Salivary concentrations of IL-8 and IL-1ra after HIIT and MICT in young, healthy adults: A randomized exercise study.

The purpose of this study was to determine whether six weeks of high intensity interval training (HIIT) would lead to greater changes in resting concentrations of salivary IL-8 and IL-1ra than moderate intensity continuous training (MICT) in young, healthy adults, and to determine whether changes in IL-8 and IL-1ra after six weeks of either HIIT or MICT were associated with changes in maximal exercise capacity (VO2max). Participants were randomly assigned to 6 weeks of HIIT (n = 12) or MICT (n = 11), matched for workload. Saliva samples were collected at the beginning (T1) and end (T2) of the intervention, and analyzed for IL-8 and IL-1ra. Participants in both groups had significant improvements in VO2max; there were no group differences in improvements. A greater reduction in IL-8 was observed in the MICT group when compared to the HIIT group (HIIT median: -9.5; MICT median: -82.3 pg/µg of protein; U = 11.5, p < 0.001). When combining the HIIT and MICT group, there were significant reductions in IL-8 from T1 to T2. There was no correlation between changes in IL-8 (r < 0.00) or IL-1ra (r = -0.013) with changes in VO2max. In conclusion, 6 weeks of exercise training leads to a reduction in IL-8; MICT may lead to greater reductions when compared to HIIT. Future research examining longer intervention periods is needed to further elucidate the effects of HIIT and MICT on different pro and anti-inflammatory cytokines.

Organic osmolytes increase expression of specific tight junction proteins in skin and alter barrier function in keratinocytes.

BACKGROUND: The epidermal barrier is important for water conservation, failure of which is evident in dry-skin conditions. Barrier function is fulfilled by the stratum corneum, tight junctions (TJs, which control extracellular water) and keratinocyte mechanisms, such as organic osmolyte transport, which regulate intracellular water homeostasis. Organic osmolyte transport by keratinocytes is largely unexplored and nothing is known regarding how cellular and extracellular mechanisms of water conservation may interact. OBJECTIVES: We aimed to characterize osmolyte transporters in skin and keratinocytes, and, using transporter inhibitors, to investigate whether osmolytes can modify TJs. Such modification would suggest a possible link between intracellular and extracellular mechanisms of water regulation in skin. METHODS: Immunostaining and quantitative polymerase chain reaction of organic osmolyte-treated organ-cultured skin were used to identify changes to organic osmolyte transporters, and TJ protein and gene expression. TJ functional assays were performed on organic osmolyte-treated primary human keratinocytes in culture. RESULTS: Immunostaining demonstrated the expression of transporters for betaine, taurine and myo-inositol in transporter-specific patterns. Treatment of human skin with either betaine or taurine increased the expression of claudin-1, claudin-4 and occludin. Osmolyte transporter inhibition abolished this response. Betaine and taurine increased TJ function in primary human keratinocytes in vitro. CONCLUSIONS: Treatment of skin with organic osmolytes modulates TJ structure and function, which could contribute to the epidermal barrier. This emphasizes a role for organic osmolytes beyond the maintenance of intracellular osmolarity. This could be harnessed to enhance topical therapies for diseases characterized by skin barrier dysfunction.

Asthma prevalence and control levels among Special Olympics athletes, and asthma-related knowledge of their coaches.

BACKGROUND: The prevalence of asthma among athletes with intellectual disabilities, and the asthma knowledge levels of their coaches, is unknown. METHODS: Special Olympics Canada athletes completed a demographic questionnaire (n = 208). Athletes who identified as having ever or current asthma completed the Asthma Control Questionnaire and the Mini Asthma Quality of Life Questionnaire and were measured for height, weight and lung function (n = 73). National level coaches (n = 27) completed a questionnaire pertaining to asthma knowledge. RESULTS: The prevalence of ever and current asthma were 35.5% (n = 73) and 21.1% (n = 44), respectively. Athletes with asthma reported that they had inadequately controlled asthma, but good quality of life. Coaches correctly answered 43% true/false questions on the survey, indicating suboptimal asthma knowledge. CONCLUSIONS: Athletes with intellectual disabilities appear to have a greater prevalence of asthma than the general population; however, coaches of these athletes appear to have limited knowledge pertaining to asthma and exercise-induced asthma.

