An exploratory study on the effect of choline and folate deficiency on levels of vascularization proteins and transcription factors in first trimester trophoblast HTR-8/SVneo cells.

AIMS: We studied the effect of choline and folate deficiencies on levels of predetermined placental proteins during early development. METHODS: We incubated HTR-8/SVneo cells under choline and folate deficiency conditions and measured levels of some placental proteins using ELISA methods. RESULTS: Concentrations of LRP2 protein in cell lysates were higher in cells incubated in choline and folate deficient media compared to the control media (mean [SD] = 2.95 [1.30] vs. 1.65 [0.27] ng/mg protein, p = 0.004). The levels of LRP2 protein in lysates of cells incubated in choline and folate deficient media were significantly higher than the concentrations in lysates of cells incubated in choline deficient but folate sufficient media (1.96 [0.28] ng/mg protein) or those incubated in choline sufficient but folate deficient media (1.77 [0.24] ng/mg protein) (p < 0.05 for both). The cellular levels of CDX2 protein were significantly higher in cells incubated in choline and folate deficient media compared to the control media (1.78 [0.60] vs. 0.99 [0.42] pg/mg protein, p = 0.002); and compared to CDX2 levels in cells incubated in choline deficient but folate sufficient media (0.87 [0.13] pg/mg protein, p < 0.001) or in choline sufficient but folate deficient media (0.96 [0.16] pg/mg protein, p < 0.001). The levels of sFLT-1 and IGF1 in culture media and that of EOMES in HTR-8/SVneo cell lysates remained unchanged under all deficiency conditions. DISCUSSION: LRP2 and CDX2 are likely to be molecular targets for early choline and folate deficiencies in human trophoblast cells. The results should be confirmed in animal models and in other models of placental cells.

Effects of Survivorship Care Plans on patient reported outcomes in ovarian cancer during 2-year follow-up - The ROGY care trial.

OBJECTIVE: The aim of this study was to assess the long-term impact of an automatically generated Survivorship Care Plan (SCP) on patient reported outcomes in ovarian cancer in routine clinical practice. Outcome measures included satisfaction with information provision and care, illness perceptions and health care utilization. METHODS: In this pragmatic cluster randomized trial, twelve hospitals in the South of the Netherlands were randomized to 'SCP care' or 'usual care'. All newly diagnosed ovarian cancer patients in the 'SCP care' arm received an SCP that was automatically generated by the oncology provider, by clicking a button in the web-based Registrationsystem Oncological GYnecology (ROGY). Ovarian cancer patients (N=174, mean age 63.3, SD=11.4; all stages) completed questionnaires directly after initial treatment and after 6, 12 and 24months. RESULTS: First questionnaires were returned from 61 (67%) ovarian cancer patients in the 'SCP care' arm and 113 (72%) patients in the 'usual care' arm. In the 'SCP care' arm, 66% (N=41) of the patients reported receipt of an SCP. No overall differences were observed between the trial arms on satisfaction with information provision, satisfaction with care or health care utilization. Regarding illness perceptions, patients in the 'SCP care' arm had lower beliefs that the treatment would help to cure their disease (overall, 6.7 vs. 7.5, P<0.01). CONCLUSIONS: SCPs did not increase satisfaction with information provision or care in ovarian cancer patients. Our trial results suggest that ovarian cancer patients may not benefit from an SCP. TRIAL REGISTRATION: clinicaltrials.gov Identifier: NCT01185626.

Using mid infrared technology as new method for the determination of the dwell time of salivary substitutes on three dimensional gingiva models.

BACKGROUND: Many people suffer from dry mouth (xerostomia) due to radiotherapy treatment of head and neck cancer, diseases like Sjogren's syndrome or as adverse effects to prescribed medications. Salivary substitute products like gels or sprays are often used for treatment. Efficacy of those oral care products are regularly assessed by validated or even not validated questionnaires. To determine the adhesion effect over time more objectively a new and sensitive method was established. The following study was designed to assess the dwell time of different oral care products in vitro. METHOD: Two different types of surfaces were covered with oral care products and washed using a definite protocol with artificial saliva salt solution. First, oral care gels or oral care sprays were spread to a polystyrene surface of 2.25 cm(2), then onto cell based three-dimensional gingiva models. The surfaces were washed ten times with artificial saliva salt solution. The resulting washing solutions were examined using mid infrared spectroscopy in order to detect ingredients of the oral care products. RESULTS: All assessed oral care gels or oral care sprays and their components were detected very sensitive. Even traces of the products were detected in the eluent and thus enabled to differentiate the dwell times of the different products. In general, the dwell time of oral care gels on polystyrene or gingiva models was longer than that of oral care sprays. The use of gingiva models improved the differentiation between different products. CONCLUSIONS: MIR spectroscopy turned out to be a sensitive method to detect salivary substitutes. Differences between single components and different products can be detected. The described method is a simple, reliable and easy process to evaluate the dwell time of oral care products in vitro and thus a useful tool to design optimised salivary substitute products. ETHICS: This is an in vitro study. No ethics or consent was required for this study.

