Chromatin modifier developmental pluripotency associated factor 4 (DPPA4) is a candidate gene for alcohol-induced developmental disorders.

BACKGROUND: Prenatal alcohol exposure (PAE) affects embryonic development, causing a variable fetal alcohol spectrum disorder (FASD) phenotype with neuronal disorders and birth defects. We hypothesize that early alcohol-induced epigenetic changes disrupt the accurate developmental programming of embryo and consequently cause the complex phenotype of developmental disorders. To explore the etiology of FASD, we collected unique biological samples of 80 severely alcohol-exposed and 100 control newborns at birth. METHODS: We performed genome-wide DNA methylation (DNAm) and gene expression analyses of placentas by using microarrays (EPIC, Illumina) and mRNA sequencing, respectively. To test the manifestation of observed PAE-associated DNAm changes in embryonic tissues as well as potential biomarkers for PAE, we examined if the changes can be detected also in white blood cells or buccal epithelial cells of the same newborns by EpiTYPER. To explore the early effects of alcohol on extraembryonic placental tissue, we selected 27 newborns whose mothers had consumed alcohol up to gestational week 7 at maximum to the separate analyses. Furthermore, to explore the effects of early alcohol exposure on embryonic cells, human embryonic stem cells (hESCs) as well as hESCs during differentiation into endodermal, mesodermal, and ectodermal cells were exposed to alcohol in vitro. RESULTS: DPPA4, FOXP2, and TACR3 with significantly decreased DNAm were discovered-particularly the regulatory region of DPPA4 in the early alcohol-exposed placentas. When hESCs were exposed to alcohol in vitro, significantly altered regulation of DPPA2, a closely linked heterodimer of DPPA4, was observed. While the regulatory region of DPPA4 was unmethylated in both control and alcohol-exposed hESCs, alcohol-induced decreased DNAm similar to placenta was seen in in vitro differentiated mesodermal and ectodermal cells. Furthermore, common genes with alcohol-associated DNAm changes in placenta and hESCs were linked exclusively to the neurodevelopmental pathways in the enrichment analysis, which emphasizes the value of placental tissue when analyzing the effects of prenatal environment on human development. CONCLUSIONS: Our study shows the effects of early alcohol exposure on human embryonic and extraembryonic cells, introduces candidate genes for alcohol-induced developmental disorders, and reveals potential biomarkers for prenatal alcohol exposure.

Health care utilisation among older people with Down syndrome compared to specific medical guidelines for health surveillance: a Swedish national register study.

BACKGROUND: Specific medical guidelines for health surveillance exist for people with Down syndrome (DS) since 25 years but knowledge of adherence to the guidelines is lacking. The guidelines were developed to avoid unnecessary suffering from preventable conditions. The aims of the study were to investigate 1) planned health care visits in relation to the co-morbidities described in specific medical guidelines as a measure of adherence, 2) unplanned health care visits as a measure of potentially unmet health care needs and 3) gender differences in health care utilisation among older people with DS. METHODS: This register-based study includes people with DS (n = 472) from a Swedish national cohort of people with intellectual disability (n = 7936), aged 55 years or more, and with at least one support according to the disability law, in 2012. Data on inpatient and outpatient specialist health care utilisation were collected from the National Patient Register for 2002-2012. RESULTS: A total of 3854 inpatient and outpatient specialist health care visits were recorded during the 11 years, of which 54.6% (n = 2103) were planned, 44.0% (n = 1695) unplanned and 1.4% (n = 56) lacked information. More than half of the visits, 67.0% (n = 2582) were outpatient health care thus inpatient 33% (n = 1272). Most planned visits (29.4%, n = 618) were to an ophthalmology clinic, and most unplanned visits to an internal medicine clinic (36.6%, n = 621). The most common cause for planned visits was cataract, found at least once for 32.8% in this cohort, followed by arthrosis (8.9%), epilepsy (8.9%) and dementia (6.6%). Pneumonia, pain, fractures and epilepsy each accounted for at least one unplanned visit for approximately one-fourth of the population (27.1, 26.9, 26.3 and 19.7% respectively). Men and women had similar numbers of unplanned visits. However, women were more likely to have visits for epilepsy or fractures, and men more likely for pneumonia. CONCLUSIONS: Increased awareness of existing specific medical guidelines for people with DS is vital for preventive measures. The relatively few planned health care visits according to the medical guidelines together with a high number of unplanned visits caused by conditions which potentially can be prevented suggest a need of improved adherence to medical guidelines.