A rationale for biopsying embryos reaching the morula stage on Day 6 in women undergoing preimplantation genetic testing for aneuploidy.

STUDY QUESTION: Is there a benefit to assessing ploidy in delayed embryos reaching the morula stage on Day 6 of development? SUMMARY ANSWER: Day-6 morulae should be considered for biopsy in women <40 years old undergoing preimplantation genetic testing for aneuploidy (PGT-A) because they are associated with acceptable, albeit reduced, euploidy and implantation rates (IRs). WHAT IS KNOWN ALREADY: Embryo development and morphology have been shown to correlate with aneuploidy and pregnancy rates. During PGT-A cycles, embryos are biopsied if they reach the blastocyst stage by Day 5 or 6, whereas slow-developing embryos are typically deselected and discarded. Determining the viability of slow-developing embryos is particularly relevant for women undergoing PGT-A who have diminished ovarian reserve and a relatively low blastocyst yield. STUDY DESIGN, SIZE, DURATION: This is a retrospective cohort study that was performed at an academic medical center. Patients who underwent IVF with PGT-A were reviewed for inclusion. PARTICIPANTS/MATERIALS, SETTING, METHODS: A total of 1615 cycles were reviewed. All cycles which involved a biopsy of a cavitating or compacted morula on Day 6 were included (n = 763). PGT-A was performed using array comparative genomic hybridization. The aneuploidy and implantation of morulae were compared to those of blastocysts originating from the same couples. MAIN RESULTS AND THE ROLE OF CHANCE: The study included 763 cycles in which 1260 morulae and 3014 blastocysts were biopsied. Women were divided into four age groups (<35, 35-37, 38-39 and ≥40 years): the prevalence of aneuploidy was consistently lower among blastocysts (40.3, 50.8, 56 and 78.3%, respectively) than among compacted morulae (68.7, 75.5, 88.9 and 98.1%, respectively) and cavitating morulae (57, 66.4, 81 and 91.6%, respectively) throughout the different age groups (P < 0.001). Of note, the majority of compacted morulae (98.1%) and cavitating morulae (91.6%) were aneuploid in women aged ≥40 years. Compacted and cavitating morulae had significantly higher rates of complex aneuploidy, which involves ≥3 chromosomes, compared with blastocysts (P < 0.001). Furthermore, euploid morulae were associated with a significantly lower IR (28.2 versus 54.6%; P = 0.002) and live birth rate (23.1 versus 55.0%; P = 0.001) compared to euploid blastocysts. LIMITATIONS REASONS FOR CAUTION: This study confirms that Day-6 morulae should not be discarded in young women undergoing PGT-A. However, a potential drawback of biopsying embryos at the morula stage is the inability to distinguish between inner cell mass and trophectoderm cell origin. The sample size of euploid morula transfer cycles in this study was limited. Thus, a larger cohort would be beneficial to validate the reassuring live birth and spontaneous abortion rates reported here. Furthermore, the reproducibility of our findings should be determined at different centers. WIDER IMPLICATIONS OF THE FINDINGS: Although Day-6 morulae are associated with higher aneuploidy rates and lower IRs compared to blastocysts, they still yielded successful pregnancies. Therefore, testing Day-6 morulae should be considered, especially for women <40 years old who are undergoing PGT-A with a small cohort of available blastocysts for biopsy. STUDY FUNDING/COMPETING INTEREST(S): The authors have nothing to disclose. They received no specific funding for this work. TRIAL REGISTRATION NUMBER: N/A.

Development of ICSI.

The first conception outside of the human body that led to the birth of Louise Brown was a tremendous accomplishment, which opened the door to the utilization of assisted reproductive techniques globally. This brought the understanding that accomplishing life in a dish required several steps, the most obvious being the timing and characteristics of fertilization. It soon became obvious in the 1980s that the most disappointing phenomenon was unexpected and complete fertilization failure. Among the approaches that were attempted to treat male factor infertility, ICSI surfaced as the technique that brought the ratio of the gametes to 1:1 and was also able to grant consistent fertilization and a higher pregnancy rate. ICSI has now been implemented for a quarter of a century, proving itself as the ultimate technique utilizing ejaculated spermatozoa independent of the semen parameters and is the sole insemination method to be used with surgically retrieved spermatozoa. There are currently various indications for ICSI that are widely adopted, rendering it the most popular insemination method worldwide. The reliability of ICSI ensures its employment in upcoming techniques involving in vitro spermatogenesis and neogametogenesis.

