater accrued disease damage (renal damage excluded), younger age, and Hispanic ethnicity (from Texas), homozygosity for the valine allele of FcgammaRIIIa (FCGR3A*GG) was a significant predictor of ESRD. Results from the multivariable logistic regression model that included all cases of renal involvement were consistent with those from the Cox model. CONCLUSIONS: Fcgamma receptor genotype is a risk factor for progression of renal disease to ESRD. Since the frequency distribution of FCGR3A alleles does not vary significantly among the ethnic groups studied, the additional factors underlying the ethnic disparities in renal disease progression remain to be elucidated.

Activation of heat shock and antioxidant responses by the natural product celastrol: transcriptional signatures of a thiol-targeted molecule.

Stress response pathways allow cells to sense and respond to environmental changes and adverse pathophysiological states. Pharmacological modulation of cellular stress pathways has implications in the treatment of human diseases, including neurodegenerative disorders, cardiovascular disease, and cancer. The quinone methide triterpene celastrol, derived from a traditional Chinese medicinal herb, has numerous pharmacological properties, and it is a potent activator of the mammalian heat shock transcription factor HSF1. However, its mode of action and spectrum of cellular targets are poorly understood. We show here that celastrol activates Hsf1 in Saccharomyces cerevisiae at a similar effective concentration seen in mammalian cells. Transcriptional profiling revealed that celastrol treatment induces a battery of oxidant defense genes in addition to heat shock genes. Celastrol activated the yeast Yap1 oxidant defense transcription factor via the carboxy-terminal redox center that responds to electrophilic compounds. Antioxidant response genes were likewise induced in mammalian cells, demonstrating that the activation of two major cell stress pathways by celastrol is conserved. We report that celastrol's biological effects, including inhibition of glucocorticoid receptor activity, can be blocked by the addition of excess free thiol, suggesting a chemical mechanism for biological activity based on modification of key reactive thiols by this natural product.

Association of retinoic acid receptor genes with meningomyelocele.

BACKGROUND: Neural tube defects (NTDs) occur in as many as 0.5-2 per 1000 live births in the United States. One of the most common and severe neural tube defects is meningomyelocele (MM) resulting from failed closure of the caudal end of the neural tube. MM has been induced by retinoic acid teratogenicity in rodent models. We hypothesized that genetic variants influencing retinoic acid (RA) induction via retinoic acid receptors (RARs) may be associated with risk for MM. METHODS: We analyzed 47 single nucleotide polymorphisms (SNPs) that span across the three retinoic acid receptor genes using the SNPlex genotyping platform. Our cohort consisted of 610 MM families. RESULTS: One variant in the RARA gene (rs12051734), three variants in the RARB gene (rs6799734, rs12630816, rs17016462), and a single variant in the RARG gene (rs3741434) were found to be statistically significant at p < 0.05. CONCLUSION: RAR genes were associated with risk for MM. For all associated SNPs, the rare allele conferred a protective effect for MM susceptibility.

THE EFFECTS OF CURCUMIN ON BODY COMPOSITION OF PATIENTS WITH ADVANCED PANCREATIC CANCER

Cachexia is very common among patients with advanced pancreatic cancer and is a marker of poor prognosis. Weight loss in cachexia is due to both adipose and muscle compartments, and sarcopenia (severe muscle depletion) is associated with worse outcomes. Curcumin has shown a myriad of biological effects, including anti-cancer and anti-inflammatory. The ability of curcumin to attenuate cachexia and muscle loss has been tested in animal models, with conflicting results so far. The hypothesis of this study was that patients with advanced pancreatic cancer treated with curcumin for two months have less fat and muscle loss as compared to matched controls not treated with this compound. A matched 1:2 case-control retrospective study was conducted with 22 patients with pancreatic cancer who were treated with curcumin on a previous protocol and 44 untreated controls with the same diagnosis matched by age, gender, time from advanced cancer, body mass index, and number of prior therapies. Data was collected regarding oncologic treatment, medication use, weights, heights, and survival. Body composition was determined by computerized tomography analyses at two timepoints separated by 60±20 days. For treated patients, the first image was at the beginning of treatment and for controls it was determined by the matching time from advanced cancer. The evolution of body composition over time was quantitatively analyzed comparing both groups. All patients lost weight both due to fat and muscle losses, and patients treated with curcumin presented greater losses both in lean adipose body mass. Use of medications, chemotherapy, age, time from advanced cancer, baseline albumin, performance status, and number of prior therapies were not independently correlated with changes in body composition variables. Patients treated with curcumin had borderline shorter survival when compared with untreated patients. Sarcopenic treated patients had significantly shorter survival than non-sarcopenic counterparts, and sarcopenia status was not associated with survival among the controls. Treated patients with shorter survival showed a tendency to lose more lean and especially fat body mass as compared to untreated patients, maybe suggesting an effect of curcumin on shifting weight loss towards the end of life by impacting its mechanisms.

