bluuebunny/crossref_metadata_2025 huggingface dataset:
Solar Lentigines are Strongly Related to Sun Exposure in Contrast to Ephelides
Authors: Bastiaens Maarten , Hoefnagel Juliette , Westendorp Rudi , Vermeer Bert‐Jan , Bouwes Bavinck Jan Nico
Published: May 2004

Solar lentigines and ephelides are different types of pigmented skin lesions predominantly present on sun‐exposed skin. Both lesions are risk indicators for melanoma and non‐melanoma skin cancer. Solar lentigines are considered as a sign of photodamage although well‐conducted epidemiological studies are lacking on this subject. Ephelides are associated with fair skin type and red hair. The aim of the present study was to investigate the relation of sun‐exposure estimates with solar lentigines and ephelides. In the Leiden Skin cancer Study 577 patients with malignant melanoma and/or non‐melanoma skin cancer and 385 individuals without a history of skin cancer were studied. The presence of solar lentigines and ephelides in the face and on the back was assessed. Data on skin type, hair color, sun‐exposure variables and cutaneous signs of photodamage were collected, by questionnaire and physical examination. Data were analyzed by chi‐square or Student t‐tests and with multivariable regression. Exposure odds ratios with 95% confidence intervals (95% CI) were calculated to estimate the relative risk for the presence of solar lentigines and ephelides dependent on signs of photodamage. The association with age was strongly positive for solar lentigines whereas it was strongly negative for ephelides (P‐values for trend <0.0001). After adjustment for age, sex and skin type, solar lentigines on the back were positively associated with cumulative (P = 0.01) and intermittent (P = 0.0002) sun exposure. After adjustment, solar lentigines on the back were also associated with a history of sunburns before the age of 20 yr (P = 0.0003) and the number of sunburns in childhood (P = 0.002). Solar lentigines in the face were significantly associated with cutaneous signs of photodamage, i.e. elastosis (odds ratio 2.4, 95% CI 1.7–3.3) and actinic keratosis (odds ratio 1.8, 95% CI 1.3–2.4) whereas ephelides were not. Ephelides in the face and on the back showed an inverse association with chronic sun exposure but after adjustment theses associations disappeared. Sunburns before the age of 20 appeared to be positively associated with ephelides on the back (P = 0.04). In contrast to lentigines, ephelides were much more associated with constitutional host factors such as fair skin and/or red hair (both P < 0.0001). This study indicates that both chronic and acute sun exposure are important in the pathogenesis of solar lentigines.
Granulocyte‐Macrophage Colony‐Stimulating Factor (GM‐CSF) Controls the Proliferation and Differentiation of Mouse Epidermal Melanocytes from Pigmented Spots Induced by Ultraviolet Radiation B
Authors: Hirobe Tomohisa , Furuya Rikako , Hara Eijiro , Horii Izumi , Tsunenaga Makoto , Ifuku Ohji
Published: May 2004

Repeated exposure of ultraviolet radiation B (UVB) on the dorsal skin of hairless mice induces the development of pigmented spots long after its cessation. The proliferation and differentiation of epidermal melanocytes in UVB‐induced pigmented spots are greatly increased, and those effects are regulated by keratinocytes rather than by melanocytes. However, it remains to be resolved what factor(s) derived from keratinocytes are involved in regulating the proliferation and differentiation of epidermal melanocytes. In this study, primary melanoblasts (c. 80%) and melanocytes (c. 20%) derived from epidermal cell suspensions of mouse skin were cultured in a basic fibroblast growth factor‐free medium supplemented with granulocyte‐macrophage colony‐stimulating factor (GM‐CSF). GM‐CSF induced the proliferation and differentiation of melanocytes in those keratinocyte‐depleted cultures. Moreover, an antibody to GM‐CSF inhibited the proliferation of melanoblasts and melanocytes from epidermal cell suspensions derived from the pigmented spots of UV‐irradiated mice, but not from control mice. Further, the GM‐CSF antibody inhibited the proliferation and differentiation of melanocytes co‐cultured with keratinocytes derived from UV‐irradiated mice, but not from control mice. The quantity of GM‐CSF secreted from keratinocytes derived from the pigmented spots of UV‐irradiated mice was much greater than that secreted from keratinocytes derived from control mice. Moreover, immunohistochemistry revealed the expression of GM‐CSF in keratinocytes derived from the pigmented spots of skin in UV‐irradiated mice, but not from normal skin in control mice. These results suggest that GM‐CSF is one of the keratinocyte‐derived factors involved in regulating the proliferation and differentiation of mouse epidermal melanocytes from UVB‐induced pigmented spots.
The Expression and Activation of Protease‐Activated Receptor‐2 Correlate with Skin Color
Authors: Babiarz‐Magee Laura , Chen Nannan , Seiberg Miri , Lin Connie B.
Published: May 2004

