bluuebunny/crossref_metadata_2025 huggingface dataset:
A test of the coincidence and duration models of melatonin action in Siberian hamsters. II. The effects of 4‐ and 8‐hr melatonin infusions on testicular development of pinealectomized juvenile Siberian hamsters (Phodopus sungorus)
Authors: Gündüz Bülent , Stetson Milton H.
Published: March 2003

In a previous paper we demonstrated that properly timed 1‐hr infusions of 50 ng melatonin effectively suppressed testicular development in juvenile Siberian hamsters. Only melatonin infused between 20:00 and 21:00 hr was effective in animals exposed to 16L (lights off 20:00 hr). In this paper we further investigate the importance of the coincidence and duration hypotheses of daily exposure of melatonin. Prepubertal Siberian hamsters received either 4‐ or 8‐hr melatonin infusions at various times either on long photoperiod (LD 16:8=16L) or on short photoperiod (LD 10:14=10L). Daily 8‐hr melatonin infusions suppressed testicular development in both photoperiods. Daily 4‐hr, 50 ng/hr, melatonin infusions at 17:00–21:00 hr inhibited testicular growth in 16L and daily 4‐hr melatonin infusions (either 50 ng/h or 50 ng/day) inhibited testicular growth at 17:00–21:00 hr in 10L. We also tested the efficacy of an interrupted melatonin infusion of long duration (8 hr). Pinealectomized prepubertal male Siberian hamsters, born on 16L, were infused with two signals of 4 hr separated by an interval of 2 hr. Melatonin‐infused groups had significantly inhibited testicular growth compared to vehicle‐infused animals. Testicular development was maximally inhibited only in those groups in which the period of melatonin sensitivity identified in the previous paper (20:00–21:00 hr) overlapped or immediately followed a period of melatonin infusion. Considering the restrictions of the experimental design employed in these studies, the results are best explained by the hypothesis that the photoperiodic gonadal response in juvenile Siberian hamsters is regulated by the coincidence in time of exogenously administered melatonin with an intrinsic rhythm of sensitivity to melatonin, which occurred at 20:00–21:00 hr. The duration of the melatonin signal alone can not explain the results.
In vitro melatonin treatment enhances cell‐mediated immune function in male prairie voles (Microtus ochrogaster)
Authors: Kriegsfeld Lance J. , Drazen Deborah L. , Nelson Randy J.
Published: March 2003

The present study was designed (1) to determine the extent to which male prairie voles (Microtus ochrogaster) alter immune status in response to short‐day lengths, (2) to evaluate the role of melatonin in coordinating these alterations in immune function, and (3) to assess the association between alterations in immune function and reproductive responsiveness to photoperiod. Male voles were housed in either long‐ or short‐day lengths for 10 wk; voles in short days were subdivided into reproductive “responders” (R) or “non‐responders” (NR) based on testicular mass at autopsy. After 10 wk of exposure to photoperiodic conditions, cell‐mediated immune function was evaluated using an in vitro splenocyte proliferation assay. The direct effects of melatonin on immune cells were evaluated by adding melatonin to one‐half of the cultures in each experimental condition. Melatonin treatment led to enhanced splenocyte proliferation for all experimental groups. Neither photoperiodic condition nor reproductive status was associated with alterations in immune function or the degree of immuno‐enhancing effects of melatonin. Taken together, the results of the present study suggest that melatonin is capable of enhancing immune function in male voles potentially by acting directly on immune cells.
Melatonin inhibits PHA‐stimulated chicken lymphocyte proliferation in vitro
Authors: Markowska Magdalena , Waloch Maria , Skwarlo‐Sońta Krystyna
Published: March 2003

Many studies have shown that melatonin plays a fundamental role in neuroimmunomodulation. There are known differences between mammals and birds in immunomodulatory function of melatonin exerted in vivo. In present study the effect of exogenous melatonin on chicken lymphoid cell proliferation in vitro was examined. Melatonin alone (10−10M–10−5M) did not exert any effect on the proliferation of the chicken thymocytes, splenocytes and lymphocytes from the bursa of Fabricius. On the other hand, melatonin addition strongly inhibited the proliferation of PHA‐activated thymocytes and splenocytes from young chickens. The effect of melatonin was hormone‐ and mitogen‐(PHA) dose‐dependent; the most pronounced effect was obtained at low PHA and melatonin concentrations i.e., 1.6–3.125 μg/100 μl of PHA and 10−10M–10−7M melatonin, respectively. Prior immunization of chickens with sheep red blood cells (SRBC) inhibited the proliferative response of splenocytes to the same dose of PHA and, consequently, the effect of melatonin addition was not observed. The results reported demonstrate a direct inhibitory effect of melatonin on in vitro activated chicken lymphocytes.
Effect of melatonin on cellular energy depletion mediated by peroxynitrite and poly (ADP‐ribose) synthetase activation in an acute model of inflammation
Authors: Dugo Laura , Serraino Ivana , Fulia Francesco , De Sarro Angela , Caputi Achille P. , Cuzzocrea Salvatore
Published: March 2003