Integrative radiomics expression predicts molecular subtypes of primary clear cell renal cell carcinoma.

AIM: To identify combined positron-emission tomography (PET)/magnetic resonance imaging (MRI)-based radiomics as a surrogate biomarker of intratumour disease risk for molecular subtype ccA and ccB in patients with primary clear cell renal cell carcinoma (ccRCC). MATERIALS AND METHODS: PET/MRI data were analysed retrospectively from eight patients. One hundred and sixty-eight radiomics features for each tumour sampling based on the regionally sampled tumours with 23 specimens were extracted. Sparse partial least squares discriminant analysis (SPLS-DA) was applied to feature screening on high-throughput radiomics features and project the selected features to low-dimensional intrinsic latent components as radiomics signatures. In addition, multilevel omics datasets were leveraged to explore the complementing information and elevate the discriminative ability. RESULTS: The correct classification rate (CCR) for molecular subtype classification by SPLS-DA using only radiomics features was 86.96% with permutation test p=7×10-4. When multi-omics datasets including mRNA, microvascular density, and clinical parameters from each specimen were combined with radiomics features to refine the model of SPLS-DA, the best CCR was 95.65% with permutation test, p<10-4; however, even in the case of generating the classification based on transcription features, which is the reference standard, there is roughly 10% classification ambiguity. Thus, this classification level (86.96-95.65%) of the proposed method represents the discriminating level that is consistent with reality. CONCLUSION: Featured with high accuracy, an integrated multi-omics model of PET/MRI-based radiomics could be the first non-invasive investigation for disease risk stratification and guidance of treatment in patients with primary ccRCC.

High prevalence of diabetes mellitus, hypertension and obesity among persons with a recorded diagnosis of intellectual disability or autism spectrum disorder.

BACKGROUND: Obesity and lack of physical activity are frequently reported in persons with intellectual disability (ID) or autism spectrum disorder (ASD). We hypothesised a higher prevalence of diabetes and hypertension in this population. METHOD: We used administrative data for all primary and specialist outpatient and inpatient healthcare consultations for people with at least one recorded diagnosis of diabetes mellitus, hypertension or obesity from 1998 to 2015. Data were drawn from the central administrative database for Stockholm County, Sweden. It was not possible to separate data for type 1 and type 2 diabetes. We stratified 26 988 individuals with IDs or ASD into three groups, with Down syndrome treated separately, and compared these groups with 1 996 140 people from the general population. RESULTS: Compared with the general population, men and women with ID/ASD had 1.6-3.4-fold higher age-adjusted odds of having a registered diagnosis of obesity or diabetes mellitus, with the exception of diabetes among men with Down syndrome. A registered diagnosis of hypertension was only more common among men with ID/ASD than in the general population. CONCLUSIONS: Diabetes and blood pressure health screening, along with efforts to prevent development of obesity already in childhood, are necessary for individuals with IDs and ASD. We believe that there is a need for adapted community-based health promotion programmes to ensure more equitable health for these populations.

Ror2, a developmentally regulated kinase, promotes tumor growth potential in renal cell carcinoma.