Skin color results from the production and distribution of melanin in the epidermis. The protease‐activated receptor‐2 (PAR‐2), expressed on keratinocytes but not on melanocytes, is involved in melanosome uptake via phagocytosis, and modulation of PAR‐2 activation affects skin color. The pattern of melanosome distribution within the epidermis is skin color‐dependent. In vitro, this distribution pattern is regulated by the ethnic origin of the keratinocytes, not the melanocytes. Therefore, we hypothesized that PAR‐2 may play a role in the modulation of pigmentation in a skin type‐dependent manner. We examined the expression of PAR‐2 and its activator, trypsin, in human skins with different pigmentary levels. Here we show that PAR‐2 and trypsin are expressed in higher levels, and are differentially localized in highly pigmented, relative to lightly pigmented skins. Moreover, highly pigmented skins exhibit an increase in PAR‐2‐specific protease cleavage ability. Microsphere phagocytosis was more efficient in keratinocytes from highly pigmented skins, and PAR‐2 induced phagocytosis resulted in more efficient microsphere ingestion and more compacted microsphere organization in dark skin‐derived keratinocytes. These results demonstrate that PAR‐2 expression and activity correlate with skin color, suggesting the involvement of PAR‐2 in ethnic skin color phenotypes.
Unusual Leucophore‐Like Cells Specifically Appear in the Lineage of Melanophores in the Periodic Albino Mutant of Xenopus laevis
Authors: Fukuzawa Toshihiko
Published: May 2004

In the periodic albino mutant (ap/ap) of Xenopus laevis, peculiar leucophore‐like cells appear in the skins of tadpoles and froglets, whereas no such cells are observed in the wild‐type (+/+). These leucophore‐like cells are unusual in (1) appearing white, but not iridescent, under incident light, (2) emitting green fluorescence under blue light, (3) exhibiting pigment dispersion in the presence of α‐melanocyte stimulating hormone (αMSH), and (4) containing an abundance of bizarre‐shaped, reflecting platelet‐like organelles. In this study, the developmental and ultrastructural characteristics of these leucophore‐like cells were compared with melanophores, iridophores and xanthophores, utilizing fluorescence stereomicroscopy, and light and electron microscopy. Staining with methylene blue, exposure to αMSH, and culture of neural crest cells were also performed to clarify the pigment cell type. The results obtained clearly indicate that: (1) the leucophore‐like cells in the mutant are different from melanophores, iridophores and xanthophores, (2) the leucophore‐like cells are essentially similar to melanophores of the wild‐type with respect to their localization in the skin and manner of response to αMSH, (3) the leucophore‐like cells contain many premelanosomes that are observed in developing melanophores, and (4) mosaic pigment cells containing both melanosomes specific to mutant melanophores and peculiar reflecting platelet‐like organelles are observed in the mutant tadpoles. These findings strongly suggest that the leucophore‐like cells in the periodic albino mutant are derived from the melanophore lineage, which provides some insight into the origin of brightly colored pigment cells in lower vertebrates.
Difference in Pathogenesis Between Vitiligo Vulgaris and Halo Nevi Associated with Vitiligo is Supported by an HLA Association Study
Authors: De Vijlder Hannah C. , Westerhof Wiete , Schreuder Geziena M. Th. , De Lange Peter , Claas Frans H. J.
Published: May 2004

Human leukocyte antigen (HLA) class II associations with two subtypes of vitiligo: vitiligo vulgaris and halo nevi associated with vitiligo were investigated. In previous studies associations between vitiligo and HLA antigens have been reported but these two subtypes have never been taken into account. However from a clinical and histological point of view, a difference in (auto)‐immune pathogenesis can be expected. This difference might be reflected in an association with different HLA alleles. Seventy‐six unrelated Dutch Caucasians, 40 with vitiligo vulgaris and 36 with halo nevi associated with vitiligo were included. A panel of randomly chosen HLA typed healthy Dutch blood donors (n = 2400) served as control population. HLA‐DR and ‐DQ typing was carried out on blood samples by amplifying genomic DNA using polymerase chain reaction followed by dot blot hybridization with sequence specific oligonucleotides. The main outcome measures were odds ratio (OR), uncorrected P‐value (Pu) and corrected P‐value. There were distinct differences in the clinical manifestations between vitiligo vulgaris and halo nevi associated with vitiligo with respect to precipitating factors, extent and progress of the disease and the association with other auto‐immune diseases in the two subtypes and their respective first degree family members. Our stratification reveals differences in HLA class II between both subtypes and between subtypes and controls. A case–control association study showed a significant positive association of HLA‐DR4 (OR = 2.787, Pu = 0.0022) and DR53 (OR = 2.249, Pu = 0.0153) and a negative association of HLA‐DR3 (OR = 0.195, Pu = 0.0024) with vitiligo vulgaris. The group with halo nevi associated with vitiligo did not show these associations, but had a significant negative association with HLA‐DR11 (OR = 0.083, Pu = 0.0067). In conclusion, the differences in HLA association within clinical subtypes of vitiligo support our suggestion that vitiligo vulgaris and halo nevi associated with vitiligo have distinct pathogenic mechanisms.
Cysteine but not Glutathione Modulates the Radiosensitivity of Human Melanoma Cells by Affecting Both Survival and DNA Damage
Authors: Kinnaert E. , Duez P. , Morandini R. , Dubois J. , Van Houtte P. , Ghanem G.
Published: May 2004