DNA single‐strand breakage and activation of the nuclear enzyme poly (ADP‐ribose) synthetase (PARS) triggers an energy‐consuming, inefficient repair cycle, which contributes to peroxynitrite‐induced cellular injury. Recently, we proposed that during an acute model (pleurisy), cellular injury is mediated by peroxynitrite formation and consequent PARS activation. Here, we investigated whether in vivo melatonin treatment inhibits cellular injury induced by peroxynitrite production and PARS activation in macrophages collected from rats subjected to carrageenan‐induced pleurisy. Macrophages harvested from the pleural cavity exhibited a significant production of peroxynitrite, as measured by the oxidation of the fluorescent dye dihydrorhodamine 123. Furthermore, carrageenan‐induced pleurisy caused a suppression of macrophage mitochondrial respiration, DNA strand breakage, activation of PARS, and reduction of cellular levels of NAD+. In vivo treatment with melatonin [12.5 or 25 or 50 mg/kg, intraperitoneally (i.p.), 30 min before carrageenan] significantly reduced peroxynitrite formation in a dose‐dependent manner and prevented the appearance of DNA damage, decrease in mitochondrial respiration, loss of cellular levels of NAD+, and PARS activation. Our study supports the view that the antioxidant and anti‐inflammatory effect of melatonin is also correlated with the inhibition of peroxynitrite production and PARS activation. In conclusion, melatonin may be a novel pharmacological approach to prevent cell injury in acute inflammation.
The effects of adrenergic agonists on c‐fos, jun‐B and nitric oxide synthase‐like immunoreactivities in cultured rat pinealocytes
Authors: Chetsawang Banthit , Govitrapong Piyarat , Møller Morten
Published: March 2003

Pineal melatonin synthesis is enhanced by the activation of adrenergic and opioid receptors. The precise mechanism of how signal transduction is affected by the activation of these receptors, leading to an increase in melatonin synthesis, is not clear. An attempt has been made to investigate the effects of the activation of these two types of receptors that might lead to the induction of immediate early genes (IEGs) and nitric oxide synthase (NOS) expressions. The present study shows that the stimulation of cultured pinealocytes by 1 μM epinephrine (an α‐ and β‐adrenergic agonist) for 2 h increased the number of c‐fos immunoreactive (IR) cells, and that this stimulatory effect was abolished by adding 10 μM prazosin (an α‐adrenergic antagonist) to the culture medium. No significant change of c‐fos‐IR cells was found when cells were treated with either 1 μM isoproterenol (a β‐adrenergic agonist) or 100 μM morphine (an opioid agonist). Neither epinephrine (1 μM), isoproterenol (1 μM) nor morphine (100 μM) altered jun‐B‐IR cells in the culture. With regard to NOS, the number of IR cells was slightly increased after 2 h of incubation by 10 μM isoproterenol, whereas morphine (100 μM) and epinephrine (10 μM) did not change the number of IR cells. The results of this experiment have demonstrated that c‐fos expression is induced by α‐adrenergic but not β‐adrenergic agonists, nor by opioid agonists. Neither α‐, β‐adrenergic nor opioid agonists had any significant effect on jun‐B, whereas NOS was slightly increased by isoproterenol.
Melatonin protects SHSY5Y neuroblastoma cells from cobalt‐induced oxidative stress, neurotoxicity and increased β‐amyloid secretion
Authors: Olivieri Gianfranco , Hess Christian , Savaskan Egemen , Ly Chuong , Meier Fides , Baysang Ginette , Brockhaus Manfred , Müller‐Spahn Franz
Published: March 2003

Heavy metals are increasingly being implicated as causative agents in neurodegenerative diseases such as Alzheimer's disease (AD). Cobalt, a positively charged transition metal, has previously been shown to be in elevated levels in the brain of AD patients compared with age‐matched controls. In this study, we investigate the effects of cobalt as an inducer of oxidative stress/cell cytotoxicity and the resultant metabolic implications for neural cells. We show that cobalt is able to induce cell cytotoxicity (reduced MTT metabolism) and oxidative stress (reduced cellular glutathione). The pre‐treatment of cells with the pineal indoleamine melatonin, prevented cell cytotoxicity and the induction of oxidative stress. Cobalt treatment of SHSY5Y cells increased the release of β‐amyloid (Aβ) compared with untreated controls (ratio Aβ 40/42). Melatonin pre‐treatment reversed the deleterious effects of cobalt. These findings are significant as cobalt is an essential nutritional requirement, usually bound to cobalamin (vitamin B12), for all animals which in the unbound form could lead to neurotoxicity.
Melatonin potentiates 5‐HT1A receptor activation in rat hypothalamus and results in hypothermia
Authors: Lin Mao‐Tsun , Chuang Jih‐Ing
Published: March 2003

Effects of melatonin on both thermoregulatory responses and hypothalamic serotonin release were assessed in unanesthetized rats at three different ambient temperatures (Ta). Systemic administration of melatonin (30–120 mg/kg, i.p) caused a decrease in both colonic temperature and hypothalamic serotonin (5‐HT) release in rats at both Ta 8 and 22°C. The hypothermia was brought about by a decrease in metabolic rate at Ta 8°C, whereas at Ta 22°C the hypothermia was produced by both a decrease in metabolic rate and an increase in cutaneous temperature. However, in the heat (Ta 31°C), neither thermoregulatory responses nor hypothalamic 5‐HT release was affected by the same amount of administered melatonin. The melatonin‐induced hypothermia and decreased 5‐HT release in the hypothalamus were attenuated by selective depletion of brain 5‐HT produced by intracerebroventricular injection of 5,7‐dihydroxytryptamine. Furthermore, the melatonin‐induced hypothermia was almost completely abolished by treatment with a 5‐HT2A receptor agonist (DOI) or a 5‐HT1A receptor antagonist [(−)‐pindolol]. The data indicate that melatonin potentiates the 5‐HT1A receptor activation in the hypothalamus and results in hypothermic effects which can be antagonized by the expected hyperthermic effect of DOI.
Protective effects of melatonin on myocardial ischemia/reperfusion injury in vivo
Authors: Lee Yen‐Mei , Chen Huey‐Rue , Hsiao George , Sheu Joen‐Rong , Wang Jhi‐Joung , Yen Mao‐Hsiung
Published: March 2003