Determinants of quality of advanced cancer care in Latin America. A look at five countries: Argentina, Brazil, Cuba, Mexico and Peru

The World Health Organization reports that nearly half a million people died of cancer in Latin America in 2001. As a growing public health problem, cancer is now either the first or second leading cause of death among adults in most Latin American nations. Despite these trends, information on the quality of care people with advanced cancer in Latin America receive has been limited. This study assessed the quality of advanced cancer care in diverse Latin American countries and institutions by surveying cancer care providers from: Argentina; Brazil; Cuba; Mexico; and Peru. This study also identified the most salient factors that influence the quality of this care at the national and institutional levels and compared these factors across countries. This study was based on the secondary analyses of data collected by the University of Texas M. D. Anderson's WHO/PAHO Collaborating Center in Supportive Cancer Care from March 2000 to November 2002. The sample for this survey was a convenience sample of physicians and nurses who treat cancer patients in these regions. Strategies for the dissemination of this survey included: mass mailings; distribution at professional meetings/conferences; collaboration with regional institutions, professional organizations and PAHO; and the posting of online surveys. The strongest predictor of providers' assessments of the quality of advanced cancer care was their ratings of access to care. This major finding reflects a shared equitable notion of quality care among providers from diverse countries and medical institutions that is highly interrelated with providing accessible care to those with advanced cancer. Higher ratings of the affordability of care, an increased reported availability of end-of-life services and opioid analgesics, practicing in either a private hospital or specialized cancer center, and practicing in Cuba were also associated with higher provider ratings of the quality of advanced cancer care. The findings of this study contribute towards the much needed body of knowledge that may guide the formulation of policies and interventions aimed at improving the care for people with advanced cancer in Latin America. ^

Targeting dysregulated nuclear proteins androgen receptor and enhancer of zeste homolog 2: Clinical implication and mechanism underline

Cancer is the most devastating disease that has tremendous impacts on public health. Many efforts have been devoted to fighting cancer through either translational or basic researches for years. Nowadays, it emerges the importance to converge these two research directions and complement to each other for battling with cancer. Thus, our study aims at both translational and basic research directions. The first goal of our study is focus on translational research to search for new agents targeting prevention and therapy of advanced prostate cancer. Hormone refractory prostate cancer is incurable and lethal. Androgen receptor (AR) mediates androgen's effect not only on the tumor initiation but also plays the major role in the relapse transition of prostate cancer. Here we demonstrate that emodin, a natural compound, can directly target AR to suppress prostate cancer cell growth in vitro and prolong the survival of C3(1)/SV40 transgenic mice in vivo. Emodin treatment resulted in repressing androgen-dependent transactivation of AR by inhibiting AR nuclear translocation. Emodin decreased the association of AR and heat shock protein 90 and increased the association of AR and MDM2, which in turn, induces AR degradation through a proteasome-mediated pathway in a ligand independent manner. Our work indicates a new mechanism for the emodin-mediated anticancer effect and justifies further investigation of emodin as a therapeutic and preventive agent for prostate cancer. The second goal of our study is try to elucidate the fundamental tumor biology of cancer progression then provide the rationale to develop more efficient therapeutic strategy. Enhancer of zeste homologue 2 (EZH2) plays an important role in many biological processes through its intrinsic methyltransferase activity to trimethylate lysine 27 in histone H3. Although overexpression of EZH2 has been shown to be involved in cancer progression, the detailed mechanisms are elusive. Here, we show that Akt phosphorylates EZH2 at serine 21 and suppresses its methyltransferase activity by impeding the binding to its substrate histone H3, resulting in a decrease of lysine 27 trimethylation and derepression of silenced genes, thus promotes cell proliferation and tumorigenicity. Our results also show that histone methylation is not permanent but regulated in a dynamic manner and that the Akt signaling pathway is involved in the regulation of this epigenetic modification through phosphorylation of EZH2, thus contributing to oncogenic processes. ^

Evaluation of the C5a anaphylatoxin and its receptor (CD88) in allergic lung disease

Allergic asthma is characterized by airflow obstruction, airway hyperresponsiveness (AHR) and chronic airway inflammation. We and others have reported that complement component C3 and the anaphylatoxin C3a receptor promote while C5 protects against the development of the biological and physiological hallmarks of allergic lung disease in mice. In this study, we assessed if the protective responses could be mediated by C5a, an activation-induced C5 cleavage product. Mice with ablation of the C5a receptor (C5aR) either by genetic deletion or by pharmacological blockade exhibited significantly exacerbated AHR compared to allergen-challenged wild-type (WT) mice. However, there were no significant differences in many of the other hallmarks of asthma such as airway infiltration by eosinophils or lymphocytes, pulmonary IL-4-producing cell numbers, goblet cell metaplasia, mucus secretion or total serum IgE levels. In contrast to elevated AHR, numbers of IL-5 and IL-13 producing pulmonary cells, and IL-5 and IL-13 protein levels, were significantly reduced in allergen-challenged C5aR-/- mice compared to allergen-challenged WT mice. Administration of a specific cysteinyl leukotriene receptor 1 (cysLT1R) antagonist before each allergen-challenge abolished AHR in C5aR-/- as well as in WT mice. Pretreatment with a C3aR antagonist dose-dependently reduced AHR in allergen-challenged WT and C5aR-/- mice. Additionally, allergen-induced upregulation of pulmonary C3aR expression was exaggerated in C5aR-/- mice compared to WT mice. In summary, deficiency or antagonism of C5aR in a mouse model of pulmonary allergy increased AHR, which was reversed or reduced by blockade of the cysLT1R and C3aR, respectively. In conclusion, this study suggests that C5a and C5aR mediate protection against AHR by suppressing cysLT and C3aR signaling pathways, which are known to promote AHR. This also supports important and opposing roles of complement components C3a/C3aR and C5a/C5aR in AHR. ^