Glutathione (GSH) and its precursor cysteine (Cys) are both known to react within any cells with oxidative species and thus play an important role in cellular defense mechanisms against oxidative stress. In melanocytes, these are also important precursors of melanogenesis by reacting non‐enzymatically with l‐dopaquinone to form the sulfur‐containing pheomelanin. Our aim was to assess pigment role in the cellular radioprotection mechanism using a human melanoma cell model of mixed‐type melanin under GSH depletion to obtain a radiosensitizing effect. The latter has been achieved either by Cys deprivation or GSH specific depletion. We first compared cell survival of Cys‐deprived and GSH‐depleted cells vs. control cells. Cys deprivation was achieved by decreasing Cys concentration in the culture medium for 24 h. In this condition, no toxicity was observed, Cys and GSH levels decreased, melanogenesis switched to a higher eumelanin synthesis and cells were significantly more resistant to 10‐Gy dose of ionizing radiations than untreated cells. Glutathione depletion was achieved with the γ‐glutamylcysteine synthetase inhibitor buthionine‐S‐sulfoximine (BSO) for 24 h at 50 μM, a concentration yielding no toxicity. In this condition, intracellular GSH level decreased but no change in pigmentation was observed and cells were slightly but significantly more sensitive to radiation than the control. We then compared DNA radio‐induced damages by Comet assay in control cells, cells treated as above and cells with stimulated pigmentation by increasing Tyr concentration in the medium. Our results showed that, when intracellular eumelanin content increased, DNA damage decreased. By contrast, DNA damage increased in cells treated with BSO alone. It is concluded that increasing the intracellular eumelanin content by the melanin precursor Tyr or by favoring the Pheo‐ to Eumelanin switch, compensates for the loss of the two intracellular radioprotectors that are GSH and Cys.
Spotlight on Spotted Mice: A Review of White Spotting Mouse Mutants and Associated Human Pigmentation Disorders
Authors: Baxter Laura L. , Hou Ling , Loftus Stacie K. , Pavan William J.
Published: May 2004

Mutation of genes that regulate neural crest‐derived melanoblast development and survival can result in reduction and/or loss of mature melanocytes. The reduction in melanocyte number in the skin and hair follicles manifests itself as areas of hypopigmentation, commonly described as white spotting in mice. To date ten genes have been identified which are associated with white‐spotting phenotypes in mouse. Seven of these genes are associated with neural crest and melanocyte disorders in humans. This review summarizes the phenotypes associated with mutation of these genes in both mouse and man. We describe our current understanding of how these genes function in development, and explore their complex roles regulating the various stages of melanocyte development.
Co‐localization of β1,6‐branched Oligosaccharides and Coarse Melanin in Macrophage–Melanoma Fusion Hybrids and Human Melanoma Cells In Vitro
Authors: Rupani Reena , Handerson Tamara , Pawelek John
Published: May 2004

Fusion hybrids between normal macrophages and Cloudman S91 melanoma cells were shown earlier to have increased metastatic potential, along with high expression of β1,6‐N‐acetylglucosaminyltransferase  V and β1,6‐branched oligosaccharides. Curiously, hybrids, but not parental melanoma cells, also produced ‘coarse melanin’– autophagic vesicles with multiple melanosomes. As β1,6‐branched oligosaccharides were known to be associated with metastasis, and coarse melanin had been described in invasive human melanomas, we looked for potential relationships between the two. Using lectin‐ and immunohistochemistry, we analyzed cell lines producing coarse melanin for β1,6‐branched oligosaccharides: gp100/pmel‐17 (a melanosomal structural component) and CD63 (a late endosome/lysosome component associated with melanoma and certain other human cancers). Cell lines used in this study were (i) hybrid 94‐H48, a highly metastatic, macrophage–melanoma experimental fusion hybrid; (ii) 6neo mouse melanoma cells, the weakly metastatic, parental fusion partner; and (iii) SKmel‐23, a human melanoma cell line derived from a metastasis. Coarse melanin granules were prominent both in hybrids and in SKmel‐23 cells, and co‐localized with stains for β1,6‐branched oligosaccharides, gp100/pmel 17, and CD63. This is the first report of this phenotype being expressed in vitro, although co‐expression of β1,6‐branched oligosaccharides and coarse melanin was recently shown to be a common and pervasive characteristic in archival specimens of human melanomas, and was most prominent in metastases. The results suggest that pathways of melanogenesis in melanoma may differ significantly from those in normal melanocytes. In vitro expression of this phenotype provides new biological systems for more detailed analyses of its genesis and regulation at the molecular genetic level.
Study on the Antioxidant Status of Vitiligo Patients of Different Age Groups in Baroda
Authors: Agrawal Deepali , Shajil E. M. , Marfatia Y. S. , Begum Rasheedunnisa
Published: May 2004

One of the major hypotheses in the pathogenesis of vitiligo is the oxidative stress hypothesis. Pollution plays a major role in the production of free radicals. Gujarat, a highly industrialized state in India has a high prevalence of vitiligo patients. No previous studies were done on the age‐dependent antioxidant status of vitiligo patients in Baroda city, Gujarat. Blood samples were collected from vitiligo patients of different age groups (5–15, 16–25, 26–35, 36–45 yr) and from age matched healthy volunteers. Antioxidant enzymes in blood such as catalase, superoxide dismutase, glutathione peroxidase and non‐enzymatic antioxidants such as reduced glutathione and plasma vitamin E were estimated. Lipid peroxidation levels in erythrocytes and the reducing equivalent system, i.e. glucose‐6‐phosphate dehydrogenase were also measured. Significant increase in superoxide dismutase activity and lipid peroxidation levels in erythrocytes was observed in all age groups of vitiligo patients as compared with age‐matched healthy controls, wherein an increase of 55% (P < 0.02) was observed in superoxide dismutase activity and lipid peroxidation levels in 36–45 yr age group. Whole blood glutathione levels, erythrocyte glutathione peroxidase and glucose‐6‐phosphate dehydrogenase activity were decreased significantly, whereas erythrocyte catalase activity and plasma vitamin E levels were not different in vitiligo patients as compared with age‐matched healthy controls. No specific age group showed a significant difference. This is the first report on the age‐dependent antioxidant status of vitiligo patients in Baroda. The disease affects individuals of any age group as shown in this study and systemic oxidative stress might precipitate the pathogenesis of vitiligo in susceptible patients.
Nitric Oxide is Toxic to Melanocytes In Vitro
Authors: Iuga Aurel O. , Qureshi Abrar A. , Lerner Ethan A.
Published: May 2004

