bluuebunny/crossref_metadata_2025 huggingface dataset:
Melatonin protects SHSY5Y neuronal cells but not cultured astrocytes from ischemia due to oxygen and glucose deprivation
Authors: Pei Zhong , Cheung Raymond Tak Fai
Published: March 2003

Abstract: As a potent free radical scavenger and antioxidant, melatonin protects brain tissue against ischemia–reperfusion injury, partly via suppression of ischemia‐induced production of nitric oxide, when given before ischemia–reperfusion or within 2 hr of onset of ischemia. In this study, we examined the neuroprotective effect of melatonin in an in vitro model of ischemia. Primary cultured astrocytes were subjected to 4 or 8 hr of oxygen–glucose deprivation (OGD), and cultured SHSY5Y human neuronal cells were exposed to 1 hr of OGD. Melatonin was added to the medium at the commencement of OGD to achieve different final concentrations, and cell death was quantified using the measurement of 3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyl‐tetrazolium bromide (MTT) at 24 hr after reversion of OGD. Treatment with melatonin did not affect the astrocytic cell death following 4 or 8 hr of OGD. The relative MTT values of the neuronal cells were (as mean ± S.E.M.) 59.1 ± 2.4% in the vehicle‐treated OGD group and 80.1 ± 2.7%, 82.5 ± 2.9%, 74.1 ± 2.3%, 64.2 ± 2.3%, 62.7 ± 2.8%, and 61.0 ± 3.9% in the OGD groups treated with melatonin at 10−3, 10−4, 10−5, 10−6, 10−7, and 10−8 m, respectively. Reduction in cell death was significant following treatment with melatonin at 10−3, 10−4, or 10−5 m. Reverse transcription‐polymerase chain reaction showed that human mt1 and MT2 membrane receptors were not expressed in the cultured neuronal cells. Our results show that melatonin co‐treatment protects cultured neuronal cells but not astrocytes against OGD‐induced cell death in a dose‐dependent manner and that the neuroprotection is independent of its known membrane receptors.
Interaction between metabolism and flow in tendon and muscle
Authors: Clark M. G. , Clerk L. H. , Newman J. M. B. , Rattigan S.
Published: March 2003

Blood vessels supplying the musculotendinous region of tendons, including interfibrillar connective tissue, represent a parallel vascular network to that supplying muscle fibers. Reciprocal control by vasomodulators and neural input determines relative flow in each network, and therefore relative supply of nutrients and hormones. Vessels supplying the tendon support fat cells and may function as a non‐nutritive network within muscle groups to regulate resting muscle energy metabolism and to act as a flow reservoir for nutrient delivery in exercise.
Stress fractures of the tibia: can personality traits help us detect the injury‐prone athlete?
Authors: Ekenman I. , Hassmén P. , Koivula N. , Rolf C. , Felländer‐Tsai L.
Published: March 2003

One of the few serious drawbacks associated with running is overuse injuries such as stress fractures of the tibia, which cause local pain and swelling, often resulting in a temporary cessation of training. Whereas some runners rarely become injured, others experience recurrent injuries even during fairly short periods of time. The aim of the present study was to compare selected personality traits in a group of runners who had sustained a previous tibial stress fracture (n=17), with a matched group of runners (n=17) who had never experienced stress fractures. The results indicated that the injured runners, especially the women, scored higher than the non‐injured runners did on inventories measuring both the Type A behavior pattern and exercise dependency. Since motivation, ambitiousness, and competitiveness are integral parts of these inventories, high scoring individuals might be part of a high‐risk population for running injuries, the more so if the individual also feels dependent on regular running for managing stress related mood states, which was the case particularly for the injured women in the present study. However, the somewhat limited number of runners who had had a confirmatory scintigram, which was a criterion for inclusion in the study, warrants a cautious interpretation of the results. The findings nevertheless suggest that in order to prevent recurrent injuries, health education professionals and clinicians ought to focus on conveying the importance of detecting precursors of injury, and the subsequent steps which should be taken to avoid developing a serious injury.
Back pain and radiological changes in the thoraco‐lumbar spine of athletes. A long‐term follow‐up
Authors: Lundin O. , Hellström M. , Nilsson I. , Swärd L.
Published: March 2003

From 1996 to 1999, back pain and radiological changes in the thoraco‐lumbar spine were investigated in 134 former top athletes, representing wrestling, gymnastics, soccer and tennis (age 27–39 years) and a group of 28 non‐athletes of comparable age. This is a long‐term follow‐up investigation of a previous radiological study of the spine with clinical correlation. Despite significantly more radiological abnormalities among the athletes, they did not report higher frequency of back pain than the non‐athletes. A decrease in disc height or new disc height reduction in one or more of the intervertebral discs between the two examinations correlated significantly with back pain at follow‐up.
Criterion related validity of self‐reported knee symptoms among athletes
Authors: Hahn T.
Published: March 2003