The induction of tumour suppressor protein P53 limits the entry of cells into the pluripotent inner cell mass lineage in the mouse embryo.

The early preimplantation embryo is susceptible to a range of exogenous stresses which result in their reduced long-term developmental potential. The P53 tumour suppressor protein is normally held at low levels in the preimplantation embryo and we show that culture stress induces the expression of a range of canonical P53-response genes (Mdm2, Bax and Cdkn1a). Culture stress caused a P53-dependent loss of cells from resulting blastocysts, and this was most evident within the inner cell mass population. Culture stress increased the proportion of cells expressing active caspase-3 and undergoing apoptosis, while inhibition of caspase-3 increased the number of cells within the inner cell mass. The P53-dependent loss of cells from the inner cell mass was accompanied by a loss of NANOG-positive epiblast progenitors. Pharmacological activation of P53 by the MDM2 inhibitor, Nutlin-3, also caused increased P53-dependent transcription and the loss of cells from the inner cell mass. This loss of cells could be ameliorated by simultaneous treatment with the P53 inhibitor, Pifithrin-α. Culture stress causes reduced signalling via the phosphatidylinositol-3-kinase signalling pathway, and blocking this pathway caused P53-dependent loss of cells from the inner cell mass. These results point to P53 acting to limit the accumulation and survival of cells within the pluripotent lineage of the blastocyst and provide a molecular framework for the further investigation of the factors determining the effects of stressors on the embryo's developmental potential.

Fathers' experiences of living with cancer: a phenomenological study.

There is a paucity of knowledge about fathers' experiences of cancer. This study explored the experiences of fathers diagnosed and living with cancer while also having parental responsibility for children. A hermeneutic phenomenological approach guided the study. Data were generated through 22 in-depth interviews with 10 fathers throughout Northern Ireland. The findings evidenced that fathers' identities are challenged and frequently re-shaped by the cancer experience, in many cases leading to an improved lifestyle behaviour. Heightened engagement with their children can provide a protective effect from the illness. On the other hand a lack of involvement led to frustration and low mood. The findings also demonstrated that father/child relationships were adversely affected by the social complexities that exist in the variances and diversity of fathers parenting roles and status. This knowledge contributes to our understanding of the complex relationships of fathers in non-traditional roles. It extends our understanding of how, when stereotyped gendered roles are ascribed to fathers it can impact on a fathers' ability to fulfil the traditional breadwinner's role. This is knowledge that will inform health care professionals and enable them to provide gendered-sensitive care that takes account of the masculine psychological responses that can shape the cancer experience.

Cost-effectiveness of secukinumab as first biologic treatment, compared with other biologics, for moderate to severe psoriasis in Germany.

BACKGROUND: Secukinumab, a fully human monoclonal antibody that selectively neutralizes interleukin 17A, has demonstrated strong and sustained efficacy in adults with moderate to severe psoriasis in clinical trials. OBJECTIVE: This analysis compared the cost per responder of secukinumab as first biologic treatment of moderate to severe psoriasis, with adalimumab, infliximab, etanercept and ustekinumab in Germany. METHODS: A 52-week decision-tree model was developed. Response to treatment was assessed based on the likelihood of achieving a predefined Psoriasis Area and Severity Index (PASI) response to separate the cohort into responders (PASI ≥75), partial responders (PASI 50 to 74) and non-responders (PASI <50). Responders at week 16 continued initial treatment, whereas partial responders and non-responders were switched to standard of care, which included methotrexate, cyclosporine, phototherapy and topical corticosteroids. Sustained response was defined as 16-week response maintained at week 52. A German healthcare system perspective was adopted. Clinical efficacy data were obtained from a mixed-treatment comparison; 2016 resource unit costs from national sources; and adverse events and discontinuation rates from the literature. We calculated cost per PASI 90 responder over week 16 and week 52, as well as cost per sustained responder between weeks 16 and 52. RESULTS: Secukinumab had the lowest cost per PASI 90 responder over 16 weeks (€18 026) compared with ustekinumab (€18 080), adalimumab (€23 499), infliximab (€29 599) and etanercept (€34 037). Over 52 weeks, costs per PASI 90 responder ranged from €42 409 (secukinumab) to €70 363 (etanercept). Likewise, secukinumab had the lowest cost per sustained 52-week PASI 90 responder (€22 690) compared with other biologic treatments. Sensitivity analyses, excluding patient copayments, showed similar results. CONCLUSIONS: First biologic treatment with secukinumab for moderate to severe psoriasis is cost-effective, with lowest cost per responder compared with other biologic treatments in Germany.