Nitric oxide is a diffusible gaseous mediator generated from l‐arginine by inducible and constitutive nitric oxide synthases. It has been associated with cytotoxic effects. Inflammatory cells and Langerhans cells can express the inducible form of nitric oxide synthase and produce large quantities of nitric oxide. The proximity of these cells to melanocytes could result in melanocyte cell death. We studied melanocyte susceptibility to nitric oxide using the nitric oxide donor compound sodium nitroprusside and nitric oxide released by the Langerhans like cell‐line XS‐52 following stimulation with lipopolysaccharide (LPS). Melanocyte lysis, quantified by chromium release in the presence of sodium nitroprusside was both time and concentration dependent. Co‐culture of LPS‐stimulated XS cells with melanocytes also resulted in melanocyte cell death. No cell death was observed when melanocytes alone were exposed to LPS. Melanocytes were killed even when the co‐cultures were performed across Transwells in which there was no direct contact between XS cells and melanocytes. XS‐induced melanocyte death was thus dependent on a diffusible factor consistent with nitric oxide. Cell death was markedly decreased in co‐cultures performed in the presence of hemoglobin, a nitric oxide quencher. The possible role that nitric oxide may play in disorders associated with loss of pigmentation is discussed.
Effects of a new cystic fibrosis transmembrane conductance regulator inhibitor on Cl− conductance in human sweat ducts
Authors: Wang X. F. , Reddy M. M. , Quinton P. M.
Published: May 2004

Effective and specific inhibition of the cystic fibrosis transmembrane conductance regulator (CFTR) Cl− channel in epithelia has long been needed to better understand the role of anion movements in fluid and electrolyte transport. Until now, available inhibitors have required high concentrations, usually in the millimolar or high micromolar range, to effect even an incomplete block of channel conductance. These inhibitors, including 5‐nitro‐2(3‐phenylpropyl‐amino)benzoate (NPPB), bumetamide, glibenclamide and DIDS, are also relatively non‐specific. Recently a new anion channel inhibitor, a thiazolidinone derivative, termed CFTRInh‐172 has been synthesized and introduced with apparently improved inhibitory properties as shown by effects on anion conductance expressed in cell lines and on secretion in vivo. Here, we assay the effect of this inhibitor on a purely salt absorbing native epithelial tissue, the freshly isolated microperfused human sweat duct, known for its inherently high expression of CFTR. We found that the inhibitor at a maximum dose limited by its aqueous solubility of 5 μm partially blocked CFTR when applied to either surface of the membrane; however, it may be somewhat more effective from the cytosolic side (∼70% inhibition). It may also partially inhibit Na+ conductance. The inhibition was relatively slow, with a half time for maximum effect of about 3 min, and showed very slow reversibility. Results also suggest that CFTR Cl− conductance (GCl) was blocked in both apical and basal membranes. The inhibitor appears to exert some effect on Na+ transport as well.
Role of nitric oxide in the reflex diuresis in rabbits during pulmonary lymphatic obstruction
Authors: McCormick K. M. , Gunawardena S. , Ravi K. , Bravo E. M. , Kappagoda C. T.
Published: May 2004

The role of nitric oxide in the reflex diuresis in response to pulmonary lymphatic drainage was examined in anaesthetized, artificially ventilated New Zealand White rabbits. Pulmonary lymphatic drainage was obstructed by raising the pressure in a pouch created from the right external jugular vein. Pulmonary lymphatic obstruction resulted in a significant increase in urine flow from an initial control value of  8.9 ± 0.5 ml (10 min)−1 to 12.1 ± 0.6 ml (10 min)−1 during lymphatic obstruction (mean ±s.e.m.; n= 17, P < 0.001). This increase in urine flow was accompanied by a significant increase in the excretion of sodium. Additionally, renal blood flow remained unchanged during the increase in urine flow caused by lymphatic obstruction. Intravenous infusion of l‐NAME, a non‐selective inhibitor of nitric oxide synthase (NOS), abolished the reflex diuresis. Furthermore, intraperitoneal administration of the relatively selective neuronal NOS blocker, 7‐nitroindazole also abolished the response. It was observed that infusion of a more soluble neuronal NOS blocker, 7‐nitroindazole sodium salt (7‐NINA), into the renal medulla also abolished the reflex diuresis. These findings suggest that the increase in urine flow in rabbits caused by pulmonary lymphatic obstruction is dependent upon the integrity of neuronal NOS activity within the renal medulla.
Effects of chronic heart failure in rats on the recovery of microvascular PO2 after contractions in muscles of opposing fibre type
Authors: McDonough Paul , Behnke Brad J. , Musch Timothy I. , Poole David C.
Published: May 2004