The objective of this study was to investigate the validity of self‐reported knee symptoms among athletes. During 12 months, 252 athletes answered a weekly questionnaire about concurrent sports activity, knee pain, knee instability, knee swelling and knee locking. Finally, the athletes answered a retrospective questionnaire summing up symptoms during the preceding 12 months. Accordingly, two 12 month period prevalences of the same period were calculated for each knee symptom. The proportion of agreement between the retrospective 12 month questionnaire and the concurrent weekly questionnaires was for knee pain, knee instability, knee swelling and knee locking, 86%, 89%, 89% and 90%, respectively. Using weekly reports as reference, the corresponding sensitivity of the 12 month period prevalence question was 0.86, 0.71, 0.66 and 0.56, respectively. The sensitivities were all significantly associated with the number of weeks since the last episode of the corresponding knee symptom. As concerns estimation of 12 month period prevalences, a retrospective summary questionnaire at the end of the period is a valid substitute for concurrent weekly questionnaire information on knee pain, knee instability and knee swelling, but only a moderate substitute for information on knee locking. A reduction in the length of the period the athlete should recall knee symptoms might improve the questionnaire validity.
Which factors predict outcome in the treatment program of anterior knee pain?
Authors: Witvrouw E. , Lysens R. , Bellemans J. , Cambier D. , Cools A. , Danneels L. , Bourgois J.
Published: March 2003

The goal of this prospective study was to determine the outcome‐predictive role of various parameters in the nonoperative treatment of chronic anterior knee pain patients. Thirty patients followed a five‐week treatment program, which consisted out of only closed kinetic chain exercises. Prior to this treatment all subjects were evaluated on muscular characteristics, subjective symptoms, weight, sex, duration of symptoms and functional performance. A multiple stepwise regression analysis revealed that the reflex response time of m. vastus medialis obliquus (VMO) (P=0.041; 0.026), and the duration of symptoms (P=0.019; 0.045) were the only two parameters which were significantly associated with the outcome (evaluated by the Kujala score) at five weeks, and at three months. The shorter the duration of symptoms, or the faster the reflex response time of VMO prior to the treatment, the better the outcome after a closed kinetic chain exercise program. The statistical significance of these parameters in this study may be seen as an indication of the importance of these variables as predictors of the outcome of a closed kinetic chain strengthening program. Using this information, it seems clinically important to begin the treatment program before the anterior knee pain becomes more chronic and treatment results become less good.
Arthroscopic treatment of impingement of the ankle reduces pain and enhances function
Authors: Rasmussen S. , Hjorth Jensen C.
Published: March 2003

A consecutive series of 105 patients with a median age of 35 (16–62) years who were operated on with arthroscopic resection for impingement of the ankle using standardized technique without distraction is presented. All patients complained of painful dorsiflexion and had failed to respond to conservative treatment. A total of 177 diagnoses were found, soft tissue impingement or synovitis in 89, anterior bony impingement in 44, chondral lesion in 20, loose bodies in 16 and osteoarthritis in eight. At follow‐up after 2 years, 65 patients were pain free while 28 patients had experienced reduction of pain. Gait was improved in 30/41 patients and 22 resumed sporting activities. The results were graded excellent in 67, good in 25, fair in six and poor in seven patients. There were four deep infections and one synovial fistula in this series. The deep infections all responded well to arthroscopic synovectomy and intravenous antibiotics. In one patient persistent symptoms were recorded. Ankle arthroscopy yielded good results in the treatment of anterior impingement of the ankle as it effectively reduced pain and enhanced function.
Tug of war: introduction to the sport and an epidemiological injury study among elite pullers
Authors: Smith Jay , Krabak Brian
Published: March 2003

The objective of this study was to describe injury patterns among elite tug of war athletes, using a survey method. The setting was the World Tug of War (TOW) Championships in 1998. All 990 male and female athletes asked to participate. Demographic data, participation history, and injury history during TOW, including injury number and type, were collected. A total of 252 (187 males and 65 females) athletes completed the survey (25% response rate). Although males had been competing longer than females (9.3 ± 6.9 years vs. 6.5 ± 5.0 years, t‐test, P < 0.005), the percentage of males (64/187 = 32%) and females (24/65 = 37%) females reporting TOW injuries was similar. Strains and sprains comprised over half of all injuries, and the back (42%), shoulder–upper limb (23%), and knee (17%) were most commonly injured. Injury patterns were similar among males and females. Elite TOW athletes represent a diverse age range (15–55 years) of males and females who have competed for 5–10 years. About one‐third will have suffered past TOW‐related injuries, typically involving sprains or strains of the low back, shoulder girdle–upper extremity, and knee. The possibility of an increased injury rate among female TOW athletes warrants further study. Considerations for future research are discussed.
Alteration in albumin level during modified muscular activity
Authors: Wagatsuma Akira , Fujimoto Koichi , Yamada Shigeru
Published: March 2003

We have previously reported that albumin protein is increased in the atrophied muscle induced by hindlimb immobilization. The purpose of this study was to evaluate the effects of several disuse models on albumin protein and mRNA levels in mice skeletal muscle and to investigate whether the elevated amount of albumin returns to control level by muscular activity increased by hindlimb remobilization. Western blot analysis revealed that hindlimb immobilization, denervation, and tenotomy, except for hindlimb unloading, significantly increased albumin levels in soleus muscles by 2.1‐, 1.9‐ and 2.0‐fold, respectively (P < 0.001). Immunohistochemical analysis showed that albumin protein accumulates in the widened extracellular space. Reverse transcription‐polymerase chain reaction (RT‐PCR) assay revealed albumin gene expression to be downregulated in all disuse models relative to control level. During hindlimb remobilization, the amounts of albumin protein appeared to remain higher level after 3 and 7 days and had returned to control level after 14 days and muscle mass, the amounts of myosin heavy chain, and actin proteins seemed to restore control levels after 21 days. These results indicate that the amount of interstitial albumin protein may be modulated by muscular activity.
Muscle performance, sex hormones and training in peri‐menopausal and post‐menopausal women
Authors: Sipilä S. , Poutamo J.
Published: March 2003

Age‐related deterioration in muscle performance is one of the major reasons for decreased functional capacity and disability in older people. In women, impaired muscle performance has already been observed during peri‐menopause in concert with rapid and dramatic decrease in ovarian hormone production. This observation suggests that female sex steroids may have an important role among other agents in regulating muscle performance in middle‐aged and older women. Previous experimental studies have shown that hormone replacement therapy (HRT) and intensive physical training have positive effects on muscle force and explosive power in healthy post‐menopausal women.

