Transcriptome Profiling of Sexual Maturation and Mating in the Mediterranean Fruit Fly, Ceratitis capitata

Sexual maturation and mating in insects are generally accompanied by major physiological and behavioural changes. Many of these changes are related to the need to locate a mate and subsequently, in the case of females, to switch from mate searching to oviposition behaviour. The prodigious reproductive capacity of the Mediterranean fruit fly, Ceratitis capitata, is one of the factors that has led to its success as an invasive pest species. To identify the molecular changes related to maturation and mating status in male and female medfly, a microarray-based gene expression approach was used to compare the head transcriptomes of sexually immature, mature virgin, and mated individuals. Attention was focused on the changes in abundance of transcripts related to reproduction, behaviour, sensory perception of chemical stimulus, and immune system processes. Broad transcriptional changes were recorded during female maturation, while post-mating transcriptional changes in females were, by contrast, modest. In male medfly, transcriptional changes were consistent both during maturation and as a consequence of mating. Of particular note was the lack of the mating-induced immune responses that have been recorded for Drosophila melanogaster, that may be due to the different reproductive strategies of these species. This study, in addition to increasing our understanding of the molecular machinery behind maturation and mating in the medfly, has identified important gene targets that might be useful in the future management of this pest.

Physiological and symptomatic responses to cycling and walking in intermittent claudication

To shed light on the potential efficacy of cycling as a testing modality in the treatment of intermittent claudication (IC), this study compared physiological and symptomatic responses to graded walking and cycling tests in claudicants. Sixteen subjects with peripheral arterial disease (resting ankle: brachial index (ABI) < 0.9) and IC completed a maximal graded treadmill walking (T) and cycle (C) test after three familiarization tests on each mode. During each test, symptoms, oxygen uptake (VO2), minute ventilation (VE), respiratory exchange ratio (RER) and heart rate (HR) were measured, and for 10 min after each test the brachial and ankle systolic pressures were recorded. All but one subject experienced calf pain as the primary limiting symptom during T; whereas the symptoms were more varied during C and included thigh pain, calf pain and dyspnoea. Although maximal exercise time was significantly longer on C than T (690 +/- 67 vs. 495 +/- 57 s), peak VO2, peak VE and peak heart rate during C and T were not different; whereas peak RER was higher during C. These responses during C and T were also positively correlated (P < 0.05) with each other, with the exception of RER. The postexercise systolic pressures were also not different between C and T. However, the peak decline in ankle pressures from resting values after C and T were not correlated with each other. These data demonstrate that cycling and walking induce a similar level of metabolic and cardiovascular strain, but that the primary limiting symptoms and haemodynamic response in an individual's extremity, measured after exercise, can differ substantially between these two modes.

Within- and between-day repeatability and variability in children's physiological responses during submaximal treadmill exercise

The purpose of this study was to verify within- and between-day repeatability and variability in children's oxygen uptake (VO^sub 2^), gross economy (GE; VO^sub 2^ divided by speed) and heart rate (HR) during treadmill walking based on self-selected speed (SS). Fourteen children (10.1 ± 1.4 years) undertook three testing sessions over 2 days in which four walking speeds, including SS were tested. Within- and between-day repeatability were assessed using the Bland and Altman method, and coefficients of variability (CV) were determined for each child across exercise bouts and averaged to obtain a mean group CV value for VO^sub 2^, GE, and HR per speed. Repeated measures analysis of variance showed no statistically significant differences in within- or between-day CV for VO^sub 2^, GE, or HR at any speed. Repeatability within- and between-day for VO^sub 2^, GE, and HR for all speeds was verified. These results suggest that submaximal VO^sub 2^ during treadmill walking is stable and reproducible at a range of speeds based on children's SS.