Impact of an Automatically Generated Cancer Survivorship Care Plan on Patient-Reported Outcomes in Routine Clinical Practice: Longitudinal Outcomes of a Pragmatic, Cluster Randomized Trial.

PURPOSE: This study was conducted to longitudinally assess the impact of an automatically generated survivorship care plan (SCP) on patient-reported outcomes in routine clinical practice. Primary outcomes were patient satisfaction with information and care. Secondary outcomes included illness perceptions and health care use. METHODS: Twelve hospitals were randomly assigned to SCP care or usual care in a pragmatic, cluster randomized trial. Newly diagnosed patients with endometrial cancer completed questionnaires after diagnosis (n = 221; 75% response), 6 months (n = 158), and 12 months (n = 147). An SCP application was built in the Web-based ROGY (Registration System Oncological Gynecology). By clicking the SCP button, a patient-tailored SCP was generated. RESULTS: In the SCP care arm, 74% of patients received an SCP. They reported receiving more information about their treatment (mean [M] = 57, standard deviation [SD] = 20 v M = 47, SD = 24; P = .03), other services (M = 35, SD = 22 v M = 25, SD = 22; P = .03), and different places of care (M = 27, SD = 25 v M = 23, SD = 26; P = .04) than the usual care arm (scales, 0 to 100). However, there were no differences regarding satisfaction with information or care. Patients in the SCP care arm experienced more symptoms (M = 3.3, SD = 2.0 v M = 2.6, SD = 1.6; P = .03), were more concerned about their illness (M = 4.4, SD = 2.3 v M = 3.9, SD = 2.1; P = .03), were more affected emotionally (M = 4.0, SD = 2.2 v M = 3.7, SD = 2.2; P = .046), and reported more cancer-related contact with their primary care physician (M = 1.8, SD = 2.0 v M = 1.1, SD = 0.9; P = .003) than those in the usual care arm (scale, 1 to 10). These effects did not differ over time. CONCLUSION: The present trial showed no evidence of a benefit of SCPs on satisfaction with information and care. Furthermore, SCPs increased patients' concerns, emotional impact, experienced symptoms, and the amount of cancer-related contact with the primary care physician. Whether this may ultimately lead to more empowered patients should be investigated further.

The impact of a cancer Survivorship Care Plan on gynecological cancer patient and health care provider reported outcomes (ROGY Care): study protocol for a pragmatic cluster randomized controlled trial.

BACKGROUND: There is a need for improvement of information provision and post-treatment care for cancer survivors. A Survivorship Care Plan (SCP) is recommended by the American Institute of Medicine and the Dutch Health Council, which is a summary of patients' course of treatment as a formal document, and includes recommendations for subsequent cancer surveillance, management of late effects, and strategies for health promotion. Until now, evidence on the effects of implementing the SCP in clinical practice is lacking. The rationale and study design of a pragmatic cluster randomized trial, aiming to assess the impact of SCP care in routine clinical practice, is presented. METHODS/DESIGN: A web-based patient registration system 'Registrationsystem Oncological GYnecology' (ROGY) is used by gynecologists in the South of the Netherlands since 2006. A personalized SCP can automatically be generated out of ROGY. In this pragmatic cluster randomized controlled trial, 12 hospitals are randomized to either 'usual care' or 'SCP care'. In patients with 'usual care', the gynecologist provides care as usual. In patients with 'SCP care', information about the tumor stage and treatment is personally discussed with the patient and a document is handed to the patient. Prospectively, all patients diagnosed with endometrial or ovarian cancer in the participating hospitals will be approached for study participation. Patients will complete questionnaires after surgery, and before additional treatment, and after 6, 12, 18 and 24 months. In addition, health care providers will be asked their opinion about implementation of SCP care. Primary outcome is defined as patient satisfaction with information provision and care. Secondary outcomes are illness perception, health-related quality of life, health care use, prevalence, course and referral rate of survivors with psychosocial distress, and health care providers' evaluation of SCP care. DISCUSSION: The ROGY Care trial will help to gain insight into the impact of SCP care on patient reported outcomes, and on the evaluation of cancer survivors and health care providers of the different elements of the SCP. Therefore, results will contribute to efforts to improve quality of care for cancer survivors. TRIAL REGISTRATION: Trial Registration: http://www.ClinicalTrials.gov. Identifier: NCT01185626 Medical Research Ethics Committee Reference Number: NL33429.008.10 Grant Reference Number: UVT2010-4743.