A treatment algorithm for couples with unexplained infertility based on sperm chromatin assessment.

OBJECTIVE: To design a reproductive treatment algorithm based on the sperm DNA fragmentation (SDF) for couples with unexplained infertility following a poor intrauterine insemination (IUI) outcome. DESIGN: Couples that failed IUI with no apparent reproductive issue in both partners were allocated to diverse reproductive treatments on the basis of SDF. SETTING: Reproductive medical center in an academic setting. PATIENT(S): Over 4 years, couples with an unexpected poor IUI outcome and no apparent female or male partner reproductive issues were recruited. INTERVENTION(S): IUI, IVF, and ICSI were performed in the standard fashion following sperm SDF assays. MAIN OUTCOMES MEASURE(S): Fertilization rate, implantation rate, pregnancy characteristics, and delivery rates. RESULT(S): A total of 354 couples with unexplained infertility and normal semen parameters underwent 1133 IUI cycles. Clinical pregnancy rate (CPR) with IUI at our center in an age-matched cohort is 23.9% while the study cohort had 1.8%. Following SDF assessment, couples with failed IUI attempts but normal SDF (SCSA 9.8 ± 4.6%; TUNEL 11.8 ± 6.2%) underwent IVF with a CPR of 12.7%; those with abnormal SDF underwent ICSI with ejaculated spermatozoa, resulting in a CPR of 18.7%. This group included couples with normal SDF that had failed IVF. Couples with abnormal SDF that failed ICSI with ejaculated spermatozoa achieved a CPR of 31.0% with surgically retrieved spermatozoa. CONCLUSION(S): Couples with unexplained infertility that present with unexpectedly poor IUI outcomes can be funneled into a treatment algorithm guided by the integrity of the sperm genome for higher chances of pregnancy using an alternate method of insemination.

Histone 3 lysine 9 acetylation is a biomarker of the effects of culture on zygotes.

Acetylation of histone proteins is a major determinant of chromatin structure and function. Fertilisation triggers a round of chromatin remodelling that prepares the genome for the first round of transcription from the new embryonic genome. In this study we confirm that fertilisation leads to a marked progressive increase in the level of histone 3 lysine 9 acetylation in both the paternally and maternally derived genomes. The culture of zygotes in simple defined media caused a marked increase in the global level of acetylation and this affected the male pronucleus more than the female. The culture created a marked asymmetry in staining between the two pronuclei that was not readily detected in zygotes collected directly from the reproductive tract and was ameliorated to some extent by optimized culture media. The increased acetylation caused by culture resulted in increased transcription of Hspa1b, a marker of embryonic genome activation. Pharmacological analyses showed the hyperacetylation of H3K9 and the increased expression of Hspa1b caused by culture were due to the altered net activity of a range of histone acetylases and deacetylases. The marked hyperacetylation of histone 3 lysine 9 caused by culture of zygotes may serve as an early biomarker for the effects of culture on the normal function of the embryo. The results also provide further evidence for an effect of the stresses associated with assisted reproductive technologies on the normal patterns of epigenetic reprogramming in the early embryo.

Intracytoplasmic sperm injection: state of the art in humans.