Inequalities in cardiovascular disease mortality : the role of behavioural, physiological and social risk factors

Background: While the relationship between socioeconomic disadvantage and cardiovascular disease (CVD) is well established, the role that traditional cardiovascular risk factors play in this association remains unclear. We examined the association between education attainment and CVD mortality and the extent to which behavioural, social and physiological factors explained this relationship. Methods: Adults (n=38 355) aged 40-69 years living in Melbourne, Australia were recruited in 1990-1994. Subjects with baseline CVD risk factor data ascertained through questionnaire and physical measurement were followed for an average of 9.4 years with CVD deaths verified by review of medical records and autopsy reports. Results: CVD mortality was higher for those with primary education only compared to those who had completed tertiary education, with a hazard ratio (HR) of 1.66 (95% confidence interval [CI] 1.11-2.49) after adjustment for age, country of birth and gender. Those from the lowest educated group had a more adverse cardiovascular risk factor profile compared to the highest educated group, and adjustment for these risk factors reduced the HR to 1.18 (95% CI 0.78-1.77). In analysis of individual risk factors, smoking and waist circumference explained most of the difference in CVD mortality between the highest and lowest education groups. Conclusions: Most of the excess CVD mortality in lower socioeconomic groups can be explained by known risk factors, particularly smoking and overweight. While targeting cardiovascular risk factors should not divert efforts from addressing the underlying determinants of health inequalities, it is essential that known risk factors are addressed effectively among lower socioeconomic groups.

Dynamics of in vitro polymer degradation of polycaprolactone-based scaffolds : accelerated versus simulated physiological conditions

The increasing use of biodegradable devices in tissue engineering and regenerative medicine means it is essential to study and understand their degradation behaviour. Accelerated degradation systems aim to achieve similar degradation profiles within a shorter period of time, compared with standard conditions. However, these conditions only partially mimic the actual situation, and subsequent analyses and derived mechanisms must be treated with caution and should always be supported by actual long-term degradation data obtained under physiological conditions. Our studies revealed that polycaprolactone (PCL) and PCL-composite scaffolds degrade very differently under these different degradation conditions, whilst still undergoing hydrolysis. Molecular weight and mass loss results differ due to the different degradation pathways followed (surface degradation pathway for accelerated conditions and bulk degradation pathway for simulated physiological conditions). Crystallinity studies revealed similar patterns of recrystallization dynamics, and mechanical data indicated that the scaffolds retained their functional stability, in both instances, over the course of degradation. Ultimately, polymer degradation was shown to be chiefly governed by molecular weight, crystallinity susceptibility to hydrolysis and device architecture considerations whilst maintaining its thermodynamic equilibrium.

Overeating as a clue to weight gain: behavioural and physiological interactions in the control of food intake

It has been estimated that 25-50% of people in most affluent societies are either obese or overweight. These disorders are the result of an imbalance between calorific intake and energy expenditure over a prolonged time period. These types of disorders are among the most common health problems in industrialized societies. Addressing these issues and offering new strategies, this thorough new study draws together contributions from interdisciplinary and international group of specialists, includes recent research on genetic influences, features discussions of epidemiological studies and covers both biological and social aspects of obesity.

Physiological investigation of periosteum structure and its application in periosteum tissue engineering