 Hormone replacement therapy in combination with physical training may exert even greater gains in muscle performance than HRT and training alone. Despite the significant mean increase in muscle force and power by HRT and/or training, a considerable variability in the individual responses is observed. The mechanism, by which female sex steroids act on muscle performance, is still unclear. Therefore, more research is needed in order to explore all the pathways, by which these steroids could act on skeletal muscle in peri‐menopausal and post‐menopausal women.
Potential of a high‐frequency correlation method to study skin blood flow
Authors: Gens Fabrice , Remenieras Jean‐Pierre , Patat Frédéric , Diridollou Stéphane , Berson Marceau
Published: March 2003

Background/aims: The vast number of existing dedicated techniques proves that skin blood flow estimation is an unsolved problem. Specificities of cutaneous vascularization (very low blood velocity, noisy environment, complexity of the vascularization architecture) result in the unsuitability of conventional ultrasonic Doppler techniques (long acquisition time, low spatial resolution). The object here was to present a high‐frequency time‐domain correlation method. In particular, the difficulties of adaptating this type of measurement (data processing, hardware problem) are pointed out.

Methods: Radio‐frequency (RF) backscattered signals, obtained with a modified version of a home‐made 20 MHz skin imaging system, are studied. Time shifts between successive windowed sections of the RF signals are determined by the mean of the cross‐correlation algorithm. A realignment procedure (to remove the artefacts caused by the movements of the patient and the manipulator) and a stationary echo cancelling procedure (to remove the signals coming from the cutaneous tissues and to permit the detection of very small vessels) are used.

Results: In vitro results show that velocity measurements as low as 0.1 mm/s are attainable with a 80 μm axial resolution, and blood vessels of 100 μm are detectable. Our technique has also been validated by means of in vivo experiment on an erysipelas located on a human leg. In this way, a 180‐μm‐diameter blood vessel has been detected on a M‐mode RF image and the corresponding velocity profile has been obtained.

Conclusion: Further improvements can be expected, and the level of performance obtained in vitro in this work should be also attainable in vivo and should then provide an effective tool for skin physiology and pathology.
Study of three complementary techniques for measuring cutaneous hydration in vivo in human subjects: NMR spectroscopy, transient thermal transfer and corneometry – application to xerotic skin and cosmetics
Authors: Girard Philippe , Beraud Anne , Sirvent Anne
Published: March 2003

Background/aims: The aim of this study was to determine the capability and the analytical quality of three different in vivo, non‐invasive, quantitative methods for measuring skin hydration: two innovative methods that have been used for more than eight years – nuclear magnetic resonance spectroscopy (NMR‐S) and transient thermal transfer (TTT) – and the more widely used and conventional corneometry.

Methods: The work presented evaluated the capability and precision, as well as cutaneous exploration depths, of the three methods. Experiments were carried out in vivo following the hydration, in kinetic terms, induced by topic application of reference moisturizing products. Spatio‐temporal efficacy of a lipolotion was also studied by the TTT method. Cases of xerotic skin were studied with TTT and corneometry.

Results: The results obtained showed better repeatability and reproducibility with the TTT and NMR‐S methods than with corneometry. NMR‐S is one of the only direct hydration measurement methods. It measures skin hydration down to the outer dermis with high precision. It is indicated for products having an action down to the deep cutaneous layers. By changing thermal power parameters, the TTT method can determine hydration to the outer, middle or deep epidermal layers. It is, therefore, possible to track the penetration of products in various layers of the epidermis. The small size of the probe enables the hydration measurement of skin sites (lips, eyelids) that were not, up to now, measurable with the two other methods. Corneometric investigations are restricted to the surface of the horny layer; measurements are easy and rapid but influenced by the composition of products applied to the skin and their phases: aqueous, oily or ionic. The xerotic skin study highlights the importance of exploration in different layers of the epidermis, as dehydration concerns not only the upper layers of the epidermis but also the medial and deep layers. With the TTT method, it has been possible to highlight the penetration dynamics of a lipolotion with, initially, an increase in the hydration in the outer epidermis, followed 3 h later by a transfer from the outer to the middle epidermis.

Conclusion: NMR‐S, TTT and corneometry represent three possible ways to assess skin hydration. Because they explore different cutaneous depths, they are more complementary than competitive. Transient thermal transfer, although a semi‐direct method, is a precise, informative, and innovative solution to evaluate skin hydration at different epidermal depths and sites.
Quantitative assessment of properties of make‐up products by video imaging: application to lipsticks
Authors: Korichi Rodolphe , Provost Robin , Heusèle Catherine , Schnebert Sylvianne
Published: March 2003

Background/aims: The different properties and visual effects of lipstick have been studied by image analysis directly on volunteers.