Inappropriate kinase expression and subsequent promiscuous activity defines the transformation of many solid tumors including renal cell carcinoma (RCC). Thus, the expression of novel tumor-associated kinases has the potential to dramatically shape tumor cell behavior. Further, identifying tumor-associated kinases can lend insight into patterns of tumor growth and characteristics. Here, we report the identification of the RTK-like orphan receptor 2 (Ror2), a new tumor-associated kinase in RCC cell lines and primary tumors. Ror2 is an orphan receptor tyrosine kinase with physiological expression normally seen in the embryonic kidney. However, in RCC, Ror2 expression correlated with expression of genes involved at the extracellular matrix, including Twist and matrix metalloprotease-2 (MMP2). Expression of MMP2 in RCC cells was suppressed by Ror2 knockdown, placing Ror2 as a mediator of MMP2 regulation in RCC and a potential regulator of extracellular matrix remodeling. The suppression of Ror2 not only inhibited cell migration, but also inhibited anchorage-independent growth in soft agar and growth in an orthotopic xenograft model. These findings suggest a novel pathway of tumor-promoting activity by Ror2 within a subset of renal carcinomas, with significant implications for unraveling the tumorigenesis of RCC.

Computer based safety training: an investigation of methods.

BACKGROUND: Computer based methods are increasingly being used for training workers, although our understanding of how to structure this training has not kept pace with the changing abilities of computers. Information on a computer can be presented in many different ways and the style of presentation can greatly affect learning outcomes and the effectiveness of the learning intervention. Many questions about how adults learn from different types of presentations and which methods best support learning remain unanswered. AIMS: To determine if computer based methods, which have been shown to be effective on younger students, can also be an effective method for older workers in occupational health and safety training. METHODS: Three versions of a computer based respirator training module were developed and presented to manufacturing workers: one consisting of text only; one with text, pictures, and animation; and one with narration, pictures, and animation. After instruction, participants were given two tests: a multiple choice test measuring low level, rote learning; and a transfer test measuring higher level learning. RESULTS: Participants receiving the concurrent narration with pictures and animation scored significantly higher on the transfer test than did workers receiving the other two types of instruction. There were no significant differences between groups on the multiple choice test. CONCLUSIONS: Narration with pictures and text may be a more effective method for training workers about respirator safety than other popular methods of computer based training. Further study is needed to determine the conditions for the effective use of this technology.

Repression of thermotolerance in Dunning R3327 prostate carcinoma cells by 2-deoxy-glucose.

The transient addition of the cytosolic energy depletor 2-deoxy-glucose to cultures of rat prostate carcinoma cells blunted the induction of Hsp70 protein following exposure to elevated temperatures in a manner that appeared to parallel its effects on energy metabolism. While the reduction in stress-induced heat-shock protein expression by treatment with 2-deoxy-glucose had no effects on the acute loss of cellular viability after exposure to heat, the acquisition of thermotolerance in response to a conditioning stimulus was specifically repressed. Therefore, 2-deoxy-glucose will be a useful tool in the investigation of mechanisms that mediate immediate versus chronic responses to cellular stress, including the specific roles played by members of the heat-shock protein family of proteins. These results might have important implications in the design of protocols for the hyperthermic treatment of tumours.

Estramustine phosphate enhances the effects of hyperthermia and induces the small heat shock protein HSP27 in the human prostate carcinoma cell line PC-3.

The antimicrotubule drug estramustine phosphate (EMP) has been shown to sensitize prostate carcinoma cells to radiation via synchronization at the G2/M phase of the cell cycle. This synchronization may also render cells more sensitive to hyperthermia, providing a rationale for multimodal treatment approaches. We have investigated the effects of EMP and hyperthermia, as well as the regulation of heat shock proteins (HSP) in the PC-3 prostatic carcinoma cell line. Cells were incubated with four doses of EMP for 48 h followed by a 1-h hyperthermia treatment ranging from 41 degrees C to 44 degrees C. Cell cycle distribution at the end of the EMP incubation was investigated by flow cytometry. Cytotoxicity was assessed by colony formation assays. HSP accumulation was investigated by Western immunoblotting. Doses of 1, 5, 10 and 15 microM EMP synchronized 27, 28, 46, and 68% of PC-3 cells at G2/M. With 5, 10 and 15 microM, a sensitizing effect of EMP was assessed at hyperthermic temperatures of 42, 43 and 44 degrees C. EMP did not alter the expression of HSP72, but substantially induced the synthesis of HSP27 in PC-3 cells. Our data show that EMP sensitizes PC-3 cells to hyperthermia induced cytotoxicity. This observation supports the rationale for multimodal treatment approaches in locally advanced prostate cancer.