Chronic heart failure (CHF) impairs muscle O2 delivery (Q̇O2) and, at a given O2 uptake (V̇O2), lowers microvascular O2 pressures (PmvO2: determined by the Q̇O2‐to‐V̇O2 ratio), which may impair recovery of high‐energy phosphates following exercise. Because CHF preferentially decreases Q̇O2 to slow‐twitch muscles, we hypothesized that recovery PmvO2 kinetics would be slowed to a greater extent in soleus (SOL: ∼84% type I fibres) than in peroneal (PER: ∼14% type I) muscles of CHF rats. PmvO2 dynamics were determined in SOL and PER muscles of control (CON: n= 6; left ventricular end‐diastolic pressure, LVEDP: ∼3 mmHg), moderate CHF (MOD: n= 7; LVEDP: ∼11 mmHg) and severe CHF (SEV: n= 4; LVEDP: ∼25 mmHg) following cessation of electrical stimulation (180 s; 1 Hz). In PER, neither the recovery PmvO2 values nor the mean response time (MRT; a weighted average of the time to 63% of the overall response) were altered by CHF (CON: 66.8 ± 8.0, MOD: 72.4 ± 11.8, SEV: 69.1 ± 9.5 s). In marked contrast, SOL PmvO2, at recovery onset, was reduced significantly in the SEV group (∼6 Torr) and PmvO2 MRT was slowed with increased severity of CHF (CON: 45.1 ± 5.3, MOD: 63.2 ± 9.4, SEV: 82.6 ± 12.3 s; P < 0.05 CON vs. MOD and SEV). These data indicate that CHF slows PmvO2 recovery following contractions and lowers capillary O2 driving pressure in slow‐twitch SOL, but not in fast‐twitch PER muscle. These results may explain, in part, the slowed recovery kinetics (phosphocreatine and V̇O2) and pronounced fatigue following muscular work in CHF patients.
Baroreceptors and the long‐term control of blood pressure
Authors: Thrasher Terry N.
Published: May 2004

The current consensus is that arterial baroreceptors are vitally important in the short term (seconds to minutes) control of mean arterial pressure (MAP) but are unimportant in determining the long‐term level of MAP. The latter statement is based primarily on two observations: first, that baroreceptors rapidly reset to the prevailing level of MAP and second, that total baroreceptor denervation has no lasting effect on the average daily MAP, although the variability of MAP is increased dramatically. However, recent studies in intact experimental animals have produced results that suggest baroreceptor resetting may not be as rapid or complete as previously thought. Furthermore, reconsideration of the responses to baroreceptor denervation suggest that the condition may accurately represent responses to short‐term baroreceptor unloading but not long‐term unloading. Results obtained using a new model of chronic baroreceptor unloading indicate that the condition results in a sustained increase in MAP. These results strongly suggest that the role of baroreceptors in the long term control of MAP needs to be revisited.
Catecholamine‐induced apoptosis and necrosis in cardiac and skeletal myocytes of the rat in vivo: the same or separate death pathways?
Authors: Goldspink David F. , Burniston Jatin G. , Ellison Georgina M. , Clark William A. , Tan Lip‐Bun
Published: May 2004

High levels of catecholamines are myotoxic but the relative amounts of apoptosis and necrosis have not been established in vivo in cardiac and skeletal muscles. Immunohistochemistry was used to detect and quantify myocyte‐specific necrosis (myosin antibody in vivo) and apoptosis (caspase‐3 antibody in vitro) in the heart and soleus muscles of male Wistar rats that had received single subcutaneous injections of isoprenaline over the range 1 μg to 5 mg [kg body weight (BW)]−1. Peak myocyte apoptosis occurred 3–6 h after, and necrosis 18 h after, a single injection of 5 mg (kg BW)−1 isoprenaline in vivo. In the heart myocyte death was mediated through the β1‐adrenergic receptor whereas myocyte death in the soleus muscle was mediated through the β2‐adrenergic receptor. Cardiomyocyte death was heterogeneously distributed throughout the heart, being greatest in the left ventricle (LV) subendocardium and peaking close to the apex, but with significantly more necrosis than apoptosis. Extensive co‐localization of caspase‐3 and myosin labelling was found in the myocytes of both the heart and the slow‐twitch soleus muscle. This, together with similar spatial distributions and responses to catecholamine doses, suggests that either caspase‐3 activation occurs in necrotic as well as apoptotic myocytes or that a large proportion of apoptotic myocytes progress to secondary necrosis in vivo.
Vagal reflex actions of atrial natriuretic peptide survive physiological but not pathological cardiac hypertrophy in rat
Authors: Hood Sally G. , Woods Robyn L.
Published: May 2004