Although many different materials, techniques and methods, including artificial or engineered bone substitutes, have been used to repair various bone defects, the restoration of critical-sized bone defects caused by trauma, surgery or congenital malformation is still a great challenge to orthopedic surgeons. One important fact that has been neglected in the pursuit of resolutions for large bone defect healing is that most physiological bone defect healing needs the periosteum and stripping off the periosteum may result in non-union or non-healed bone defects. Periosteum plays very important roles not only in bone development but also in bone defect healing. The purpose of this project was to construct a functional periosteum in vitro using a single stem cell source and then test its ability to aid the repair of critical-sized bone defect in animal models. This project was designed with three separate but closely-linked parts which in the end led to four independent papers. The first part of this study investigated the structural and cellular features in periostea from diaphyseal and metaphyseal bone surfaces in rats of different ages or with osteoporosis. Histological and immunohistological methods were used in this part of the study. Results revealed that the structure and cell populations in periosteum are both age-related and site-specific. The diaphyseal periosteum showed age-related degeneration, whereas the metaphyseal periosteum is more destructive in older aged rats. The periosteum from osteoporotic bones differs from normal bones both in terms of structure and cell populations. This is especially evident in the cambial layer of the metaphyseal area. Bone resorption appears to be more active in the periosteum from osteoporotic bones, whereas bone formation activity is comparable between the osteoporotic and normal bone. The dysregulation of bone resorption and formation in the periosteum may also be the effect of the interaction between various neural pathways and the cell populations residing within it. One of the most important aspects in periosteum engineering is how to introduce new blood vessels into the engineered periosteum to help form vascularized bone tissues in bone defect areas. The second part of this study was designed to investigate the possibility of differentiating bone marrow stromal cells (BMSCs) into the endothelial cells and using them to construct vascularized periosteum. The endothelial cell differentiation of BMSCs was induced in pro-angiogenic media under both normoxia and CoCl2 (hypoxia-mimicking agent)-induced hypoxia conditions. The VEGF/PEDF expression pattern, endothelial cell specific marker expression, in vitro and in vivo vascularization ability of BMSCs cultured in different situations were assessed. Results revealed that BMSCs most likely cannot be differentiated into endothelial cells through the application of pro-angiogenic growth factors or by culturing under CoCl2-induced hypoxic conditions. However, they may be involved in angiogenesis as regulators under both normoxia and hypoxia conditions. Two major angiogenesis-related growth factors, VEGF (pro-angiogenic) and PEDF (anti-angiogenic) were found to have altered their expressions in accordance with the extracellular environment. BMSCs treated with the hypoxia-mimicking agent CoCl2 expressed more VEGF and less PEDF and enhanced the vascularization of subcutaneous implants in vivo. Based on the findings of the second part, the CoCl2 pre-treated BMSCs were used to construct periosteum, and the in vivo vascularization and osteogenesis of the constructed periosteum were assessed in the third part of this project. The findings of the third part revealed that BMSCs pre-treated with CoCl2 could enhance both ectopic and orthotopic osteogenesis of BMSCs-derived osteoblasts and vascularization at the early osteogenic stage, and the endothelial cells (HUVECs), which were used as positive control, were only capable of promoting osteogenesis after four-weeks. The subcutaneous area of the mouse is most likely inappropriate for assessing new bone formation on collagen scaffolds. This study demonstrated the potential application of CoCl2 pre-treated BMSCs in the tissue engineering not only for periosteum but also bone or other vascularized tissues. In summary, the structure and cell populations in periosteum are age-related, site-specific and closely linked with bone health status. BMSCs as a stem cell source for periosteum engineering are not endothelial cell progenitors but regulators, and CoCl2-treated BMSCs expressed more VEGF and less PEDF. These CoCl2-treated BMSCs enhanced both vascularization and osteogenesis in constructed periosteum transplanted in vivo.