Effect of heat stress on LPS-induced febrile response in D-galactosamine-sensitized rats.

We have previously reported that heat conditioning augments lipopolysaccharide (LPS)-induced fever in rats, which is accompanied by an accumulation of heat shock protein (HSP) in the liver and the reduction of the plasma level of tumor necrosis factor (TNF-alpha) (Kluger MJ, Rudolph K, Soszynski D, Conn CA, Leon LR, Kozak W, Wallen ES, and Moseley PL. Am J Physiol Regulatory Integrative Comp Physiol 273: R858-R863, 1997). In the present study we have tested whether inhibition of protein synthesis in the liver can reduce the effect of this heat conditioning on the LPS-induced febrile response in the rat. D-galactosamine (D-gal) was used to selectively inhibit liver protein synthesis. D-gal (500 mg/kg) or PBS as control was administered intraperitoneally 1 h before heat stress. LPS (50 microg/kg ip) was injected 24 h post-heat exposure. Treatment with D-gal blunted the febrile response to LPS. Moreover, heat-conditioned rats treated first with D-gal and subsequently with LPS demonstrated a profound fall in core temperature 10--18 h post-LPS. A significant increase of serum TNF-alpha accompanied this effect of D-gal on fever. Heat-conditioned animals receiving D-gal showed an inhibition in inducible HSP-70 in the liver. These data support the role of hepatic function in modulating the febrile response to LPS.

Brefeldin A induces p53-independent apoptosis in primary cultures of human prostatic cancer cells.

PURPOSE: The objective of this study was to investigate growth-inhibitory and apoptotic activity of the experimental antitumor drug, brefeldin A (BFA), on primary cultures of human epithelial cells derived from prostatic adenocarcinomas. MATERIALS AND METHODS: Clonal assays were performed to evaluate the effects of BFA on growth of prostatic cancer cell strains. Loss of cell viability in response to BFA was assessed by trypan blue exclusion. Induction of apoptosis by BFA was evaluated by morphologic criteria, electrophoretic assay of DNA fragmentation, and a cell death ELISA. Immunoblots were used to monitor p53 and pRB expression in response to BFA. RESULTS: BFA was growth-inhibitory at a half-maximal concentration of 5 ng./ml. (18 nM). Morphological manifestations of apoptosis were evident by 24 hours of treatment. Cell viability declined and the cell death ELISA indicated an 18-fold increase in apoptosis in BFA-treated versus untreated cells at 48 hours. DNA fragmentation was also seen at 48 hours. Levels of p53 were not altered by BFA, but pRB was maintained in a hypophosphorylated state by BFA treatment. CONCLUSIONS: BFA is a potent inducer of apoptosis in prostatic cancer cells via a p53-independent mechanism. Cells derived from low-grade as well as high-grade cancers responded similarly to BFA. Since p53-mediated pathways of apoptosis may frequently be abrogated in prostatic cancer cells, agents such as BFA that induce p53-independent cell death may be promising candidates for chemotherapeutic agents.

Effects of bright light on age-related changes in the locomotor activity of Syrian hamsters.

Syrian hamsters display age-related changes in the expression of circadian rhythms and in responsiveness of the circadian system to photic and non-photic stimuli. This study characterized the effects of age on the locomotor activity rhythm of middle-aged and old hamsters and evaluated the effects of strengthening the entraining light signal. Compared with young (4.5 mo) animals, middle-aged (11.25 mo) and old (16 mo) animals displayed increased daily bouts of activity (P < 0.001) and reduced total daily activity and activity rhythm amplitude (P < 0.05) in 14:10-h light-dark cycles. After the light intensity was increased from 300 to 1,500 lx during the light cycle, middle-aged hamsters demonstrated decreased daily activity bouts (P < 0.05) and increased total daily activity (P < or = 0.01) and activity rhythm amplitude (P < or = 0.001) compared with controls maintained in 300 lx. The pattern of changes in the activity rhythm of old experimental animals was similar to trends observed in middle-aged experimental hamsters, although not as robust. Thus age-related changes in the activity rhythm are occurring by middle age in hamsters, and the provision of stronger entraining signals may lead to more stable circadian organization.