Atrial natriuretic peptide (ANP) enhances cardiac vagal baroreflexes in normotensive animals. In spontaneously hypertensive rats (SHRs) this effect of ANP was absent. The reflex actions of ANP were preserved if hypertrophy was completely prevented in SHRs. However even a small amount of cardiac hypertrophy, with no hypertension, in SHRs was accompanied by a loss of the reflex bradycardic actions of ANP. In the present study, we investigated whether pathophysiological cardiac hypertrophy, induced by one‐kidney, one‐clip renovascular hypertension (1K‐1C; n= 6), or physiological cardiac hypertrophy induced by chronic spontaneous, wheel‐running exercise training (n= 7), similarly prevented vagal reflex actions of ANP. Cardiac baroreceptor‐activated bradycardia was measured during rapid ramp increases (∼5 s) in blood pressure after bolus doses of methoxamine or vehicle in conscious, chronically instrumented rats during infusions of ANP (50 pmol kg−1 min−1). Compared with uninephrectomised control rats (n= 10), rats with 1K‐1C had cardiac hypertrophy (∼55% increase in left ventricle:body weight (LV:BW) ratio; P < 0.05) and blunted vagal baroreflex gain (−0.93 ± 0.18 versus−0.50 ± 0.13 beats min−1 mmHg−1; P < 0.05). ANP did not augment baroreflex function in 1K‐1C. Compared with their sedentary controls (n= 7), exercise‐trained rats with cardiac hypertrophy (∼20% increase LV:BW ratio; P < 0.05) also had blunted ramp baroreflex bradycardia (−1.28 ± 0.23 versus−0.57 ± 0.09 beats min−1 mmHg−1; P < 0.05). In contrast, ANP more than doubled baroreflex bradycardia in exercise‐trained rats (P < 0.05). The aetiology of cardiac hypertrophy therefore influenced whether ANP retained its vagal baroreflex enhancing properties.
Lactate metabolism: a new paradigm for the third millennium
Authors: Gladden L. B.
Published: May 2004

For much of the 20th century, lactate was largely considered a dead‐end waste product of glycolysis due to hypoxia, the primary cause of the O2 debt following exercise, a major cause of muscle fatigue, and a key factor in acidosis‐induced tissue damage. Since the 1970s, a ‘lactate revolution’ has occurred. At present, we are in the midst of a lactate shuttle era; the lactate paradigm has shifted. It now appears that increased lactate production and concentration as a result of anoxia or dysoxia are often the exception rather than the rule. Lactic acidosis is being re‐evaluated as a factor in muscle fatigue. Lactate is an important intermediate in the process of wound repair and regeneration. The origin of elevated [lactate] in injury and sepsis is being re‐investigated. There is essentially unanimous experimental support for a cell‐to‐cell lactate shuttle, along with mounting evidence for astrocyte–neuron, lactate–alanine, peroxisomal and spermatogenic lactate shuttles. The bulk of the evidence suggests that lactate is an important intermediary in numerous metabolic processes, a particularly mobile fuel for aerobic metabolism, and perhaps a mediator of redox state among various compartments both within and between cells. Lactate can no longer be considered the usual suspect for metabolic ‘crimes’, but is instead a central player in cellular, regional and whole body metabolism. Overall, the cell‐to‐cell lactate shuttle has expanded far beyond its initial conception as an explanation for lactate metabolism during muscle contractions and exercise to now subsume all of the other shuttles as a grand description of the role(s) of lactate in numerous metabolic processes and pathways.
Exposure to cAMP and β‐adrenergic stimulation recruits CaV3 T‐type channels in rat chromaffin cells through Epac cAMP‐receptor proteins
Authors: Novara M. , Baldelli P. , Cavallari D. , Carabelli V. , Giancippoli A. , Carbone E.
Published: May 2004

T‐type channels are expressed weakly or not at all in adult rat chromaffin cells (RCCs) and there is contrasting evidence as to whether they play a functional role in catecholamine secretion. Here we show that 3–5 days after application of pCPT‐cAMP, most RCCs grown in serum‐free medium expressed a high density of low‐voltage‐activated T‐type channels without altering the expression and characteristics of high‐voltage‐activated channels. The density of cAMP‐recruited T‐type channels increased with time and displayed the typical biophysical and pharmacological properties of low‐voltage‐activated Ca2+ channels: (1) steep voltage‐dependent activation from −50 mV in 10 mm Ca2+, (2) slow deactivation but fast and complete inactivation, (3) full inactivation following short conditioning prepulses to −30 mV, (4) effective block of Ca2+ influx with 50 μm Ni2+, (5) comparable permeability to Ca2+ and Ba2+, and (6) insensitivity to common Ca2+ channel antagonists. The action of exogenous pCPT‐cAMP (200 μm) was prevented by the protein synthesis inhibitor anisomycin and mimicked in most cells by exposure to forskolin and 1‐methyl‐3‐isobutylxanthine (IBMX) or isoprenaline. The protein kinase A (PKA) inhibitor H89 (0.3 μm) and the competitive antagonist of cAMP binding to PKA, Rp‐cAMPS, had weak or no effect on the action of pCPT‐cAMP. In line with this, the selective Epac agonist 8CPT‐2Me‐cAMP nicely mimicked the action of pCPT‐cAMP and isoprenaline, suggesting the existence of a dominant Epac‐dependent recruitment of T‐type channels in RCCs that may originate from the activation of β‐adrenoceptors. Stimulation of β‐adrenoceptors occurs autocrinally in RCCs and thus, the neosynthesis of low‐voltage‐activated channels may represent a new form of ‘chromaffin cell plasticity’, which contributes, by lowering the threshold of action potential firing, to increasing cell excitability and secretory activity during sustained sympathetic stimulation and/or increased catecholamine circulation.
Maternal nutrient restriction alters gene expression in the ovine fetal heart
Authors: Han Hyung‐Chul , Austin Kathleen J. , Nathanielsz Peter W. , Ford Stephen P. , Nijland Mark J. , Hansen Thomas R.
Published: May 2004