The production of oxygen free radicals has been strongly implicated as an important pathophysiological mechanism mediating myocardial ischemia/reperfusion (I/R) injury. Various antioxidants have cardioprotective effects. Melatonin, an indoleamine synthesized by the pineal gland, is a potent antioxidant and a direct free radical scavenger. This is the first in vivo study to evaluate the effect of melatonin (0.5, 1.0, and 5.0 mg/kg, i.v. bolus) on myocardial I/R injury in anesthetized Sprague–Dawley rats. Results demonstrate that pretreatment with intermediate or high doses of melatonin (1.0 and 5.0 mg/kg) at 10 min before left coronary artery occlusion markedly suppressed ventricular tachycardia (VT) and ventricular fibrillation (VF), while reducing the total number of premature ventricular contractions and total duration of VT and VF that occurred during the 45‐min ischemic period. Pretreatment with melatonin dramatically improved survival rate of rats when compared with the I/R‐only group. After 1‐hr reperfusion, melatonin caused a significant reduction in infarct size when compared with I/R‐only group. Meanwhile, pretreatment with melatonin (1.0 mg/kg) significantly reduced superoxide anion production and myeloperoxidase activity; the latter is an index of neutrophil infiltration in the ischemic myocardium. Additionally, pretreatment with melatonin (1.0 and 5.0 mg/kg) significantly attenuated ventricular arrhythmias and mortality elicited by reperfusion following 5‐min ischemia. In conclusion, melatonin suppresses ischemia‐ and reperfusion‐induced ventricular arrhythmias and reduces infarct size resulting from I/R injury. The pronounced cardioprotective activity of melatonin may be mediated by its antioxidant activity and by its capacity for neutrophil inhibition in myocardial I/R.
Melatonin reduces non‐adrenergic, non‐cholinergic relaxant neurotransmission by inhibition of nitric oxide synthase activity in the gastrointestinal tract of rodents in vitro
Authors: Storr M. , Koppitz P. , Sibaev A. , Saur D. , Kurjak M. , Franck H. , Schusdziarra V. , Allescher H.‐D.
Published: March 2003

The aim of the present study was to investigate the effects of melatonin on non‐adrenergic, non‐cholinergic (NANC) relaxant neurotransmission in the gastrointestinal tract, which is mainly mediated by nitrergic and peptidergic mechanisms. Melatonin (10−7–10−3 M) had no effect on the basal tonus of the rat gastric fundus smooth muscle. Relaxant responses following electrical stimulation(40 V; 0.5 ms pulse duration; 10 s stimulation duration) under NANC conditions on a 5‐hydroxytryptamine (5‐HT, 10−7 M) contraction plateau were elicited at frequencies in the range of 0.5–16 Hz. Melatonin significantly reduced these inhibitory NANC responses (16 Hz without melatonin: –103 ± 6.3%; melatonin 10−5 M: –80.4 ± 7.5%; melatonin 10−4 M: –39.1 ± 17.1%). Intracellular recording was carried out in a mouse colonic preparation. Electrical neural stimulation of the mouse colonic neurons caused biphasic intracellular hyperpolarization in smooth‐muscle cells. The initial fast component is apamin‐sensitive, and the following slow component is dependent on nitrergic mechanisms, as it is abolished in the presence of NG‐nitro‐L‐arginine (L‐NNA). Melatonin significantly reduced the nitric oxide‐dependent slow component of neurally transmitted hyperpolarization, whereas the initial fast component was left unchanged. In a synaptosomal preparation of the enteric nervous system of rat intestine, enzymatic nitric oxide synthase (NOS) activity was significantly reduced by melatonin at concentrations ranging from 10−7 to 10−4 M (basal preparation including cofactors: 61.2 ± 9.4 fmol/mg; melatonin 10−4 M: 39.2 ± 6.9 fmol/mg). Reverse transcriptase‐polymerase chain reaction (RT‐PCR) studies were conducted to investigate the melatonin receptors (mt1, MT2 and MT3) present in the esophagus, stomach and ileum of the rat. The presence of mt1 mRNA expression alone, but not of mRNA expression for MT2 or MT3, was demonstrated in the tissues. In conclusion, this study demonstrates that melatonin reduces the functional inhibitory NANC response. It shows that this effect may be the result of a reduction of the nitrergic component of the smooth‐muscle inhibitory junction potential (IJP) and related to direct inhibition of NOS activity in enteric synaptosomes. The presence of mt1 receptor transcripts adds supportive evidence for a possible physiological role of melatonin within the enteric nervous system.
Interferon‐gamma independent oxidation of melatonin by macrophages
Authors: Rodrigues Maria R. , Rodriguez Dunia , Henrique Catalani Luiz , Russo Momtchilo , Campa Ana
Published: March 2003

Abstract: Mononuclear phagocytes appear to synthesize kynurenine‐like products from the oxidation of biologically active indole compounds including melatonin, catalyzed by interferon (IFN)‐γ‐inducible enzyme indoleamine 2,3‐dioxygenase (IDO). Concanavalin A (Con A) is a plant lectin that induces interferon‐gamma (IFN‐γ) production by T cells. In this study we investigated whether Con A‐primed peritoneal macrophages are able to oxidize melatonin to N1‐acetyl‐N2‐formyl‐5‐methoxykynuramine (AFMK). The AFMK production was accompanied by chemiluminescence. It was found that Con A‐primed but not resident macrophages produce AFMK. Surprisingly, Con A‐primed macrophages from IFN‐γ‐deficient mice were as effective as macrophages from IFN‐γ‐sufficient mice in oxidizing melatonin. Moreover, addition of an inhibitor of IDO (1‐methyltryptophan) did not affect melatonin oxidation. Con A‐primed but not resident macrophages have a significant content of myeloperoxidase (MPO) and inhibition of MPO by azide completely blocked chemiluminescence and AFMK production. Thus, our findings provide evidence that melatonin oxidation by macrophages may occur through a mechanism dependent of MPO and independent of IFN‐γ and IDO activity.
Estrogen‐dependent tensile properties of the rabbit knee medial collateral ligament
Authors: Räsänen T. , Messner K.
Published: March 2003