Effects of combined treatment of chemotherapeutics and hyperthermia on survival and the regulation of heat shock proteins in Dunning R3327 prostate carcinoma cells.

BACKGROUND: Hyperthermia can enhance the clinical response of chemotherapeutic agents in prostate cancer, but optimal sequencing of this combination therapy needs to be developed. Given the role of heat shock proteins (HSPs) in the development of resistance (thermotolerance) to subsequent hyperthermic stresses as well as to certain chemotherapeutics, the study of HSP regulation is important in the establishment of effective schedules in multimodal treatment strategies. METHODS: In this study we evaluated the effects of the chemotherapeutic agents cisplatin, 5-fluorouracil, and adriamycin in combination with hyperthermia. (43 degrees C, 1 h) on clonogenic survival and inducible HSP70 regulation in Dunning rat adenocarcinoma of the prostate. HSP70 was analyzed by Western blot and by measuring beta-galactosidase produced by cells stably transfected with a gene construct containing the E. coli beta-galactosidase gene driven by the Drosophila HSP70 promoter. RESULTS: Colony formation assays revealed a sensitizing effect of hyperthermia when simultaneously combined with each chemotherapeutic agent, resulting in a potentiated cytotoxicity compared to subsequenced treatments. Thermotolerant cells showed a significantly better survival when treated with adriamycin alone, but also when each chemotherapeutic agent was combined with hyperthermia. This enhanced survival was correlated with inducible HSP70 accumulation. The chemotherapeutics modified the HSP70 promoter activation induced by hyperthermia, suggesting changes in the development of cellular thermotolerance. CONCLUSIONS: Our data reveal synergistic cytotoxic effects of the synchronous application of chemotherapeutic agents and hyperthermia on this model of prostate cancer. Furthermore, they demonstrate that the induction of HSPs in thermotolerant cells, as measured by HSP70 induction, results in a modulation the chemotherapeutic-mediated cytotoxicity. Therefore, HSP70 is a useful marker of cellular resistance in multimodal approaches combining hyperthermia and chemotherapeutic agents in the treatment of locally advanced prostate carcinoma.

Oxidants differentially regulate the heat shock response.

Cells, animals, and humans respond to hyperthermia through the synthesis of a family of proteins termed heat shock proteins (HSPs). Because hyperthermic stress may also result in mitochondrial uncoupling and the generation of reactive oxygen species, we wondered whether oxidant stress was sufficient to increase cellular levels of HSP70. HSP70 was detected in cells heated or treated with menadione but not in those treated with hydrogen peroxide or xanthine/xanthine oxidase. We speculate that oxidant stress from menadione exposure is qualitatively different from exposure from hydrogen peroxide or xanthine/xanthine oxidase.

Effect of heat stress on LPS-induced fever and tumor necrosis factor.

Exposure to heat stress leads to both short-term and long-term effects on morbidity. Male rats were exposed to a high ambient temperature of 40 degrees C, which resulted in biotelemetered core body temperature rising to approximately 42 degrees C. This treatment led to a marked enhancement in lipopolysaccharide (LPS)-induced fever at 24 h after exposure to heat stress. The increase in fever was accompanied by a significant suppression in the circulating concentration of tumor necrosis factor. Heat-shock protein-70 measured in liver was elevated by the heat exposure (but not further elevated by the injection of LPS). An enhanced fever to LPS and other inflammatory stimuli found in heat-stressed human subjects could explain the apparent increase in susceptibility to disease.

Aging and photoperiod affect entrainment and quantitative aspects of locomotor behavior in Syrian hamsters.

