Topical Polyethylene Glycol as a Novel Chemopreventive Agent for Oral Cancer via Targeting of Epidermal Growth Factor Response

Head and neck squamous cell carcinoma (HNSCC) is a major cause of morbidity and mortality underscoring the need for safe and effective chemopreventive strategies. Targeting epidermal growth factor receptor (EGFR) is attractive in that it is an early critical event in HNSCC pathogenesis. However, current agents lack efficacy or have unacceptable toxicity. Several groups have demonstrated that the over-the-counter medication, polyethylene glycol (PEG) has remarkable chemopreventive efficacy against colon carcinogenesis. Importantly, we reported that this effect is mediated through EGFR internalization/degradation. In the current study, we investigated the chemopreventive efficacy of this agent against HNSCC, using both the well validated animal model 4-NQO (4-nitroquinoline 1-oxide) rat model and cell culture with the human HNSCC cell line SCC-25. We demonstrated that daily topical application of 10% PEG-8000 in the oral cavity (tongue and cavity wall) post 4NQO initiation resulted in a significant reduction in tumor burden (both, tumor size and tumors/tumor bearing rat) without any evidence of toxicity. Immunohistochemical studies depicted decreased proliferation (number of Ki67-positive cells) and reduced expression of EGFR and its downstream effectors cyclin D1 in the tongue mucosa of 4NQO-rats treated with PEG. We showed that EGFR was also markedly downregulated in SCC-25 cells by PEG-8000 with a concomitant induction of G1-S phase cell-cycle arrest, which was potentially mediated through upregulated p21cip1/waf1. In conclusion, we demonstrate, for the first time, that PEG has promising efficacy and safety as a chemopreventive efficacy against oral carcinogenesis

Interferon-inducible gene 202b controls CD8+ T cell-mediated suppression in anti-DNA Ig peptide-treated (NZB × NZW) F1 lupus mice

Administration of an artificial peptide (pConsensus) based on anti-DNA IgG sequences that contain major histocompatibility complex class I and class II T-cell determinants, induces immune tolerance in NZB/NZW F1 female (BWF1) mice. To understand the molecular basis of CD8+ Ti-mediated suppression, we previously performed microarray analysis to identify genes that were differentially expressed following tolerance induction with pCons. CD8+ T cells from mice tolerized with pCons showed more than two-fold increase in Ifi202b mRNA, an interferon inducible gene, versus cells from untolerized mice. Ifi202b expression increased through weeks 1–4 after tolerization and then decreased, reapproaching baseline levels at 6 weeks. In vitro polyclonal activation of tolerized CD8+ T cells significantly increased Ifi202b mRNA expression. Importantly, silencing of Ifi202b abrogated the suppressive capacity of CD8+ Ti cells. This was associated with decreased expression of Foxp3, and decreased gene and protein expression of transforming growth factor (TGF)β and interleukin-2 (IL-2), but not of interferon (IFN)-γ, IL-10, or IL-17. Silencing of another IFN-induced gene upregulated in tolerized CD8+ T cells, IFNAR1, had no effect on the ability of CD8+ T cells to suppress autoantibody production. Our findings indicate a potential role for Ifi202b in the suppressive capacity of peptide-induced regulatory CD8+ Ti cells through effects on the expression of Foxp3 and the synthesis of TGFβ

Mixing in a swarm of bubbles rising in a confined cell measured by mean of PLIF with two different dyes

The present contribution reports an experimental study of the mixing of a passive scalar of very low diffusivity in a homogeneous swarm of inertial bubbles rising in a thin gap. A patch of fluorescent dye is injected within the swarm, and we observe the evolution of its mass in a given region of observation. We analyse the effect of the liquid agitation on the mixing mechanisms varying the gas volume fraction from 1.3 to 7.5 %, while the Reynolds number of the bubbles, Re = 450, their Weber number, We = 0.7, and the gapto-bubble diameter ratio, w/d = 0.25, are kept approximately constant. Here, the in-plane local mass of dye is measured by using a two-dyes planar laser-induced fluorescence (PLIF) technique that has been adapted to fix the problem of multiple light reflections at the bubble interfaces. Indeed, they induce both temporal and spatial variations of the captured light intensity that are superimposed to the effective fluorescence signal and prevent from using a standard PLIF technique. The analysis of the instantaneous concentration fields reveals the dominant role of the bubble wakes in the scalar transport. It is shown that mixing in this planar confined geometry is very efficient and enhanced by the increasing gas volume fraction. The present study also highlights that the mixing is not governed by a Fickian law of diffusion

Mutation analysis of "Endoglin" and "Activin receptor-like kinase" genes in German patients with hereditary hemorrhagic telangiectasia and the value of rapid genotyping using an allele-specific PCR-technique