The influence of oophorectomy or continuous administration of estradiol on the tensile properties of the rabbit knee medial collateral ligament was investigated. Young postpubertal female New Zealand white rabbits were either oophorectomized or underwent a sham operation. The sham‐operated animals received in addition a daily dosage of 4 mg 17β‐estradiol. After 5 months the animals were killed, and the material properties of the bone‐ligament‐bone complex in one knee were determined using a material testing machine and video system, and compared to non‐treated control animals. There was no difference in elastic modulus between the groups. However, the ligaments from low‐dose estrogen‐treated animals had a smaller cross‐sectional area and a higher ultimate tensile strength than those from controls or oophorectomized rabbits (P<0.04–0.0003).
Can the hamstring muscles protect the anterior cruciate ligament during a side‐cutting maneuver?
Authors: Simonsen E. B. , Magnusson S. P. , Bencke J. , Næsborg H. , Havkrog M. , Ebstrup J. F. , Sørensen H.
Published: March 2003

Because anterior cruciate ligament (ACL) injuries are common in European handball the present study assessed knee joint shear forces to estimate ACL loading in six elite female handball players during a side‐cutting maneuver. A pilot investigation in three dimensions showed that peak moments occurred in the sagittal plane at a high velocity. Therefore, analysis of the movement was performed in two dimensions using high‐speed cinematography, ground reaction forces, and electromyography (EMG). Film and force plate data allowed for calculation of net joint moments (inverse dynamics), estimates of instantaneous muscle‐tendon lengths, contraction velocities, and peak loading of the ACL. During the breaking phase of the maneuver the peak knee joint moment was 239 Nm (99–309), which yielded an ACL‐load of 520 N (215–673). The corresponding peak EMG amplitudes for the hamstring muscles were 34–39% of maximum EMG. During the breaking phase the quadriceps muscle contracted eccentrically with a velocity of 216–253% fiber length/s. In constrast, the hamstring muscles contracted concentrically with a velocity of 222–427% fiber length/s. These results suggest that a side‐cutting maneuver produces loads that are insufficient to rupture the ACL. Furthermore, the rapid concentric hamstring contraction suggests that even during maximal activation, the ability of the hamstrings to reduce the ACL load is marginal.
Effects of walking on bone quality as determined by ultrasound in the elderly
Authors: Yanagimoto Y. , Oshida Y. , Sato Y.
Published: March 2003

In the present study, measurements of broadband ultrasound attenuation (BUA) were used as indications of bone quality in elderly residents of an old people’s home. To investigate the possibility of using number of steps walked as an indicator of the role played by exercise on bone quality, we studied the relationship between BUA and the number of steps, taken as a measure of the number of impacts against the ground placing a direct mechanical stress on the bones. The subjects were 59 healthy elderly women with a mean age of 78±8 years who had no impairments in walking or daily life. A statistical analysis of the relationship between age, number of steps, BUA, stride length, and walking speed revealed that number of steps and BUA declined with age, and that BUA increased with a greater number of steps. Among other factors, age was found to be negatively correlated with weight (r=−0.343, P<0.01), BUA (r=−0.542, P<0.001), total number of steps (r=−0.524, P<0.001) and past exercise habits (r=−0.425, P<0.001). There were positive correlations between BUA and total number of steps (r=0.606, P<0.001), walking speed (r=0.460, P<0.001), stride length (r=0.373, P<0.01) and past exercise habits (r=0.429, P<0.001). These results show that the benefit to bone quality increases the more a person walks in daily life. The above demonstrates that degree of walking activity is positively correlated to bone quality among the elderly. These results also suggest that, when investigating the relationship between exercise and bone quality, one should look not only at distance and walking speed, but also number of steps as the number of impacts against the ground that place a direct mechanical stress on the bones. This may prove to be a useful indicator of bone quality in future use.
Structure of the tendon connective tissue
Authors: Kannus P.
Published: March 2003

Tendons consist of collagen (mostly type I collagen) and elastin embedded in a proteoglycan‐water matrix with collagen accounting for 65–80% and elastin approximately 1–2% of the dry mass of the tendon. These elements are produced by tenoblasts and tenocytes, which are the elongated fibroblasts and fibrocytes that lie between the collagen fibers, and are organized in a complex hierarchical scheme to form the tendon proper. Soluble tropocollagen molecules form cross‐links to create insoluble collagen molecules which then aggregate progressively into microfibrils and then into electronmicroscopically clearly visible units, the collagen fibrils. A bunch of collagen fibrils forms a collagen fiber, which is the basic unit of a tendon. A fine sheath of connective tissue called endotenon invests each collagen fiber and binds fibers together. A bunch of collagen fibers forms a primary fiber bundle, and a group of primary fiber bundles forms a secondary fiber bundle. A group of secondary fiber bundles, in turn, forms a tertiary bundle, and the tertiary bundles make up the tendon. The entire tendon is surrounded by a fine connective tissue sheath called epitenon. The three‐dimensional ultrastructure of tendon fibers and fiber bundles is complex. Within one collagen fiber, the fibrils are oriented not only longitudinally but also transversely and horizontally. The longitudinal fibers do not run only parallel but also cross each other, forming spirals. Some of the individual fibrils and fibril groups form spiral‐type plaits. The basic function of the tendon is to transmit the force created by the muscle to the bone, and, in this way, make joint movement possible. The complex macro‐ and microstructure of tendons and tendon fibers make this possible. During various phases of movements, the tendons are exposed not only to longitudinal but also to transversal and rotational forces. In addition, they must be prepared to withstand direct contusions and pressures. The above‐described three‐dimensional internal structure of the fibers forms a buffer medium against forces of various directions, thus preventing damage and disconnection of the fibers.
Sonography of tendons
Authors: Rasmussen O. S.
Published: March 2003