The application of normal, SV40 T-antigen-immortalised and tumour-derived oral keratinocytes, under serum-free conditions, to the study of the probability of cancer progression as a result of environmental exposure to chemicals.

In vitro models are currently not considered to be suitable replacements for animals in experiments to assess the multiple factors that underlie the development of cancer as a result of environmental exposure to chemicals. An evaluation was conducted on the potential use of normal keratinocytes, the SV40 T-antigen-immortalised keratinocyte cell line, SVpgC2a, and the carcinoma cell line, SqCC/Y1, alone and in combination, and under standardised serum-free culture conditions, to study oral cancer progression. In addition, features considered to be central to cancer development as a result of environmental exposure to chemicals, were analysed. Genomic expression, and enzymatic and functional data from the cell lines reflected many aspects of the transition of normal tissue epithelium, via dysplasia, to full malignancy. The composite cell line model develops aberrances in proliferation, terminal differentiation and apoptosis, in a similar manner to oral cancer progression in vivo. Transcript and protein profiling links aberrations in multiple gene ontologies, molecular networks and tumour biomarker genes (some proposed previously, and some new) in oral carcinoma development. Typical specific changes include the loss of tumour-suppressor p53 function and of sensitivity to retinoids. Environmental agents associated with the aetiology of oral cancer differ in their requirements for metabolic activation, and cause toxic effects to cells in both the normal and the transformed states. The results suggest that the model might be useful for studies on the sensitivity of cells to chemicals at different stages of cancer progression, including many aspects of the integrated roles of cytotoxicity and genotoxicity. Overall, the properties of the SVpgC2a and SqCC/Y1 cell lines, relative to normal epithelial cells in monolayer or organotypic culture, support their potential applicability to mechanistic studies on cancer risk factors, including, in particular, the definition of critical toxicity effects and dose-effect relationships.

In vitro reproduction of clinical hallmarks of eczematous dermatitis in organotypic skin models.

Dermatitis is a group of highly pruritic chronic inflammatory skin diseases which represents a major public-health problem worldwide. The prevalence of dermatitis has increased in recent years affecting up to 20% of the general population. Acute skin lesions are characterized by extensive degrees of intercellular edema of the epidermis (spongiosis) and a marked perivenular inflammatory cell infiltrate in the dermis. Keratinocytes within eczematous lesions exhibit a modified expression of proinflammatory cytokines, chemokines and cell-surface molecules. The pathophysiological puzzle of dermatitis is far from being elucidated completely, but skin infiltration of activated memory/effector T cells are thought to play the pivotal role in the pathogeneses. The aim of this study was the set-up of organotypic models mimicking the symptoms of eczematous dermatitis to provide a tool for therapeutic research in vitro. Therefore activated T cells (ATs) were integrated in organotypic skin and epidermis equivalents (SE, EE). These models enabled the reproduction of several clinical hallmarks of eczematous dermatitis: (1) T cells induce keratinocyte apoptosis, which leads to a reduced expression of the adhesion molecule E-cadherin (E-cad) and disruption of the epidermal barrier. (2) Expression of intercellular adhesion molecule-1 (ICAM-1) allows the attachment of leukocytes to epidermal cells. (3) Upregulation of neurotrophin-4 (NT-4) in the epidermis is thought to mediate pruritus in lesions by supporting nerve outgrowth. (4) Elevated levels of pro-inflammatory cytokines (IL-1alpha and IL-6) and chemokines (IL-8, IP-10, TARC, MCP-1, RANTES and eotaxin) amplify the inflammatory response and lead to an influx of secondary immunocells into the skin. The therapeutics dexamethasone and FK506 markedly reduce cytokines/chemokines production and epidermal damaging in these models. These data underline that activated memory/effector T cells induce eczematous changes in this HaCaT cell based organotypic skin equivalent. Furthermore it can be concluded that these models make it possible to investigate targets of therapeutics in skin.