<p>Abstract</p> <p>Background</p> <p>Hereditary hemorrhagic telangiectasia (HHT), also known as Rendu-Osler-Weber syndrome, is an autosomal dominant disorder which is clinically characterised by recurrent epistaxis, mucocutaneous telangiectasia and visceral arteriovenous malformations. Genetic linkage studies identified two genes primarily related to HHT: endoglin (<it>ENG</it>) on chromosome 9q33-34 and activin receptor-like kinase1 (<it>ACVRL1</it>) on chromosome 12q13. We have screened a total of 41 unselected German patients with the suspected diagnosis of HHT. Mutation analysis for the <it>ENG </it>and <it>ACVRL1 </it>genes in all patients was performed by PCR amplification. Sequences were then compared to the HHT database <url>http://www.hhtmutation.org</url> sequences of the <it>ENG </it>mRNA (accession no. BC014271.2) and the <it>ACVRL1 </it>mRNA (accession no. NM000020.1).</p> <p>Results</p> <p>We identified 15 different mutations in 18 cases by direct sequencing. Among these mutations, one novel <it>ENG </it>mutation could be detected which has not yet been described in the literature before. The genotype-phenotype correlation was consistent with a higher frequency of pulmonary arteriovenous malformations in patients with <it>ENG </it>mutations than in patients with <it>ACVRL1 </it>mutations in our collective.</p> <p>Conclusion</p> <p>For rapid genotyping of mutations and SNPs (single nucleotide polymorphisms) in <it>ENG </it>and <it>ACVRL1</it>, allele-specific PCR methods with sequence-specific primers (PCR-SSP) were established and their value analysed.</p

Antibiotic Treatment of the Tick Vector Amblyomma americanum Reduced Reproductive Fitness

BACKGROUND: The lone star tick Amblyomma americanum is a common pest and vector of infectious diseases for humans and other mammals in the southern and eastern United States. A Coxiella sp. bacterial endosymbiont was highly prevalent in both laboratory-reared and field-collected A. americanum. The Coxiella sp. was demonstrated in all stages of tick and in greatest densities in nymphs and adult females, while a Rickettsia sp. was less prevalent and in lower densities when present. METHODOLOGY/PRINCIPAL FINDINGS: We manipulated the numbers of both bacterial species in laboratory-reared A. americanum by injecting engorged nymphs or engorged, mated females with single doses of an antibiotic (rifampin or tetracycline) or buffer alone. Burdens of the bacteria after molting or after oviposition were estimated by quantitative polymerase chain reaction with primers and probes specific for each bacterial species or, as an internal standard, the host tick. Post-molt adult ticks that had been treated with rifampin or tetracycline had lower numbers of the Coxiella sp. and Rickettsia sp. and generally weighed less than ticks that received buffer alone. Similarly, after oviposition, females treated previously with either antibiotic had lower burdens of both bacterial species in comparison to controls. Treatment of engorged females with either antibiotic was associated with prolonged time to oviposition, lower proportions of ticks that hatched, lower proportions of viable larvae among total larvae, and lower numbers of viable larvae per tick. These fitness estimators were associated with reduced numbers of the Coxiella sp. but not the Rickettsia sp. CONCLUSION/SIGNIFICANCE: The findings indicate that the Coxiella sp. is a primary endosymbiont, perhaps provisioning the obligately hematophagous parasites with essential nutrients. The results also suggest that antibiotics could be incorporated into an integrated pest management plan for control of these and other tick vectors of disease

Reduced Apaf-1 expression in human cutaneous melanomas

Malignant melanoma is a life-threatening skin cancer due to its highly metastatic character and resistance to radio- and chemotherapy. It is believed that the ability to evade apoptosis is the key mechanism for the rapid growth of cancer cells. However, the exact mechanism for failure in the apoptotic pathway in melanoma cells is unclear. p53, the most frequently mutated tumour suppressor gene in human cancers, is a key apoptosis inducer. However, p53 mutation is only found in 15–20% of melanoma biopsies. Recently, it was found that Apaf-1, a downstream target of p53, is inactivated in metastatic melanoma. Specifically, loss of heterozygosity (LOH) of the Apaf-1 gene was found in 40% of metastatic melanoma. To determine if loss of Apaf-1 expression is indeed involved in melanoma progression, we employed the tissue microarray technology and examined Apaf-1 expression in 70 human primary malignant melanoma biopsies by immunohistochemistry. Our data showed that Apaf-1 expression is significantly reduced in melanoma cells compared with normal nevi (χ2=6.02, P=0.014). Our results also revealed that loss of Apaf-1 was not associated with the tumour thickness, ulceration or subtype, patient's gender, age and 5-year survival. In addition, our in vitro apoptosis assay revealed that overexpression of Apaf-1 can sensitise melanoma cells to anticancer drug treatment. Taken together, our data indicate that Apaf-1 expression is significantly reduced in human melanoma and that Apaf-1 may serve as a therapeutic target in melanoma