Recent advances in ultrasound technology and the development of high‐resolution ultrasound transducers have enabled detailed depiction of superficial musculoskeletal structures. While in the past considered as complementary to magnetic resonance (MR) imaging, modern ultrasound has clearly become competitive. It is now the imaging modality of choice for evaluating tendon pathology. The major advantages of ultrasound include dynamic evaluation of structures, low cost and wide availability. The main disadvantage is a high degree of operator dependency. This article discusses the current ultrasound characteristics of normal tendon, the artefacts that may mimic tendon disease and reviews the applications of ultrasound to the evaluation of tendon disorders, including tendon tears, tendinitis, tenosynovitis, ganglion cysts and bursitis.
Body proportions, body composition and pubertal development of children in competitive sports
Authors: Damsgaard R. , Bencke J. , Matthiesen G. , Petersen J. H. , Müller J.
Published: March 2003

The aim of this study was primarily to investigate anthropometric variables, body composition and pubertal development in children aged 9–13 participating in competitive sports. Secondly, the influence of age, sport, training hours and pubertal development/maternal menarcheal age on body composition and pubertal development was explored. A total of 183 (96 girls, 87 boys) children performing swimming (Sw), tennis (Te), European team handball (TH), and gymnastics (Gy) took part in the study. Anthropometric measurements and pubertal development were determined. The participants completed a questionnaire regarding hours of training per week and maternal menarcheal age. Significant differences in stature (z‐scores) were found in both boys (Sw=0.06; Te=0.04; TH=0.05; Gy=−0.66, P<0.004) and girls (Sw=0.12; Te=0.19; TH=0.25; Gy=−0.96, P<0.004). In girls, sum of skinfolds in millimetres (Sw=33.4; Te=33.3; TH=41.0; Gy=27.2, P<0.02) and body mass index z‐scores (SW=0.00; Te=−0.27; TH=0.35; Gy=−0.25, P<0.001) were different between the sports. A regression analysis revealed that in girls, age and maternal menarcheal age were significantly associated with pubertal development (P<0.005 and P<0.01), respectively, and sport was associated with the sum of skinfolds (P<0.05). In boys, only age was significantly associated with pubertal development (P<0.005). In conclusion, anthropometric and body composition differences exist in athletes of both sexes from different sports but are more evident in females. Most importantly, we did not find any effect of training on body composition or pubertal development, confirming previous data that children in competitive sports are selected due to constitutional factors.
Flexibility and passive resistance of the hamstrings of young adults using two different static stretching protocols
Authors: Chan S. P. , Hong Y. , Robinson P. D.
Published: March 2003

This investigation determined the effects of a static stretching program with different stretching protocols on the flexibility and passive resistance of the hamstrings of young adults. Forty healthy subjects (24 males and 16 females) aged 18 to 30 years were randomly assigned to one of four groups. The two training groups underwent static stretch training of the hamstrings either with a four‐week protocol or with an eight‐week protocol. The other two groups acted as control groups. A significant increase in flexibility of hamstrings was found in both of the two training groups (P<0.05). No difference was found in the range of motion gained between the two training groups. An increase in passive resistance at the corresponding maximal joint angle was only demonstrated in the four‐week training group (P<0.05). Both protocols are effective in terms of improving flexibility of hamstrings. However, if injury is reduced when there is relatively lower passive resistance at the end‐of‐range, then the eight‐week training regimen would be recommended.
Chronic stretching and running economy
Authors: Nelson A. G. , Kokkonen J. , Eldredge C. , Cornwell A. , Glickman‐Weiss E.
Published: March 2003

Research demonstrates an inverse relationship between the range of motion of selected joint movements (flexibility) and running economy. Since stretching exercises have been shown to increase joint range of motion, stretching exercises may be contraindicated for endurance running performance. Hence, this study investigated the influence of a 10‐week program of stretching exercises on the oxygen costs of a 10 min sub‐maximal (approx. 70% peak VO2) treadmill run. Thirty‐two (16 female, 16 male) physically active, treadmill accommodated, college students participated in the study. All participants maintained their current activity level, with half the participants (8 female, 8 male) adding a 40 min, 3 days per week session of thigh and calf muscle stretching exercises. After 10 weeks, the stretching group (STR) exhibited a significant (P<0.05) increase (3.1±2.2 cm) in the sit‐and‐reach, while the non‐stretching group (CON) experienced no significant (P>0.05) change (0.0±0.4 cm). However, neither the STR nor the CON exhibited a significant (P>0.05) change in the O2 cost for the submaximal run. It is concluded, therefore, that a chronic stretching program does not necessarily negatively influence running economy.
Analytic quantification of phototype‐related regional skin complexion
Authors: Hermanns Jean François , Petit Ludivine , Hermanns‐Lê Trinh , Piérard Gérald E.
Published: March 2003

Background/aims: Narrow‐band spectrophotometry that yields melanin (M) and erythema (E) indexes is a convenient method for assessing skin colour. The objective of the study was to assess the phenotype‐associated body site differences in skin complexion.

Method: The Mexameter MX16 was used to record the skin colour in 137 subjects. Evaluations were made on the inner arm, volar forearm, dorsal forearm and forehead.