Among infertile couples, 25% involve both male and female factors, while male factor alone accounts for another 25% due to oligo-, astheno-, teratozoospermia, a combination of the three, or even a complete absence of sperm cells in the ejaculate and can lead to a poor prognosis even with the help of assisted reproductive technology (ART). Intracytoplasmic sperm injection (ICSI) has been with us now for a quarter of a century and in spite of the controversy generated since its inception, it remains in the forefront of the techniques utilized in ART. The development of ICSI in 1992 has drastically decreased the impact of male factor, resulting in millions of pregnancies worldwide for couples who, without ICSI, would have had little chance of having their own biological child. This review focuses on the state of the art of ICSI regarding utility of bioassays that evaluate male factor infertility beyond the standard semen analysis and describes the current application and advances in regard to ICSI, particularly the genetic and epigenetic characteristics of spermatozoa and their impact on reproductive outcome.

The cost of systemic corticosteroid-induced morbidity in severe asthma: a health economic analysis.

BACKGROUND: Treatment of severe asthma may include high dose systemic-steroid therapy which is associated with substantial additional morbidity. This study estimates the additional healthcare costs associated with steroid-induced morbidity by comparing three patients groups: those with severe asthma, moderate asthma and no asthma. METHODS: Patients with severe asthma (n = 808, GINA step 5 treatment) were matched by age and gender with patients with mild/moderate asthma (n = 3,975, GINA step 2 and 3 treatment) and a non-asthma control cohort (with a diagnosis of rhinitis; n = 2,412) from the Optimum Patient Care Research Database (OPCRD), a nationally representative primary care database. Prescribed drugs and publicly funded healthcare activity were monetised and annual costs per patient estimated. Regression analyses were used to estimate the additional healthcare cost associated with steroid-induced morbidity. RESULTS: Average healthcare costs per person per year range from £2603 - £4533 for the severe asthma cohort, to £978 - £2072 for the mild/moderate asthma cohort, to £560 - £1324 for the non-asthma control cohort, depending on the costing scenario. Differences in induced morbidity costs were evident between patients with asthma differentiated by steroid exposure. In relation to prescription drugs used to treat steroid-induced co-morbidities, females with severe asthma and high steroid exposure cost approximately £789 more per year than a corresponding female with no asthma, while males cost approximately £744 more than their counterparts with no asthma. Estimates were extrapolated to all healthcare costs. CONCLUSIONS: This study provides the first robust estimates of the additional cost of healthcare related to steroid-induced morbidity relative to patients with no steroid exposure. The study will help inform use of steroid-sparing strategies in this patient group.

Mapping global changes in nuclear cytosine base modifications in the early mouse embryo.

Reprogramming epigenetic modifications to cytosine is required for normal embryo development. We used improved immunolocalization techniques to simultaneously map global changes in the levels of 5'-methylcytosine (5meC) and 5'-hydroxymethylcytosine (5hmC) in each cell of the embryo from fertilization through the first rounds of cellular differentiation. The male and female pronuclei of the zygote showed similar staining levels, and these remained elevated over the next three cell cycles. The inner cells of the morula showed a progressive reduction in global levels of both 5meC and 5hmC and further losses occurred in the pluripotent inner cell mass (ICM) of the blastocyst. This was accompanied by undetectable levels of DNA methyltransferase of each class in the nuclei of the ICM, while DNA methyltransferase 3B was elevated in the hypermethylated nuclei of the trophectoderm (TE). Segregation of the ICM into hypoblast and epiblast was accompanied by increased levels in the hypoblast compared with the epiblast. Blastocyst outgrowth in vitro is a model for implantation and showed that a demethylated state persisted in the epiblast while the hypoblast had higher levels of both 5meC and 5hmC staining. The high levels of 5meC and 5hmC evident in the TE persisted in trophoblast and trophoblast giant cells after attachment of the blastocyst to the substratum in vitro. This study shows that global cytosine hypomethylation and hypohydroxymethylation accompanied the formation of the pluripotent ICM and this persisted into the epiblast after blastocyst outgrowth, and each differentiated lineage formed in the early embryo showed higher global levels of 5meC and 5hmC.

DAT isn't all that: cocaine reward and reinforcement require Toll-like receptor 4 signaling.