Aging affects the regulation of diurnal and circadian rhythmicity. We tested the hypothesis that the age-related difference in the phase angle of entrainment of the locomotor activity rhythm to a light-dark (LD) cycle would be greater under LD 6:18 than LD 14:10. We also analyzed changes in quantitative aspects of wheel-running behavior according to age group. Young (9-wk-old), middle-aged (11- to 12-mo-old), and old (15- to 17-mo-old) male golden hamsters were entrained to a 14:10 LD cycle followed by re-entrainment to a 6:18 LD cycle. Fourteen days after the start of locomotor recording in LD 14:10 and again after 27 days in LD 6:18, the phase of activity onset, the total number of wheel revolutions performed per day, the peak intensity of wheel-running activity, the duration of the active period, and the level of fragmentation of locomotor activity were quantitated. We also studied the temporal distribution of the largest bout of wheel-running activity among the age groups in both photoperiods. Short days induced testicular regression at a similar rate among young, middle-aged, and old hamsters. The data are discussed in terms of the effects of age on overall circadian organization in the seasonally changing environment.

A computer program to aid in visual concept development in dentistry.

Beginning dental students normally receive their first exposure to the study of tooth forms (morphology) through a dental anatomy laboratory course in which they are required to reproduce tooth morphology, usually with wax. The fabrication of a tooth in wax requires proper visual recognition skills and fine eye-hand coordination. Many students struggle with one or both of these. A computer program, designed to teach recognition concepts, was delivered to three groups of beginning freshman dental students in conjunction with their dental anatomy laboratory course while a group of their classmates served as the controls. This study investigated (1) instructional design and interface improvement and (2) the best method to implement the computer program. Experimental and control groups all received normal daily critiques of their course project work. After completion of the computer program, all groups were tested with a recognition-based examination as well as with a practical examination, requiring the reproduction of a tooth in wax. All experimental groups scored better than the control group on both examinations. Results indicated that computer-based instruction may be a useful means to foster visual concept development. An expanded program, using better graphics, animation and movies is currently under development.

Aging alters the serotonergic modulation of light-induced phase advances in golden hamsters.

Recent findings have raised the possibility that some of the age-related changes in the circadian system and the response of the circadian pacemaker to environmental stimuli may involve central serotonergic mechanisms. The present study compared the effects ofpretreatment with the serotonin agonist 8-hydroxy-2(di-n-propylamino)tetralin (5 mg/kg ip) on the magnitude of light-induced phase advances in young (2-4 mo) and old (18-20 mo) golden hamsters. The ability of this serotonin agonist to attenuate the photic phase resetting of circadian locomotor rhythmicity in young animals was decreased by 46% in old hamsters (P < 0.05). These results suggest that deficits in the mechanisms for serotonergic control of circadian function may interfere with the optimal adaptation of the senescent organism to its temporal environment.

Beta-galactosidase as a marker of HSP70 promoter induction in Dunning R3327 prostate carcinoma cells.

Hyperthermia is known to improve the response of tumors to radiation or chemotherapeutic treatment when combined in multimodal strategies. The cellular response to hyperthermia is associated with the synthesis of heat shock proteins (HSP). To study the stress response in prostate cancer we have developed a clone of Dunning R3327 rat prostate carcinoma cells stably transfected with a gene construct containing the E. coli beta-galactosidase gene driven by the Drosophila HSP70 promoter. The measurement of beta-galactosidase serves as a rapid and semiquantitative assay of HSP70 gene activation. The Dunning cell clone showed evidence of incorporation of the HSP70/beta-galactosidase construct within the genomic DNA by Southern blot analysis. When compared to mock-transfected control cells, the clone showed minimal baseline beta-galactosidase activity, which significantly increased following a hyperthermic stress. The time course of beta-galactosidase elevation following heat stress paralleled the time course of cellular HSP70 elevation by Western blot analysis. These stably transfected Dunning R3327 cells may provide a useful tool to study the effects of hyperthermia, radiation, and chemotherapeutic agents on the cellular stress response and in the establishment of HSP70 as a marker of cellular resistance in the multimodal treatment of prostate cancer.