The mechanism of maturation of insulin-like growth factor-1

This study, to elucidate the role of des(1-3)IGF-I in the maturation of IGF-I,used two strategies. The first was to detect the presence of enzymes in tissues, which would act on IGF-I to produce des(1-3)IGF-I, and the second was to detect the potential products of such enzymic activity, namely Gly-Pro-Glu(GPE), Gly-Pro(GP) and des(l- 3)IGF-I. No neutral tripeptidyl peptidase (TPP II), which would release the tripeptide GPE from IGF-I, was detected in brain, urine nor in red or white blood cells. The TPPlike activity which was detected, was attributed to a combined action of a dipeptidyl peptidase (DPP N) and an aminopeptidase (AP A). A true TPP II was, however, detected in platelets. Two purified TPP II enzymes were investigated but they did not release GPE from IGF-I under a variety of conditions. Consequently, TPP II seemed unlikely to participate in the formation of des(1-3)IGF-I. In contrast, an acidic tripeptidyl peptidase activity (TPP I) was detected in brain and colostrum, the former with a pH optimum of 4.5 and the latter 3.8. It seems likely that such an enzyme would participate in the formation of des( 1-3 )IGF-I in these tissues in vitro, ie. that des(1-3)IGF-I may have been produced as an artifact in the isolation of IGF-I from brain and colostrum in acidic conditions. This contrasts with suggestions of an in vivo role for des(1-3)IGF-I, as reported by others. The activity of a dipeptidyl peptidase N (DPP N) from urine, which should release the dipeptide GP from IGF-I, was assessed under a variety of conditions and with a variety of additives and potential enzyme stimulants, but there was no release of GP. The DPP N also exhibited a transferase activity with synthetic substrates in the presence of dipeptides, at lower concentrations than previously reported for other acceptors or other proteolytic enzymes. In addition, a low concentration of a product,possibly the tetrapeptide Gly-Pro-Gly-Leu, was detected with the action of the enzyme on IGF-I in the presence of the dipeptide Gly-Leu. As part of attempts to detect tissue production of des(1-3)IGF-I, a monoclonal antibody (MAb ), directed towards the GPE- end ofiGF-I was produced by immunisation with a 10-mer covalently attached to a carrier protein. By the use of indirect ELISA and inhibitor studies, the MAb was shown to selectively recognise peptides with anNterminal GPE- sequence, and applied to the indirect detection of des(1-3)IGF-I. The concentration of GPE in brain, measured by mass spectrometry ( MS), was low, and the concentration of total IGF-I (measured by ELISA with a commercial polyclonal antibody [P Ab]) was 40 times higher at 50 nmol/kg. This also, was not consistent with the action of a tripeptidyl peptidase in brain that converted all IGF-I to des(1-3)IGF-I plus GPE. Contrasting ELISA results, using the MAb prepared in this study, suggest an even higher concentration of intact IGF-I of 150 nmollkg. This would argue against the presence of any des( 1-3 )IGF-I in brain, but in turn, this indicates either the presence of other substances containing a GPE amino-terminus or other cross reacting epitope. Although the results of the specificity studies reported in Chapter 5 would make this latter possibility seem unlikely, it cannot be completely excluded. No GP was detected in brain by MS. No GPE was detected in colostrum by capillary electrophoresis (CE) but the interference from extraneous substances reduced the detectability of GPE by CE and this approach would require further, prior, purification and concentration steps. A molecule, with a migration time equal to that of the peptide GP, was detected in colostrum by CE, but the concentration (~ 10 11mo/L) was much higher than the IGF-I concentration measured by radio-immunoassay using a PAb (80 nmol/L) or using a Mab (300-400 nmolL). A DPP IV enzyme was detected in colostrum and this could account for the GP, derived from substrates other than IGF-1. Based on the differential results of the two antibody assays, there was no indication of the presence of des(1-3)IGF-I in brain or colostrum. In the absence of any enzyme activity directed towards the amino terminus of IGF-I and the absence any potential products, IGF-I, therefore, does not appear to "mature" via des(1-3)IGF-I in the brain, nor in the neutral colostrum. In spite of these results which indicate the absence of an enzymic attack on IGF-I and the absence of the expected products in tissues, the possibility that the conversion of IGF-I may occur in neutral conditions in limited amounts, cannot be ruled out. It remains possible that in the extracellular environment of the membrane, a complex interaction of IGF-I, binding protein, aminopeptidase(s) and receptor, produces des(1- 3)IGF-I as a transient product which is bound to the receptor and internalised.