The initial reinforcing properties of drugs of abuse, such as cocaine, are largely attributed to their ability to activate the mesolimbic dopamine system. Resulting increases in extracellular dopamine in the nucleus accumbens (NAc) are traditionally thought to result from cocaine's ability to block dopamine transporters (DATs). Here we demonstrate that cocaine also interacts with the immunosurveillance receptor complex, Toll-like receptor 4 (TLR4), on microglial cells to initiate central innate immune signaling. Disruption of cocaine signaling at TLR4 suppresses cocaine-induced extracellular dopamine in the NAc, as well as cocaine conditioned place preference and cocaine self-administration. These results provide a novel understanding of the neurobiological mechanisms underlying cocaine reward/reinforcement that includes a critical role for central immune signaling, and offer a new target for medication development for cocaine abuse treatment.

Healthcare utilisation among cancer survivors over 50 years of age.

There are now over 104,000 people living in Ireland with a cancer diagnosis. Using The Irish Longitudinal Study on Ageing (TILDA), healthcare utilisation of cancer survivors (aged 50 +) was compared with those without a history of cancer across service providers. Our cancer variable was stratified by time since diagnosis (2-5, 6-10, 11+ years) and type (breast, prostate, colorectal and a miscellaneous group of other cancers). While the probability of cancer survivors accessing GP services was not significant different to respondents without a history of cancer, the probability of an outpatient specialist office visit was 19.5, 11.8 and 14.0 percentage points higher, respectively for those 2-5years, 6-10 years and 11 years or more after their cancer diagnosis and was statistically significant. In Ireland, the pattern of GP and specialist use appears less well defined compared to other European countries. This suggests an overarching policy response is required for cancer survivorship care.

Clinical Consensus Guidelines on the Application of Cerebrospinal Fluid Biomarkers for Alzheimer's Disease Diagnosis: Recommendations of the Irish Network for Biomarkers in Neurodegeneration.

It is accepted that a lumbar puncture (LP) and cerebrospinal fluid (CSF) biomarker analysis support the routine diagnostic work-up for the differential diagnosis of dementia due to Alzheimer's disease (AD) within certain patient cohorts1. These tests, which measure CSF protein concentrations of amyloid-ß42 (Aß42), total tau (t-tau) and phospho tau (p-tau), were recently validated, accredited and made available clinically for the first time in Ireland. A working group, comprising Irish clinical and scientific researchers, met to review a) the validation results; b) international consensus opinions, and c) research and clinical evidence as to the clinical utility of CSF biomarker analysis for AD dementia diagnosis. The outcome of this meeting was the formulation of a consensus statement paper for the benefit of health care professionals involved in the diagnosis and management of dementia to ensure appropriate use of these biomarker tests in clinical settings in Ireland.

Short- and long-term effects of gestational diabetes mellitus on healthcare cost: a cross-sectional comparative study in the ATLANTIC DIP cohort.

AIMS: This paper examines the association between gestational diabetes mellitus and costs of care during pregnancy and 2-5 years post pregnancy. METHODS: Healthcare utilization during pregnancy was measured for a sample of 658 women drawn from the Atlantic Diabetes in Pregnancy (ATLANTIC DIP) network. Healthcare utilization 2-5 years post pregnancy was assessed for a subsample of 348 women via a postal questionnaire. A vector of unit costs was applied to healthcare activity to calculate the costs of care at both time points. Differences in cost for women with gestational diabetes mellitus compared with those with normal glucose tolerance during the pregnancy were examined using univariate and multivariate regression analyses. RESULTS: Gestational diabetes mellitus was independently associated with an additional €817.60 during pregnancy (€1192.1 in the gestational diabetes mellitus group, €511.6 in the normal glucose tolerance group), in the form of additional delivery and neonatal care costs, and an additional €680.50 in annual healthcare costs 2-5 years after the index pregnancy (€6252.4 in the gestational diabetes mellitus group, €5434.8 in the normal glucose tolerance group). CONCLUSIONS: These results suggest that gestational diabetes mellitus is associated with increased costs of care during and post pregnancy. They provide indication of the associated cost that can be avoided or reduced by the screening, prevention and management of gestational diabetes mellitus in pregnancy. These estimates are useful for further studies that examine the cost and cost-effectiveness of such programmes.

Differential expression and functionality of ATP-binding cassette transporters in the human hair follicle.