Adequate maternal nutrient supply is critical for normal fetal organogenesis. We previously demonstrated that a global 50% nutrient restriction during the first half of gestation causes compensatory growth of both the left and right ventricles of the fetal heart by day 78 of gestation. Thus, it was hypothesized that maternal nutrient restriction significantly altered gene expression in the fetal cardiac left ventricle (LV). Pregnant ewes were randomly grouped into control (100% national research council (NRC) requirements) or nutrient‐restricted groups (50% NRC requirements) from day 28 to day 78 of gestation, at which time fetal LV were collected. Fetal LV mRNA was used to construct a suppression subtraction cDNA library from which 11 cDNA clones were found by differential dot blot hybridization and virtual Northern analysis to be up‐regulated by maternal nutrient restriction: caveolin, stathmin, G‐1 cyclin, α‐actin, titin, cardiac ankyrin repeat protein (CARP), cardiac‐specific RNA‐helicase activated by MEF2C (CHAMP), endothelial and smooth muscle derived neuropilin (ESDN), prostatic binding protein, NADH dehydrogenase subunit 2, and an unknown protein. Six of these clones (cardiac α‐actin, cyclin G1, stathmin, NADH dehydrogenase subunit 2, titin and prostatic binding protein) have been linked to cardiac hypertrophy in other species including humans. Of the remaining clones, caveolin, CARP and CHAMP have been shown to inhibit remodelling of hypertrophic tissue. Compensatory growth of fetal LV in response to maternal undernutrition is concluded to be associated with increased transcription of genes related to cardiac hypertrophy, compensatory growth or remodelling. Counter‐regulatory gene transcription may be increased, in part, as a response to moderating the degree of cardiac remodelling. The short‐ and long‐term consequences of these changes in fetal heart gene expression and induction of specific homeostatic mechanisms in response to maternal undernutrition remain to be determined.
Postnatal microbial colonization programs the hypothalamic–pituitary–adrenal system for stress response in mice
Authors: Sudo Nobuyuki , Chida Yoichi , Aiba Yuji , Sonoda Junko , Oyama Naomi , Yu Xiao‐Nian , Kubo Chiharu , Koga Yasuhiro
Published: May 2004

Indigenous microbiota have several beneficial effects on host physiological functions; however, little is known about whether or not postnatal microbial colonization can affect the development of brain plasticity and a subsequent physiological system response. To test the idea that such microbes may affect the development of neural systems that govern the endocrine response to stress, we investigated hypothalamic–pituitary–adrenal (HPA) reaction to stress by comparing germfree (GF), specific pathogen free (SPF) and gnotobiotic mice. Plasma ACTH and corticosterone elevation in response to restraint stress was substantially higher in GF mice than in SPF mice, but not in response to stimulation with ether. Moreover, GF mice also exhibited reduced brain‐derived neurotrophic factor expression levels in the cortex and hippocampus relative to SPF mice. The exaggerated HPA stress response by GF mice was reversed by reconstitution with Bifidobacterium infantis. In contrast, monoassociation with enteropathogenic Escherichia coli, but not with its mutant strain devoid of the translocated intimin receptor gene, enhanced the response to stress. Importantly, the enhanced HPA response of GF mice was partly corrected by reconstitution with SPF faeces at an early stage, but not by any reconstitution exerted at a later stage, which therefore indicates that exposure to microbes at an early developmental stage is required for the HPA system to become fully susceptible to inhibitory neural regulation. These results suggest that commensal microbiota can affect the postnatal development of the HPA stress response in mice.
Tetrahydrobiopterin improves endothelial dysfunction and vascular oxidative stress in microvessels of intrauterine undernourished rats
Authors: Franco Maria do Carmo P. , Fortes Zuleica B. , Akamine Eliana H. , Kawamoto Elisa M. , Scavone Cristoforo , De Britto Luiz Roberto Giorgetti , Muscara Marcelo N. , Teixeira Simone A. , Tostes Rita C. A. , Carvalho Maria Helena C. , Nigro Dorothy
Published: May 2004

In the present study, we investigated the effects of the exogenous application of tetrahydrobiopterin on the endothelium‐dependent vasorelaxation and superoxide anion generation in the mesenteric microvessels of intrauterine undernourished rats. In addition, we investigated the presence of peroxynitrite in these rats by evaluation of nitrotyrosine‐containing proteins, a stable end‐product of peroxynitrite oxidation. For this, female pregnant Wistar rats were fed either normal or 50% of the normal intake diets during the whole gestational period. Male offspring (16 weeks of age) were studied to assess microvascular reactivity, superoxide production using a hydroethidine staining assay, nitric oxide synthase (NOS) activity and nitric oxide (NO) production. Western blot analysis was used to quantify nitrotyrosine‐containing proteins and relative multiplex RT‐PCR analysis for endothelial NOS (eNOS) mRNA expression. Superfusion with tetrahydrobiopterin significantly decreased superoxide generation and improved vascular function. Intrauterine malnutrition induced a decrement of NOS activity and NO production without affecting the gene expression of eNOS. However, incubation with tetrahydrobiopterin significantly improved NO production after stimulation with acetylcholine or bradykinin in intrauterine undernourished rats. The fact that the nitrotyrosine‐containing proteins were increased could, at first sight, suggest that the peroxynitrite is the mediator responsible for the excessive oxidation and depletion of tetrahydrobiopterin. Our study shows that exogenous application of tetrahydrobiopterin leads to a significant improvement of endothelium‐dependent vasodilatation, enhanced NO production and decreased superoxide generation in microvessels of intrauterine undernourished rats. Since we found a decrease in NOS activity without an alteration in the gene expression of eNOS, we suggest that impaired NOS‐dependent responses of mesenteric arterioles are related to the impairment of tetrahydrobiopterin pathways.
Mitochondrial calcium sequestration and protein kinase C cooperate in the regulation of cortical F‐actin disassembly and secretion in bovine chromaffin cells
Authors: Cuchillo‐Ibáñez I. , Lejen T. , Albillos A. , Rosé S. D. , Olivares R. , Villarroya M. , García A. G. , Trifaró J.‐M.
Published: May 2004