Methods: After controlling the volunteer's position mechanically using an ophthalmic table and visually using an acquirement mask, which is an indicator of luminance and guide marks, we carried out video colour images of the make‐up area. From these images, we quantified the colour, gloss, covering power, long‐lasting effect and streakiness, using computer science programs.

Results/Conclusion:

Quantitative colorimetric assessment requires the transformation of the RGB components obtained by a video colour camera into CIELAB colorimetric space. The expression of each coordinate of the L*a*b* space according to R,G,B was carried out by a statistical method of polynomial approximations. A study, using 24 colour images extracted from a Pantone® palette, showed a very good correlation with a Minolta Colorimeter® CR 300. The colour assessment on volunteers required a segmentation method by maximizing the entropy. The aim was to separate the colour information sent back by the skin to the make‐up area. It was very useful to precisely delimit the contour between the skin and the product in the case of almost identical colours and to evaluate the streakiness. From this colour segmentation, an algorithm was studied to search for the shades most represented in the overall colour of the make‐up area. The capacity to replicate what the consumer perceives of the make‐up product, to carry out studies without having any contact with the skin surface, and the constant improvement of software and video acquirement systems all make video imaging a very useful tool in the quantitative assessment of the properties and visual effects of a make‐up product.
The use of high‐frequency diagnostic ultrasound to investigate the effect of hormone replacement therapy on skin thickness
Authors: Chen L. , Dyson M. , Rymer J. , Bolton P. A. , Young S. R.
Published: March 2003

Background/aims: Previous investigations have suggested that hormone replacement therapy (HRT) could have a positive effect on the maintenance of skin thickness post‐menopause. Previous skin measurement devices have proved variable in their accuracy and ease of use. This investigation assessed the effect of HRT on the skin in a noninvasive way, using high‐frequency diagnostic ultrasound.

Method: The study was a cross‐sectional observational study, carried out at a menopause and gynaecology outpatient’s clinic. A total of 84 women (comprising 34 HRT users, 25 post‐menopausal controls, and 25 premenopausal controls) took part in the study. Each volunteer was scanned using diagnostic ultrasound on the arm. Skin thickness measurements were made from each scan using computerised image analysis.

Results: Skin thickness was shown to be greater in the HRT group than in the post‐menopausal controls (P<0.01).

Conclusions: High‐frequency diagnostic ultrasound proved to be a useful clinical tool and showed that HRT appears to help maintain skin thickness in menopause.
MHC Class II Presentation of Endogenous Tumor Antigen by Cellular Vaccines Depends on the Endocytic Pathway but not H2‐M
Authors: Qi Ling , Ostrand‐Rosenberg Suzanne
Published: March 2003

We have developed cell‐based cancer vaccines that activate anti‐tumor immunity by directly presenting endogenously synthesized tumor antigens to CD4+ T helper lymphocytes via MHC class II molecules. The vaccines are non‐conventional antigen‐presenting cells because they express MHC class II, do not express invariant chain or H‐2M, and preferentially present endogenous antigen. To further improve therapeutic efficacy we have studied the intracellular trafficking pathway of MHC class II molecules in the vaccines using endoplasmic reticulum‐localized lysozyme as a model antigen. Experiments using endocytic and cytosolic pathway inhibitors (chloroquine, primaquine, and brefeldin A) and protease inhibitors (lactacystin, LLnL, E64, and leupeptin) indicate antigen presentation depends on the endocytic pathway, although antigen degradation is not mediated by endosomal or proteasomal proteases. Because H2‐M facilitates presentation of exogenous antigen via the endocytic pathway, we investigated whether transfection of vaccine cells with H‐2M could potentiate endogenous antigen presentation. In contrast to its role in conventional antigen presentation, H‐2M had no effect on endogenous antigen presentation by vaccine cells or on vaccine efficacy. These results suggest that antigen/MHC class II complexes in the vaccines may follow a novel route for processing and presentation and may produce a repertoire of class II‐restricted peptides different from those presented by professional APC. The therapeutic efficacy of the vaccines, therefore, may reside in their ability to present novel tumor peptides, consequently activating tumor‐specific CD4+ T cells that would not otherwise be activated.
Control of MHC Class I Traffic from the Endoplasmic Reticulum by Cellular Chaperones and Viral Anti‐Chaperones
Authors: Gruhler Albrecht , Früh Klaus
Published: March 2003

MHC class I molecules assemble with peptides in the endoplasmic reticulum (ER). To ensure that only peptide‐loaded MHC molecules leave the ER, empty molecules are retained by ER‐resident chaperones, most notably the MHC‐specific tapasin. ER exit of class I MHC is also controlled by viruses, but for the opposite purpose of preventing peptide presentation to T cells. Interestingly, some viral proteins are able to retain MHC class I molecules in the ER despite being transported. By contrast, other viral proteins exit the ER only upon binding to class I MHC, thereby rerouting newly synthesized class I molecules to intracellular sites of proteolysis. Thus, immune escape can be achieved by reversing, inhibiting or redirecting the chaperone‐assisted MHC class I folding, assembly and intracellular transport.
ADPKD: A Human Disease Altering Golgi Function and Basolateral Exocytosis in Renal Epithelia
Authors: Charron Audra J. , Bacallao Robert L. , Wandinger‐Ness Angela
Published: March 2003