BACKGROUND: ATP-binding cassette (ABC) transporters are involved in the active transport of an extremely diverse range of substrates across biological membranes. These transporters are commonly implicated in the development of multidrug resistance and are also involved in numerous physiological and homeostatic processes, including lipid transport, cell migration and differentiation. OBJECTIVES: To close the knowledge gap in the expression of ABC transporters in the human hair follicle (HF). METHODS: Quantitative polymerase chain reaction (qPCR) of ABC genes and immunofluorescence microscopy analysis of cryosections of human HFs. RESULTS: By qPCR analysis, numerous members of the ABC transporter superfamily, such as ABCB1, ABCG2 and ABCA12, were found to be transcribed in full-length human scalp HFs. Immunofluorescence microscopy demonstrated that the intrafollicular protein expression of different xenobiotic ABC transporters (ABCB1, ABCC1, ABCC4, ABCG2) varies greatly, with ABCG2 expression restricted primarily to the epithelial stem cell region of the outer root sheath (bulge), whereas expression of ABCB1, ABCC1 and ABCC4 was more widespread. Lipid transporters ABCA1, ABCA12 and ABCA4 were almost uniformly expressed throughout the HF epithelium. Functional ABCB1/G2 activity was demonstrated by exclusion of the substrate dye, Hoechst 33342. In the bulge, this was reversed by ABCB1 and ABCG2 inhibition. CONCLUSIONS: These data encourage further investigation of ABC transporters as potentially important regulators of HF epithelial biology. Clinically, pharmacological modulation of the activity of selected intrafollicular ABC transporters may permit novel therapeutic interventions, such as protecting HF stem cells from chemotherapy-induced damage, counteracting cholesterol-associated hypertrichosis, and manipulating the intrafollicular prostaglandin balance in androgenetic alopecia.

Determinants of valid measurements of global changes in 5'-methylcytosine and 5'-hydroxymethylcytosine by immunolocalisation in the early embryo.

A classical model of epigenetic reprogramming of methyl-cytosine-phosphate-guanine (CpG) dinucleotides within the genome of the early embryo involves a process of active demethylation of the paternally derived genome immediately following fertilisation, creating marked asymmetry in global cytosine methylation levels in male and female pronuclei, followed by passive demethylation of the maternally derived genome over subsequent cell cycles. This model has dominated thinking in developmental epigenetics over recent decades. Recent re-analyses of the model show that demethylation of the paternally derived genome is more modest than formerly thought and results in overall similar levels of methylation of the paternal and maternal pronuclei in presyngamal zygotes, although there is little evidence for a pervasive process of passive demethylation during the cleavage stage of development. In contrast, the inner cell mass of the blastocyst shows some loss of methylation within specific classes of loci. Improved methods of chemical analysis now allow global base-level analysis of modifications to CpG dinucleotides within the cells of the early embryo, yet the low cost and convenience of the immunolocalisation techniques mean that they still have a valuable place in the analysis of the epigenetics of embryo development. In this review we consider the key strengths and weaknesses of this methodology and some factors required for its valid use and interpretation.

Survival signalling in the preimplantation embryo.

The development of the preimplantation embryo (from fertilisation until the formation of the differentiated blastocyst) occurs without a requirement for exogenous mitogenic or survival signals. This distinguishes the behaviour of cells in the early embryo from all other normal cells. The discovery that fertilisation triggers the production and release of potent bioactive mediators by the embryo that act back on membrane receptors demonstrated the presence of closed autocrine embryotrophic loops. It is now clear that these ligands act in concert with paracrine mediators normally present within the reproductive tract to support the normal development of the embryo. These ligands act via receptors expressed by the embryo to activate signalling transduced by 1-o-phosphatidylinositol-3-kinase and the resultant formation of phosphatidylinositol-3,4,5-trisphosphate. This polyphosphorylated membrane phospholipid acts as a docking site for proteins possessing the PH domain. These include PDK1, AKT and phospholipase C. The activation of these proteins accounts for the initiation of new transcription from the embryonic genome to form a pro-survival, anti-apoptotic transcriptome and the post-transcriptional activation of pro-survival signalling within embryonic cells. This includes the attenuation of action of pro-apoptotic signals, such as P53. The production of embryotrophic ligands after fertilisation bootstraps development by the activation of transcription from the embryonic genome, followed by the activation of pro-survival settings within embryo cells.