Results: The inner arm and volar forearm had similar values for each of the colorimetric values. The dorsal forearm exhibited the highest M values, and the forehead had the highest E values. Significant positive linear correlations were found between M values from the distinct body sites. The E values showed a correlation only between the inner arm and the volar forearm.

Conclusion: Regional skin colour differences are obvious. Sun‐exposed skin does not react in the same way on the forearm and forehead. Regional variations in M can be predicted from the innate M value. In contrast, the regional variability in E is highly variable among subjects.
Effects of ageing on dermal echogenicity
Authors: Gniadecka M.
Published: March 2003

Background/aims: Changes in the dermis associated with ageing can be detected by high‐frequency skin ultrasonography. In photoaged skin, this technique shows a subepidermal low echogenic band (SLEB) that is probably an ultrasound manifestation of elastosis and oedema in the papillary dermis. Since some authors found an association between age and SLEB thickness or its echogenicity on exposed sites, it has been proposed to use these parameters to quantify skin photoageing.

Methods: To determine whether SLEB can be used as a quantitative marker of ageing, its prevalence was determined on forearm skin in a group of 55 individuals (age 18–57 years). The size of SLEB has been measured by quantifying the number of low echogenic pixels in the subepidermal area, which is an accurate method for assessing SLEB severity.

Results: The prevalence of SLEB increased with age, but SLEB was also present in young subjects. The echogenicity of the subepidermal area did not show any age dependence. However, when a ratio of echogenicity between upper and lower dermis was calculated, a linear dependence on age was found.

Conclusions: This study indicates that skin echogenicity measured as a ratio between the upper and lower dermis may be used to objectively estimate photoageing.
A noninvasive method to assess skin irritation and compromised skin conditions using simple tape adsorption of molecular markers of inflammation
Authors: Perkins Mary A. , Osterhues Marcia A. , Farage Miranda A. , Robinson Michael K.
Published: March 2003

Background/aims: We have developed a simple noninvasive method to assess inflammatory changes in human skin, even in the absence of visible clinical irritation. Our approach is based on a simple tape (Sebutape®) adsorption method to recover molecular mediators of skin inflammation (e.g., cytokines). This procedure has been used to investigate baseline cytokine levels on skin, to assess normal skin condition and to evaluate changes due to chemical insult, existing dermatitis, or sun exposure.

Methods: In clinical studies, Sebutape was applied to normal appearing uncompromised skin, as well as to compromised (diaper or heat rash), chemically treated (sodium laurel sulfate), or sun‐exposed skin. Sebutape was applied to the skin for a 1 min collection interval. Tapes were extracted in saline using a 10 min sonication, and the extracts were analyzed for human interleukin‐1α (IL‐1α), IL‐1 receptor antagonist (IL‐1RA) and IL‐8 using commercial immunoassay test kits. The cytokine levels recovered from each tape extract were normalized to total protein (TP) levels. In infant product use tests, the severity of skin irritation (diaper and heat rash or erythema) was also assessed using a visual grading scale.

Results: The method itself caused minimal, if any, skin damage. Additionally, Sebutape was shown to quantitatively adsorb detectable levels of cytokine from normal‐appearing (control) or compromised (e.g., rashed or chemically treated) skin. In infant studies, significant increases in IL‐1α levels were found in skin exhibiting diaper rash, heat rash and erythema compared with normal appearing control skin sites. When these results were normalized to total protein levels recovered from each tape, the significance was maintained. A positive correlation (r2=0.82) existed between IL‐1RA levels and diaper rash severity. Significant increases in IL‐8 levels were recovered from diaper rash versus control skin sites. There were differences in baseline cytokine levels in normal skin related to body site and sun exposure. The IL‐1RA/IL‐1α ratios for sun‐exposed skin of the face and lower leg were significantly (P<0.05) higher (3–6‐fold) than those for skin sites that typically receive minimal sun exposure (i.e., underarm, upper leg and upper back). There was a significant increase in IL‐1α and a directional increase in IL‐8 levels in adult skin sites treated with the irritant, sodium lauryl sulfate, even in the absence of visible skin irritation (erythema).

Conclusion: Our results demonstrate that this method is a useful noninvasive technique for assessing skin inflammatory events. In addition, the method is simple and easily applied in a clinical setting, whether on infants or adults.
Adipsin and the Glucose Transporter GLUT4 Traffic to the Cell Surface via Independent Pathways in Adipocytes
Authors: Millar Caroline A. , Meerloo Timo , Martin Sally , Hickson Gilles R.X. , Shimwell Neil J. , Wakelam Michael J.O. , James David E. , Gould Gwyn W.
Published: March 2003

Insulin increases the exocytosis of many soluble and membrane proteins in adipocytes. This may reflect a general effect of insulin on protein export from the trans Golgi network. To test this hypothesis, we have compared the trafficking of the secreted serine protease adipsin and the integral membrane proteins GLUT4 and transferrin receptors in 3T3‐L1 adipocytes. We show that adipsin is secreted from the trans Golgi network to the endosomal system, as ablation of endosomes using transferrin‐HRP conjugates strongly inhibited adipsin secretion. Phospholipase D has been implicated in export from the trans Golgi network, and we show that insulin stimulates phospholipase D activity in these cells. Inhibition of phospholipase D action with butan‐1‐ol blocked adipsin secretion and resulted in accumulation of adipsin in trans Golgi network‐derived vesicles. In contrast, butan‐1‐ol did not affect the insulin‐stimulated movement of transferrin receptors to the plasma membrane, whereas this was abrogated following endosome ablation. GLUT4 trafficking to the cell surface does not utilise this pathway, as insulin‐stimulated GLUT4 translocation is still observed after endosome ablation or inhibition of phospholipase D activity. Immunolabelling revealed that adipsin and GLUT4 are predominantly localised to distinct intracellular compartments. These data suggest that insulin stimulates the activity of the constitutive secretory pathway in adipocytes possibly by increasing the budding step at the TGN by a phospholipase D‐dependent mechanism. This may have relevance for the secretion of other soluble molecules from these cells. This is not the pathway employed to deliver GLUT4 to the plasma membrane, arguing that insulin stimulates multiple pathways to the cell surface in adipocytes.
Membrane Tethering and Fusion in the Secretory and Endocytic Pathways
Authors: Waters M. Gerard , Hughson Frederick M.
Published: March 2003