Epithelial cells explanted from autosomal dominant polycystic kidney disease (ADPKD) tissue exhibit impaired exocytosis, specifically between the Golgi and basolateral membrane (Charron A, Nakamura B, Bacallo R, Wandinger‐Ness A. Compromised cytoarchitecture and polarized trafficking in autosomal dominant polycystic kidney disease cells. J Cell Biol 2000; 148: 111–124.). Here the defect is shown to result in the accumulation of the basolateral transport marker vesicular stomatitis virus (VSV) G protein in the Golgi complex. Golgi complex morphology is consequently altered in the disease cells, evident in the noticeable fenestration and dilation of the cisternae. Further detailed microscopic evaluation of normal kidney and ADPKD cells revealed that ineffective basolateral exocytosis correlated with modulations in the localization of select post‐Golgi transport effectors. The cytosolic coat proteins p200/myosin II and caveolin exhibited enhanced association with the cytoskeleton or the Golgi of the disease cells, respectively. Most cytoskeletal components with known roles in vesicle translocation or formation were normally arrayed with the exception of Golgi β‐spectrin, which was less prevalent on vesicles. The rab8 GTPase, important for basolateral vesicle targeting, was redistributed from the perinuclear Golgi region to disperse vesicles in ADPKD cells. At the basolateral membrane of ADPKD cells, there was a notable loss of the exocyst components sec6/sec8 and an unidentified syntaxin. It is postulated that dysregulated basolateral transport effector function precipitates the disruption of basolateral exocytosis and dilation of the ADPKD cell Golgi as basolateral cargo accumulates within the cisternae.
Watching Proteins in the Wild: Fluorescence Methods to Study Protein Dynamics in Living Cells
Authors: Chamberlain Chester , Hahn Klaus M.
Published: March 2003

The advent of GFP imaging has led to a revolution in the study of live cell protein dynamics. Ease of access to fluorescently tagged proteins has led to their widespread application and demonstrated the power of studying protein dynamics in living cells. This has spurred development of next generation approaches enabling not only the visualization of protein movements, but correlation of a protein's dynamics with its changing structural state or ligand binding. Such methods make use of fluorescence resonance energy transfer and dyes that report changes in their environment, and take advantage of new chemistries for site‐specific protein labeling.
Dsl1p, an Essential Protein Required for Membrane Traffic at the Endoplasmic Reticulum/Golgi Interface in Yeast
Authors: VanRheenen Susan M. , Reilly Barbara A. , Chamberlain Stormy J. , Waters M. Gerard
Published: March 2003

To identify novel factors required for ER to Golgi transport in yeast we performed a screen for genes that, when mutated, confer a dependence on a dominant mutant form of the ER to Golgi vesicle docking factor Sly1p, termed Sly1‐20p. DSL1, a novel gene isolated in the screen, encodes an essential protein with a predicted molecular mass of 88 kDa. DSL1 is required for transport between the ER and the Golgi because strains bearing mutant alleles of this gene accumulate the pre‐Golgi form of transported proteins at the restrictive temperature. Two strains bearing temperature‐sensitive alleles of DSL1 display distinct phenotypes as observed by electron microscopy at the restrictive temperature; although both strains accumulate ER membrane, one also accumulates vesicles. Interestingly, the inviability of strains bearing several mutant alleles of DSL1 can be suppressed by expression of either Erv14p (a protein required for the movement of a specific protein from the ER to the Golgi), Sec21p (the γ‐subunit of the COPI coat protein complex coatomer), or Sly1‐20p. Because the strongest suppressor is SEC21, we proposed that Dsl1p functions primarily in retrograde Golgi to ER traffic, although it is possible that Dsl1p functions in anterograde traffic as well, perhaps at the docking stage, as suggested by the suppression by SLY1‐20.
Type IVB Secretion by Intracellular Pathogens
Authors: Sexton Jessica A. , Vogel Joseph P.
Published: March 2003

A growing number of pathogens are being found to possess specialized secretion systems which they use in various ways to subvert host defenses. One class, called type IV, are defined as having homology to the conjugal transfer systems of naturally occurring plasmids. It has been proposed that pathogens with type IV secretion systems have acquired and adapted the conjugal transfer systems of plasmids and now use them to export toxins. Several well‐characterized intracellular pathogens, including Legionella pneumophila, Coxiella burnetii, Brucella abortus, and Rickettsia prowazekii, contain type IV systems which are known or suspected to be of critical importance in their ability to cause disease. Specifically, these systems are believed to be the key factors determining intracellular fate, and thus the ability to replicate and cause disease.
Low pH Inhibits Compensatory Endocytosis at a Step Between Depolarization and Calcium Influx
Authors: Smith Robert M. , Baibakov Boris , Lambert Nevin A. , Vogel Steven S.
Published: March 2003

Cell function can be modulated by the insertion and removal of ion channels from the cell surface. The mechanism used to keep channels quiescent prior to delivery to the cell surface is not known. In eggs, cortical vesicle exocytosis inserts voltage‐gated calcium channels into the cell surface. Calcium influx through these channels triggers compensatory endocytosis. Secretory vesicles contain high concentrations of calcium and hydrogen ions. We propose that lumenal hydrogen ions inhibit vesicular calcium channel gating prior to exocytosis, discharge of lumenal protons upon vesicle‐plasma membrane fusion enables calcium channel gating. Consistent with this hypothesis we find that cortical vesicle lumens are acidic, and exocytosis releases lumenal hydrogen ions. Acidic extracellular pH reversibly blocks endocytosis, and the windows of opportunity for inhibition with a calcium‐channel blocker or hydrogen ions are indistinguishable. Calcium ionophore treatment circumvents the low pH block, suggesting that calcium influx, or an upstream step, is obstructed. Inhibition of calcium influx by preventing membrane depolarization is unlikely, as elevation of the extracellular potassium concentration failed to overcome the pH block, and low extracellular pH was found to depolarize the membrane potential. We conclude that low pH inhibits endocytosis at a step between membrane depolarization and calcium influx.
SifA, a Type III Secreted Effector of Salmonella typhimurium, Directs Salmonella‐Induced Filament (Sif) Formation Along Microtubules
Authors: Brumell John H. , Goosney Danika L. , Finlay B. Brett
Published: March 2003