An overview of physical growth and maturation

An understanding of physical growth and maturation is relevant to many disciplines, including exercise and sport science, anthropology, human biology, medicine, psychology and education. Growth and maturation is governed by a complex interaction between genetic and environmental factors. There is increasing evidence that physical activity plays an important role in normal growth, development, health and well-being of children and youth, however, caution is required in the activity setting so that growth and maturation is not jeopardised. To appreciate the impact of physical activity and/or exercise on growth and maturation, a thorough understanding of the general principles of auxology is useful. Following an introduction to terminology, an overview of physical growth and development is provided in the context of morphological changes. Detailed information is provided regarding individual variability in growth and development along with sexual dimorphism. A small degree of sexual dimorphism exists at birth however striking differences develop during the pubertal years. Sexual dimorphism in body composition is largely regulated by endocrine factors with critical roles played by growth hormone and gonadal steroids.

Human atrial beta1L-adrenoceptor but not beta3-adrenoceptor activation increases force and Ca2+ current at physiological temperature

BACKGROUND AND PURPOSE It has been proposed that BRL37344, SR58611 and CGP12177 activate b3-adrenoceptors in human atrium to increase contractility and L-type Ca2+ current (ICa-L). b3-adrenoceptor agonists are potentially beneficial for the treatment of a variety of diseases but concomitant cardiostimulation would be potentially harmful. It has also been proposed that (-)-CGP12177 activates the low affinity binding site of the b1-adrenoceptor in human atrium. We therefore used BRL37344, SR58611 and (-)-CGP12177 with selective b-adrenoceptor subtype antagonists to clarify cardiostimulant b-adrenoceptor subtypes in human atrium. EXPERIMENTAL APPROACH Human right atrium was obtained from patients without heart failure undergoing coronary artery bypass or valve surgery. Cardiomyocytes were prepared to test BRL37344, SR58611 and CGP12177 effects on ICa-L. Contractile effects were determined on right atrial trabeculae. KEY RESULTS BRL37344 increased force which was antagonized by blockade of b1- and b2-adrenoceptors but not by blockade of b3-adrenoceptors with b3-adrenoceptor-selective L-748,337 (1 mM). The b3-adrenoceptor agonist SR58611 (1 nM–10 mM) did not affect atrial force. BRL37344 and SR58611 did not increase ICa-L at 37°C, but did at 24°C which was prevented by L-748,337. (-)-CGP12177 increased force and ICa-L at both 24°C and 37°C which was prevented by (-)-bupranolol (1–10 mM), but not L-748,337. CONCLUSIONS AND IMPLICATIONS We conclude that the inotropic responses to BRL37344 are mediated through b1- and b2-adrenoceptors. The inotropic and ICa-L responses to (-)-CGP12177 are mediated through the low affinity site b1L-adrenoceptor of the b1-adrenoceptor. b3-adrenoceptor-mediated increases in ICa-L are restricted to low temperatures. Human atrial b3-adrenoceptors do not change contractility and ICa-L at physiological temperature.

Ureaplasma colonization of amniotic fluid and efficacy of antenatal corticosteroids for preterm lung maturation in sheep

Objective: To assess the efficacy of maternal betamethasone for improving preterm lung function, in the presence of inflammation induced by amniotic fluid ureaplasma colonization. ----- ----- Study design: Ewes bearing single fetuses were randomized to receive an intra-amniotic injection of Ureaplasma parvum (serovar 6; 2×107 colony forming units) or vehicle at 86±2 days of pregnancy (mean±SD: term is 150d), followed by maternal intramuscular betamethasone (0.5mg/kg) or saline, either 2 or 7 days before delivery of lambs at 123±1d. ----- ----- Results: Amniotic fluid IL-8 was elevated by ureaplasmas (p=0.049) but unaffected by betamethasone. Lung inflammation induced by ureaplasmas was not affected by betamethasone. Lung compliance was increased by ureaplasma colonization (p=0.009) and betamethasone (p=0.042), and effects were additive. Lung surfactant was increased by ureaplasma colonization (p<0.001) and betamethasone 7 days (p=0.001), but not 2 days, before delivery. ----- ----- Conclusion: Inflammation improves preterm lung function due to increases in surfactant. Antenatal corticosteroids further augment lung function, through an apparently independent mechanism.