Aqueous extracts from dietary supplements influence the production of inflammatory cytokines in immortalized and primary T lymphocytes

<p>Abstract</p> <p>Background</p> <p>Congaplex^®and Immuplex^®are dietary supplements that have been traditionally used to support immune system function. The purpose of these experiments was to determine whether Congaplex^®and Immuplex^®affect immune function using primary and immortalized T lymphocytes.</p> <p>Methods</p> <p>Immortalized CEM and Jurkat T lymphocytes and primary peripheral mononuclear blood cells (PBMCs) were treated with the aqueous extracts from Congaplex^®and Immuplex^®to determine the effects of these products on cytokine production in activated T lymphocytes.</p> <p>Results</p> <p>Congaplex^®enhanced phytohemagglutinin/phorbol 12-myristate 13-acetate (PHA/PMA) stimulation of both CEM and Jurkat cells as measured by the production of cytokines, while Immuplex^®suppressed PHA/PMA-induced production of cytokines, with the exception of interleukin (IL)-8 which was enhanced by Immuplex^®. <it>In vitro </it>treatment of PBMCs from 10 healthy donors with Congaplex^®or Immuplex^®decreased PHA-stimulated production of interferon (IFN)-γ but increased the production of IL-13.</p> <p>Conclusions</p> <p>While the effects of Congaplex^®and Immuplex^®differed in these two models, these data demonstrate that the aqueous extracts from these two dietary supplements can affect the inflammatory response of T lymphocytes.</p

Activity of the multikinase inhibitor dasatinib against ovarian cancer cells

BackgroundHere, we explore the therapeutic potential of dasatinib, a small-molecule inhibitor that targets multiple cytosolic and membrane-bound tyrosine kinases, including members of the Src kinase family, EphA2, and focal adhesion kinase for the treatment of ovarian cancer.MethodsWe examined the effects of dasatinib on proliferation, invasion, apoptosis, cell-cycle arrest, and kinase activity using a panel of 34 established human ovarian cancer cell lines. Molecular markers for response prediction were studied using gene expression profiling. Multiple drug effect/combination index (CI) isobologram analysis was used to study the interactions with chemotherapeutic drugs.ResultsConcentration-dependent anti-proliferative effects of dasatinib were seen in all ovarian cancer cell lines tested, but varied significantly between individual cell lines with up to a 3 log-fold difference in the IC(50) values (IC(50) range: 0.001-11.3 micromol l(-1)). Dasatinib significantly inhibited invasion, and induced cell apoptosis, but less cell-cycle arrest. At a wide range of clinically achievable drug concentrations, additive and synergistic interactions were observed for dasatinib plus carboplatin (mean CI values, range: 0.73-1.11) or paclitaxel (mean CI values, range: 0.76-1.05). In this study, 24 out of 34 (71%) representative ovarian cancer cell lines were highly sensitive to dasatinib, compared with only 8 out of 39 (21%) representative breast cancer cell lines previously reported. Cell lines with high expression of Yes, Lyn, Eph2A, caveolin-1 and 2, moesin, annexin-1, and uPA were particularly sensitive to dasatinib.ConclusionsThese data provide a clear biological rationale to test dasatinib as a single agent or in combination with chemotherapy in patients with ovarian cancer

Early Category-Specific Cortical Activation Revealed by Visual Stimulus Inversion

Visual categorization may already start within the first 100-ms after stimulus onset, in contrast with the long-held view that during this early stage all complex stimuli are processed equally and that category-specific cortical activation occurs only at later stages. The neural basis of this proposed early stage of high-level analysis is however poorly understood. To address this question we used magnetoencephalography and anatomically-constrained distributed source modeling to monitor brain activity with millisecond-resolution while subjects performed an orientation task on the upright and upside-down presented images of three different stimulus categories: faces, houses and bodies. Significant inversion effects were found for all three stimulus categories between 70–100-ms after picture onset with a highly category-specific cortical distribution. Differential responses between upright and inverted faces were found in well-established face-selective areas of the inferior occipital cortex and right fusiform gyrus. In addition, early category-specific inversion effects were found well beyond visual areas. Our results provide the first direct evidence that category-specific processing in high-level category-sensitive cortical areas already takes place within the first 100-ms of visual processing, significantly earlier than previously thought, and suggests the existence of fast category-specific neocortical routes in the human brain