Studies of intracellular trafficking over the past decade or so have led to striking advances in our understanding of the molecular processes by which transport intermediates dock and fuse. SNARE proteins play a central role, assembling into complexes that bridge membranes and may catalyze membrane fusion directly. In general, different SNARE proteins operate in different intracellular trafficking pathways, so recent reports that SNARE assembly in vitro is promiscuous have come as something of a surprise. We propose a model in which proper SNARE assembly is under kinetic control, orchestrated by members of the Sec1 protein family, small GTP‐binding Rab proteins, and a diverse assortment of tethering proteins.
Reconstitution of Clathrin‐Independent Endocytosis at the Apical Domain of Permeabilized MDCK II cells: Requirement for a Rho‐Family GTPase
Authors: Garred Øystein , Kjersti Rodal Siv , Van Deurs Bo , Sandvig Kirsten
Published: March 2003

This paper studies the endocytosis of ricin at the apical pole of polarized MDCK II cells after permeabilization of the cells basolaterally with streptolysin O. Ricin endocytosis after the addition of cytosol with an ATP‐regenerating system was 2–3‐fold higher than after the addition of a transport medium. A similar increase in ricin endocytosis was obtained by reconstitution of dialyzed cytosol with the nonhydrolyzable GTP analog, GTPγS, in the presence of an ATP‐regenerating system. The nonhydrolyzable GDP analog, GDPβS, did not increase ricin uptake. In contrast to the data obtained with ricin, GTPγS was found to inhibit apical transferrin uptake in MDCK II cells transfected with the human transferrin receptor, and the data thus imply that GTPγS supports clathrin‐independent endocytosis. Electron microscopy (EM) demonstrated that free endocytic vesicles were formed from the apical pole of permeabilized MDCK II cells in the presence of GTPγS and that both a ricin–HRP conjugate, HRP, and cationized gold were endocytosed. Ricin endocytosis in the presence of intact cytosol, as well as GTPγS‐stimulated ricin uptake, was inhibited by Clostridium botulinum C3 transferase, an enzyme found to inactivate Rho proteins. The data demonstrate that apical clathrin‐independent endocytosis functions in the presence of GTPγS, and suggest that one or more of the small GTP binding proteins of the Rho family is involved in regulation of the apical clathrin‐independent endocytosis in MDCK II cells.
NBD‐Labeled Phosphatidylcholine and Phosphatidylethanolamine are Internalized by Transbilayer Transport across the Yeast Plasma Membrane
Authors: Grant Althea M. , Hanson Pamela K. , Malone Lynn , Wylie Nichols J.
Published: March 2003

The internalization and distribution of fluorescent analogs of phosphatidylcholine (M‐C6‐NBD‐PC) and phosphatidylethanolamine (M‐C6‐NBD‐PE) were studied in Saccharomyces cerevisiae. At normal growth temperatures, M‐C6‐NBD‐PC was internalized predominantly to the vacuole and degraded. M‐C6‐NBD‐PE was internalized to the nuclear envelope/ER and mitochondria, was not transported to the vacuole, and was not degraded. At 2°C, both were internalized to the nuclear envelope/ER and mitochondria by an energy‐dependent, N‐ethylmaleimide‐sensitive process, and transport of M‐C6‐NBD‐PC to and degradation in the vacuole was blocked. Internalization of neither phospholipid was reduced in the endocytosis‐defective mutant, end4‐1. However, following pre‐incubation at 37°C, internalization of both phospholipids was inhibited at 2°C and 37°C in sec mutants defective in vesicular traffic. The sec18/NSF mutation was unique among the sec mutations in further blocking M‐C6‐NBD‐PC translocation to the vacuole suggesting a dependence on membrane fusion. Based on these and previous observations, we propose that M‐C6‐NBD‐PC and M‐C6‐NBD‐PE are transported across the plasma membrane to the cytosolic leaflet by a protein‐mediated, energy‐dependent mechanism. From the cytosolic leaflet, both phospholipids are spontaneously distributed to the nuclear envelope/ER and mitochondria. Subsequently, M‐C6‐NBD‐PC, but not M‐C6‐NBD‐PE, is sorted by vesicular transport to the vacuole where it is degraded by lumenal hydrolases.
Nuclear Localization of the Saccharomyces cerevisiae HMG Protein NHP6A Occurs by a Ran‐Independent Nonclassical Pathway
Authors: Yen Yi‐Meng , Roberts Paul M. , Johnson Reid C.
Published: March 2003