A unique feature of Salmonella enterica serovar typhimurium (S. typhimurium) is its ability to enter into (invade) epithelial cells and elongate the vacuole it occupies into tubular structures called Salmonella‐induced filaments (Sifs). This phenotype is dependent on SifA, a Salmonella virulence factor that requires the SPI‐2‐encoded type III secretion system for delivery into host cells. Previous attempts to study SifA and other type III secreted proteins have been limited by a lack of suitable reagents. We examined SifA function by expressing SifA with two internal hemagglutinin epitope tags. By employing subcellular fractionation techniques, we determined that translocated SifA was membrane associated in infected HeLa cells. Confocal microscopy revealed that SifA associated with the Salmonella vacuole and with Sifs. Our analysis also revealed that microtubules serve as a scaffold for Sifs, and that SifA colocalizes with microtubules at sites of interaction between lysosomal glycoprotein‐containing vesicles and Sifs. Treatment with the microtubule inhibitor nocodazole blocked Sif formation but did not prevent SifA translocation into the Salmonella vacuole. While polymerized actin has been observed on Sifs, this phenotype was transient and did not play a role in promoting or maintaining Sif formation. Our findings demonstrate the essential role of microtubule dynamics in the formation of Sifs and the utility of this epitope tagging strategy for the study of bacterial type III secreted proteins.
Clathrin‐Dependent or Not: Is It Still the Question?
Authors: Johannes Ludger , Lamaze Christophe
Published: March 2003

Whether the endocytic uptake of a given molecule is mediated through clathrin‐coated pits or not is a classical criterion used to characterize its endocytic pathway(s). Hence, clathrin‐dependent endocytosis has been associated with highly selective and efficient uptake, whereas clathrin‐independent endocytosis appeared to be confined to bulk uptake of fluid‐phase markers. This scholastic view has recently been challenged using newly developed molecular tools that allow for the first time a functional and mechanistic analysis of these less well‐characterized clathrin‐independent pathways, including caveolar uptake and macropinocytosis. Furthermore, several studies point to a critical role of lateral lipid asymmetry – lipid rafts/microdomains – in membrane sorting. We will discuss the potential role of these structures in endocytosis and the possibility that differential sorting at the plasma membrane predisposes the ensuing intracellular fate of a given molecule as well as its physiological function.
An ER Membrane Protein, Sop4, Facilitates ER Export of the Yeast Plasma Membrane [H+]ATPase, Pma1
Authors: Luo Wen‐jie , Gong Xiao‐hua , Chang Amy
Published: March 2003

We have analyzed the mechanism by which Sop4, a novel ER membrane protein, regulates quality control and intracellular transport of Pma1–7, a mutant plasma membrane ATPase. At the restrictive temperature, newly synthesized Pma1–7 is targeted for vacuolar degradation instead of being correctly delivered to the cell surface. Loss of Sop4 at least partially corrects vacuolar mislocalization, allowing Pma1–7 routing to the plasma membrane. Ste2–3 is a mutant pheromone receptor which, like Pma1–7, is defective in targeting to the cell surface, resulting in a mating defect. sop4Δ suppresses the mating defect of ste2–3 cells as well as the growth defect of pma1–7. Visualization of newly synthesized Pma1–7 in sop4Δ cells by indirect immunofluorescence reveals delayed export from the ER. Similarly, ER export of wild‐type Pma1 is delayed in the absence of Sop4 although intracellular transport of Gas1 and CPY is unaffected. These observations suggest a model in which a selective increase in ER residence time for Pma1–7 may allow it to achieve a more favorable conformation for subsequent delivery to the plasma membrane. In support of this model, newly synthesized Pma1–7 is also routed to the plasma membrane upon release from a general block of ER‐to‐Golgi transport in sec13–1 cells.
The Overexpression of GMAP‐210 Blocks Anterograde and Retrograde Transport Between the ER and the Golgi Apparatus
Authors: Pernet‐Gallay Karin , Antony Claude , Johannes Ludger , Bornens Michel , Goud Bruno , Rios Rosa M.
Published: March 2003