Pilot study on the effect of parenteral vitamin E on ischemia and reperfusion induced liver injury: a double blind, randomized, placebo-controlled trial.

BACKGROUND AND AIMS: Liver surgery usually involves ischemia and reperfusion (I/R) which results in oxidative stress and cell damage. The administration of antioxidants should diminish or prevent this damage. The purpose of this study was to investigate the effect of the antioxidant vitamin E on I/R injury. METHODS: We carried out a placebo-controlled double-blind study on 68 patients undergoing elective, tumor-related, partial liver resection. 47 patients were qualified for the per protocol population based evaluation. The patients were randomly assigned to two groups. The day before surgery one group received three infusions containing vitamin E (600 IU=540 mg vitamin E emulsion). The other group received three infusions of placebo. RESULTS: Length of stay in the intensive care unit (ICU) was significantly shorter in the verum group than in the placebo group (P<0.05). There were signs of improvement for AUC AST (P<0.05), ALT and GLDH in the verum group after surgery. Serum vitamin E concentration increased after administration of vitamin E infusion and declined in both treatment groups after surgery (P<0.01). In the verum group vitamin E deficiency was prevented while vitamin E concentration remained low in the placebo group (P<0.01). CONCLUSIONS: The findings from this study indicate that preoperative administration of vitamin E is safe and that this treatment may have beneficial effects by reducing the impact of I/R injury in liver surgery.

3,4-Dihydroxymandelic acid, a noradrenalin metabolite with powerful antioxidative potential.

The decarboxylated noradrenaline metabolite 3,4-dihydroxymandelic acid [DHMA, 2-(3,4-dihydroxyphenyl)-2-hydroxyacetic acid] occurs in different mammalian tissues, especially in the heart. To elucidate the physiological function of DHMA, the antioxidative and radical scavenging activity was determined by physicochemical and cell-based test systems. In the 2,2-diphenyl-1-picrylhydrazyl assay it shows a 4-fold higher radical scavenging activity compared to the standard antioxidants ascorbic acid, tocopherol, and butylated hydroxytoluene. DHMA is also a very potent superoxide radical scavenger and shows a 5-fold smaller IC(50) value compared to standard ascorbic acid. Again, in most cases the antioxidative power of DHMA against bulk lipid oxidation determined by accelerated autoxidation of oils is much higher than for the standard antioxidants. In soybean oil and squalene a DHMA/alpha-tocopherol mixture (1:1 w/w) shows a synergistic effect. Last but not least, 0.001 and 0.0005% levels of DHMA protect human primary fibroblasts against H(2)O(2)-induced oxidative stress as determined by the 2',7'-dichlorofluorescein assay.

Expression of retinoid-related genes in serum-free cultures of normal, immortalized and malignant human oral keratinocytes.

Retinoids are used in the clinical treatment of oral squamous carcinoma, including both early and late stages. Inter-individual variation in responsiveness, including a common insensitivity of advanced stages, suggest that changes in retinoid-related functions might characterize tumor development. To investigate a genetic basis for this hypothesis, an in vitro multi-step model of carcinogenesis involving normal (NOK), SV40 T antigen-immortalized (SVpgC2a) and malignant (SqCC/Y1) oral keratinocytes was analysed under identical culture conditions using micro-array technique (Affymetrix HG_U95A chip) for expression of 52 genes related to retinoid metabolism and actions. The variable detection of between 22-26 transcripts in the cell lines, involving binding/transport factors, receptors, transcriptional activators/repressors and responsive genes, indicated specificity in regards to the expression of known retinoid-related genes in oral keratinocytes. The transformed cell lines variably exhibited differences as compared to NOK, i.e., lower transcript levels for cellular retinol binding protein, the cellular retinoic acid binding protein II (CRABP II) and retinoic acid receptor gamma, whereas in contrast, the levels of CRABP I were higher. Transcripts for proteins interacting with nuclear retinoid receptors were similarly expressed among the cell types, whereas transcripts for retinoid-metabolizing enzymes were generally not detected. Finally, transcripts of retinoid-responsive genes, including RARRES3, RI58, NN8-4AG and midkine, were variably expressed. The overall results imply selective expression of retinoid-related functions in normal and transformed keratinocytes, and that cell transformation can impair the capacity for binding and storage of retinol as well as retinoic acid-mediated signalling. These multiple alterations are consistent with possible retinoid insensitivity during oral carcinogenesis.