The Saccharomyces cerevisiae non‐histone protein 6‐A (NHP6A) is a member of the high‐mobility group 1/2 protein family that bind and bend DNA of mixed sequence. NHP6A has only one high‐mobility group 1/2 DNA binding domain and also requires a 16‐amino‐acid basic tail at its N‐terminus for DNA binding. We show in this report that nuclear accumulation of NHP6A is strictly correlated with its DNA binding properties since only nonhistone protein 6 A–green fluorescent protein chimeras that were competent for DNA binding were localized to the nucleus. Despite the requirement for basic residues within the N‐terminal segment for DNA binding and nuclear accumulation, this region does not appear to contain a nuclear localization signal. Moreover, NHP6A does not bind to the yeast nuclear localization signal receptor SRP1 and nuclear targeting of NHP6A does not require the function of the 14 different importins. Unlike histone H2B1 which contains a classical nuclear localization signal, entry of NHP6A into the nucleus was found to be independent of Ran as judged by coexpression of Ran GTPase mutants and was shown to occur at 0 °C after a 15‐min induction. These unusual properties lead us to suggest that NHP6A entry into the nucleus proceeds by a nonclassical Ran‐independent pathway.
Characterization of Salmonella‐Induced Filaments (Sifs) Reveals a Delayed Interaction Between Salmonella‐Containing Vacuoles and Late Endocytic Compartments
Authors: Brumell John H. , Tang Patrick , Mills Scott D. , Finlay B. Brett
Published: March 2003

Salmonella typhimurium is a facultative intracellular pathogen that colonizes host cells throughout the course of infection. A unique feature of this pathogen is its ability to enter into (invade) epithelial cells and elongate the vacuole within which it resides into tubular structures called Salmonella‐induced filaments (Sifs). In this study we sought to characterize the mechanism of Sif formation by immunofluorescence analysis using subcellular markers. The late endosomal lipid lysobisphosphatidic acid associated in a punctate pattern with the Salmonella‐containing vacuole, starting 90 min after infection and increasing thereafter. Lysobisphosphatidic acid‐rich vesicles were also found to interact with Sifs, at numerous sites along the tubules. Similarly, cholesterol‐rich vesicles were also found in association with intracellular bacteria and Sifs. The lysosomal hydrolase cathepsin D was present in Sifs, both in a punctate pattern and, at later times, predominantly in an uninterrupted linear pattern. Rab7 associated with Sifs and expression of the N125I dominant negative mutant of this GTPase inhibited Sif formation. Transfection of HeLa cells with a vector encoding SifA fused to the green fluorescent protein caused swelling and aggregation of lysobisphosphatidic acid‐containing compartments, suggesting that this virulence factor directs membrane fusion events involving late endosomes. Our findings demonstrate that Sif formation involves fusion of late endocytic compartments with the Salmonella‐containing vacuole, and suggest that SifA modulates this event.
Yeast Ribosomes Bind to Highly Purified Reconstituted Sec61p Complex and to Mammalian p180
Authors: Morrow Michael W. , Brodsky Jeffrey L.
Published: March 2003

To determine whether the yeast Sec61p translocation pore is a high‐affinity ribosome receptor in the endoplasmic reticulum, we isolated the Sec61p complex using an improved protocol in which contaminants found previously to be associated with the complex are absent. The purified complex, which contains Sec61p with an amino terminal hexahistidine tag, was active since it rescued a sec61–3 post‐translational translocation defect in a reconstituted system. Co‐reconstitution of the Sec61p and Sec63p complexes into liposomes failed to support post‐translational translocation, suggesting that Sec62p is required for this process. By Scatchard analysis, the purified Sec61p complex bound to yeast ribosomes when reconstituted into liposomes with a KD of 5.6 nm, a value similar to the KD obtained when ribosome binding to total microsomal protein was measured (2.7 nm). In addition, a mammalian protein, p180, which has been proposed to be a ribosome receptor, was expressed in yeast, and endoplasmic reticulum‐derived microsomes isolated from this strain exhibited ∼2.3‐fold greater binding to yeast ribosomes. Despite this increase in ribosome binding, neither co‐ nor post‐translational translocation was compromised in vivo. In sum, our data suggest that the Sec61p complex is a ribosome receptor in the yeast endoplasmic reticulum membrane.
Dyneins Have Run Their Course in Plant Lineage
Authors: Lawrence Carolyn J. , Morris N. Ronald , Meagher Richard B. , Dawe R. Kelly
Published: March 2003

Flowering plant genomes lack flagellar and cytoplasmic dyneins as well as the proteins that make up the dynactin complex. The mechanisms for organizing the Golgi apparatus, establishing spindle poles, and moving nuclei, vesicles, and chromosomes in flowering plants must be fundamentally different from those in other systems where these processes are dependent upon dynein and dynactin.
Melanophilin, the Product of the Leaden Locus, is Required for Targeting of Myosin‐Va to Melanosomes
Authors: Provance D. William , James Ted L. , Mercer John A.
Published: March 2003

The formation of complex subcellular organelles requires the coordinated targeting of multiple components. Melanosome biogenesis in mouse melanocytes is an excellent model system for studying the coordinated function of multiple gene products in intracellular trafficking. To begin to order events in melanosome biogenesis and distribution, we employed the classical coat‐color mutants ashen, dilute, and leaden, which affect melanosome distribution, but not melanin synthesis. The loci have been renamed Rab27a, Myo5a, and Mlph for their gene products. While each of the three loci has been shown to be required for melanosome distribution, the point(s) at which each acts is unknown. We have utilized primary melanocytes to examine the interdependencies between rab27a, myosin‐Va, and melanophilin. The localization of rab27a to melanosomes did not require the function of either myosin‐Va or melanophilin, but leaden function was required for the association of myosin‐Va with melanosomes. In leaden melanocytes permeabilized before fixation, myosin‐Va immunoreactivity was greatly attenuated, suggesting that myosin‐Va is free in the cytoplasm. Finally, we have complemented both the leaden and ashen phenotypes by cell fusion and observed redistribution of mature melanosomes in the absence of both protein and melanin synthesis. Together, our data suggest a model for the initial assembly of the machinery required for melanosome distribution.