Golgi Microtubule‐Associated Protein (GMAP)‐210 is a peripheral coiled‐coil protein associated with the cis‐Golgi network that interacts with microtubule minus ends. GMAP‐210 overexpression has previously been shown to perturb the microtubule network and to induce a dramatic enlargement and fragmentation of the Golgi apparatus (Infante C, Ramos‐Morales F, Fedriani C, Bornens M, Rios RM. J Cell Biol 1999; 145: 83–98). We now report that overexpressing GMAP‐210 blocks the anterograde transport of both a soluble form of alkaline phosphatase and the hemagglutinin protein of influenza virus, an integral membrane protein, between the endoplasmic reticulum and the cis/medial (mannosidase II‐positive) Golgi compartment. Retrograde transport of the Shiga toxin B‐subunit is also blocked between the Golgi apparatus and the endoplasmic reticulum. As a consequence, the B‐subunit accumulates in compartments positive for GMAP‐210. Ultrastructural analysis revealed that, under these conditions, the Golgi complex is totally disassembled and Golgi proteins as well as proteins of the intermediate compartment are found in vesicle clusters distributed throughout the cell. The role of GMAP‐210 on membrane processes at the interface between the endoplasmic reticulum and the Golgi apparatus is discussed in the light of the property of this protein to bind CGN membranes and microtubules.
Fusion and Fission Events in the Endocytic Pathway of Dictyostelium
Authors: Maniak Markus
Published: March 2003

The endocytic pathway in Dictyostelium appears as a short circuit between endocytosis and exocytosis. Within the hour that elapses between internalization of nutrients and release of remnants, digestion by lysosomal enzymes occurs. Meanwhile, the maturing endosome undergoes a complex series of fusion and fission events, which change its character profoundly and which are far from being fully understood. This review attempts to order the dynamic events into a sequence of stages that is most consistent with present knowledge.
Management of HIV and AIDS in the African context
Authors: Wood R
Published: March 2003

The initial response to the African HIV epidemic was to concentrate on the prevention of new infections. There is now an urgent need to address the health care requirements of large numbers of already infected individuals. The spectrum of disease in the African setting is dominated by tuberculosis, bacterial and protozoan infections. In much of Africa, health services are overwhelmed by the care of terminally ill AIDS patients. In the absence of specific HIV therapy, health care resources are being increasingly utilised, but with little survival benefit for the individual. Resources available for treating patients vary considerably between the richer and poorer countries of the continent. Primary prevention of opportunistic infections and maternal child transmission are at present affordable and cost‐effective interventions. Whilst antiretroviral therapies may presently be unaffordable in much of Africa, they represent a modality that can have a major effect on HIV survival. The challenge is to improve the health and longevity of HIV‐infected individuals with the rational use of the limited health resources available in Africa today.
Recurrent aphthous stomatitis and gene polymorphisms for the inflammatory markers TNF‐α, TNF‐β and the vitamin D receptor: no association detected
Authors: Bazrafshani MR , Hajeer AH , Ollier WER , Thornhill MH
Published: March 2003

OBJECTIVE: To investigate the possibility that minor recurrent aphthous stomatitis (MiRAS) is associated with the inheritance of specific gene polymorphisms for markers associated with macrophage driven inflammation, i.e. tumor necrosis factor‐α (TNF‐α), TNF‐β or the vitamin D receptor (VDR).

SETTING: MiRAS is a common, painful, ulcerative condition of the mouth. Its etiology is unknown although mononuclear inflammatory cells are thought to play an important role. There is no clear genetic mode of inheritance, however, many patients report a positive family history and disease concordance is significantly higher in monozygotic than dizygotic twins, suggesting a polygenic mode of inheritance.

METHODS: Ninety‐five MiRAS patients and an ethnically matched control population were genotyped for TNFA‐308, TNFB NcoI and VDR (intron 8 and exon 9) polymorphisms. Chi‐square analysis was used to compare the allele frequencies and genotypes of cases and controls.

RESULTS: No significant association was identified between inheritance of specific alleles or genotypes of the TNFA‐308, TNFB NcoI and VDR (intron 8 and exon 9) polymorphisms and susceptibility to MiRAS.

CONCLUSIONS: Inheritance of specific gene polymorphisms for TNF‐α, TNF‐β or VDR does not appear to be a significant factor in determining susceptibility to MiRAS.
RNA chaperone activity of the Sm‐like Hfq protein
Authors: Moll Isabella , Leitsch David , Steinhauser Tanja , Bläsi Udo
Published: March 2003

The Escherichia coli Sm‐like host factor I (Hfq) protein is thought to function in post‐transcriptional regulation by modulating the function of small regulatory RNAs. Hfq also interferes with ribosome binding on E. coli ompA messenger RNA, indicating that Hfq also interacts with mRNAs. In this study, we have used stimulation of group I intron splicing in vivo and a modified in vitro toeprinting assay to determine whether Hfq acts as an RNA chaperone. Hfq was able to rescue an RNA ‘folding trap’ in a splicing defective T4 bacteriophage td gene in vivo. Enzymatic analysis showed that Hfq affects the accessibility of the ompA start codon, as well as other bases within the ribosome‐binding site, explaining its negative effect on ribosome binding. We also show that the Hfq‐induced structural changes in ompA mRNA are maintained after proteolytic digestion of the protein, which classifies Hfq as an RNA chaperone.
Cross-talk between glucocorticoid and retinoic acid signals involving glucocorticoid receptor interaction with the homoeodomain protein Pbx1
Authors: SUBRAMANIAM Nanthakumar , CAMPIÓN Javier , RAFTER Ingalill , OKRET Sam
Published: March 2003

Glucocorticoid (GC) signalling influences the response of the cell to a number of other signals via a mechanism referred to as ‘cross-talk'. This cross-talk may act at several levels, including an interaction between the transcription factors involved in the signalling pathways. In the present paper, we demonstrate a novel functional interaction between GC and all-trans-retinoic acid (RA) signalling. We show that, in P19 embryonal carcinoma cells, GCs potentiate RA-induced expression of the murine Hoxb-1 gene through an autoregulatory element, b1-ARE, recognized by the Pbx1 and HOXB1 homoeodomain proteins. The synergistic effect of GC did not involve GC receptor (GR) binding to the b1-ARE, and the GC—GR complex alone was unable to activate transcription via the element. Furthermore, the ability of the GR to transactivate was not required, excluding expression of a GC-induced protein as the mechanism for the GC/RA synergy. Additional transfection experiments showed that the Pbx1/HOXB1 heterodimer was the target for the GC effect. Furthermore, functional dissection of the GR demonstrated that the DNA-binding domain (DBD) of the GR was required for the synergy. A physical interaction between the GR and Pbx1 proteins was demonstrated in vivo by co-immunoprecipitation experiments. These results are compatible with a model in which the GC/RA synergy is mediated by a direct interaction between the GR and Pbx1. On the basis of the ubiquitous expression of both GR and Pbx1, a number of genes regulated by Pbx are likely to be important targets for GC-mediated ‘cross-talk'.
Nucleoside diphosphate kinase of Saccharomyces cerevisiae, Ynk1p: localization to the mitochondrial intermembrane space
Authors: AMUTHA Boominathan , PAIN Debkumar
Published: March 2003

Nucleoside diphosphate kinase (NDPK) is a highly conserved multifunctional enzyme. It catalyses the transfer of γ phosphates from nucleoside triphosphates to nucleoside diphosphates by a mechanism that involves formation of an autophosphorylated enzyme intermediate. The phosphate is usually supplied by ATP. NDPK activity in different subcellular compartments may regulate the crucial balance between ATP and GTP or other nucleoside triphosphates. NDPKs are homo-oligomeric proteins and are predominantly localized in the cytosol. In this paper, we demonstrate that in Saccharomyces cerevisiae a small fraction of total NDPK activity encoded by YNK1 is present in the intermembrane space (IMS) of mitochondria, and the corresponding protein Ynk1p in the IMS represents approx. 0.005% of total mitochondrial proteins. Ynk1p, synthesized as a single gene product, must therefore be partitioned between cytoplasm and mitochondrial IMS fractions. A mechanism for this partitioning is suggested by our observations that interaction with a 40kDa protein of the translocase of outer mitochondrial membrane (Tom40p), occurs preferentially with unfolded, unphosphorylated forms of Ynk1p. A population of newly translated, but not yet folded or autophosphorylated, Ynk1p intermediates may be imported into the IMS of mitochondria and trapped there by subsequent folding and oligomerization. Within the small volume of the IMS, Ynk1p may be more concentrated and may be required to supply GTP to several important proteins in this compartment.
Responses of carboxylating enzymes, sucrose metabolizing enzymes and plant hormones in a tropical epiphytic CAM orchid to CO2 enrichment
Authors: Li C. R. , Gan L. J. , Xia K. , Zhou X. , Hew C. S.
Published: March 2003

After exposure to a doubled CO2 concentration of 750 µL L−1 for 2 months, average relative growth rate (RGR) of Mokara Yellow increased 25%. The two carboxylating enzymes, ribulose‐1,5‐bisphosphate carboxylase/oxygenase (Rubisco) and phosphoenolpyruvate carboxylase (PEPCase), responded differently to CO2 enrichment. There was a significant daytime down‐regulation in Rubisco activity in the leaves of CO2‐enriched plants. However, PEPCase activity in CO2‐enriched plants was much higher in the dark period, although it was slightly lower during the daytime than that at ambient CO2. Leaf sucrose–phosphate synthase (SPS) and sucrose synthase (SS) activities in CO2‐enriched plants increased markedly, along with a night‐time increase in total titratable acidity and malate accumulation. There was a remarkable increase in the levels of indole‐3‐acetic acid (IAA), gibberellins A1 and A3 (GA1+3), isopentenyladenosine (iPA) and zeatin riboside (ZR) in the expanding leaves of plants grown at elevated CO2. It is suggested that (1) the down‐regulation of Rubisco and up‐regulation of SPS and SS are two important acclimation processes that are beneficial because it enhanced both photosynthetic capacity at high CO2 and reduced resource investment in excessive Rubisco capacity; (2) the increased levels of plant hormones in CO2‐enriched M. Yellow might play an important role in controlling its growth and development.
Field Vole Microtus agrestis abundance and Hen Harrier Circus cyaneus diet and breeding in Scotland
Authors: Redpath Stephen M. , Thirgood Simon J. , Clarke Roger
Published: March 2003

In many parts of the global range, voles form an important part of the diet of Hen Harriers Circus cyaneus, and breeding numbers are correlated with the abundance of these small mammals. In Scotland, however, little information is available on harrier diet in the spring and our understanding of causes of variation in harrier breeding density is complicated by human interference. In this paper we explore the relationship between Field Vole Microtus agrestis abundance and harrier spring diet, density and productivity in southern Scotland. Over three years, voles occurred on average in 67% of pellets, and 79% in years of high and intermediate vole abundance. From 1992, the number of breeding harriers increased following protection from illegal persecution. After accounting for this trend, harrier numbers correlated strongly with vole abundance. Harrier clutch size was also correlated with vole abundance. Although fledging success tended to be greater in years of vole increase than in years of vole decline, fledging success was not significantly correlated with the relative abundance of voles, or with the abundance of Meadow Pipits or Red Grouse chicks.