Mitochondria play an important role in the homeostasis of intracellular Ca2+ and regulate its availability for exocytosis. Inhibitors of mitochondria Ca2+ uptake such as protonophore CCCP potentiate the secretory response to a depolarizing pulse of K+. Exposure of cells to agents that directly (cytochalasin D, latrunculin B) or indirectly (PMA) disrupt cortical F‐actin networks also potentiate the secretory response to high K+. The effects of cytochalasin D and CCCP on secretion were additive whereas those of PMA and CCCP were not; this suggests different mechanisms for cytochalasin D and CCCP and a similar mechanism for PMA and CCCP. Mitochondria were the site of action of CCCP, because the potentiation of secretion by CCCP was observed even after depletion of Ca2+ from the endoplasmic reticulum. CCCP induced a small increase in the cytosolic Ca2+ concentration ([Ca2+]c) that was not modified by the protein kinase C (PKC) inhibitor chelerythrine. Both CCCP and PMA induced cortical F‐actin disassembly, an effect abolished by chelerythrine. In addition, rotenone and oligomycin A, two other mitochondrial inhibitors, also evoked cortical F‐actin disassembly and potentiated secretion; again, these effects were blocked by chelerythrine. CCCP also enhanced the phosphorylation of PKC and myristoylated alanine‐rich C kinase substance (MARCKS), and these were also inhibited by chelerythrine. The results suggest that the rapid sequestration of Ca2+ by mitochondria would protect the cell from an enhanced PKC activation and cortical F‐actin disassembly, thereby limiting the magnitude of the secretory response.
Fluctuations and Bistabilities on Catalyst Nanoparticles
Authors: Johánek V. , Laurin M. , Grant A. W. , Kasemo B. , Henry C. R. , Libuda J.
Published: May 2004

We show that coverage fluctuations on catalyst particles can drastically alter their macroscopic catalytic behavior. Scrutinizing the occurrence of kinetic bistabilities, it is demonstrated by molecular beam experiments on model catalysts that macroscopically observable bistabilities vanish completely with decreasing particle size, as previously predicted by theory. The effect is attributed to fluctuation-induced transitions between two kinetic reaction regimes, with a transition rate controlled by both particle size and surface defects. These results suggest that fluctuation-induced effects represent a general phenomenon affecting the reaction kinetics on nanostructured surfaces.
Use of Monoclonal Antibodies That Recognize p60 for Identification of
Listeria monocytogenes
Authors: Yu Kang-Y. , Noh Youngsoon , Chung Minsub , Park Hong-J. , Lee Namseok , Youn Moonyeon , Jung Byeong Y. , Youn Byung-S.
Published: May 2004

Listeria monocytogenes
causes major food-borne outbreaks of disease worldwide. Specific identification of this microorganism is of utmost importance to public health and industry.
Listeria
species are known to secrete a 60-kDa protein collectively termed p60, which is encoded by the
iap
(invasion-associated protein) gene and secreted in large quantities into the growth media. p60 is a highly immunogenic murein hydrolase that is essential for cell division. Due to these properties, p60 is an ideal diagnostic target for the development of immunological detection systems for
L. monocytogenes
. We report here two independent lines of monoclonal antibody (MAb): p6007, which specifically recognizes
L. monocytogenes
p60, and p6017, which reacts with a wide range of
Listeria
p60 proteins. By combining these antibodies with a polyclonal antibody, we developed efficient sandwich enzyme-linked immunosorbent assay (ELISA) systems which can specifically identify
L. monocytogenes
or generally detect
Listeria
species. Since an excess amount of the peptide corresponding to PepA or PepD did not interfere with the ELISA, and direct ELISAs were unable to detect both peptides, we concluded that the epitope presumed to be recognized by p6007 or p6017 could be distinguished from PepA and PepD as described by Bubert et al. (Appl. Environ. Microbiol. 60:3120-3127, 1997). To our best knowledge, this is the first example of an immunological identification system that uses p60-recognizing MAbs.
Intraperitoneal Injection of Lipopolysaccharide Induces Dynamic Migration of Gr-1highPolymorphonuclear Neutrophils in the Murine Abdominal Cavity
Authors: Miyazaki Shuichi , Ishikawa Fumio , Fujikawa Toshihiko , Nagata Shigekazu , Yamaguchi Keizo
Published: May 2004

Intraperitoneal injection of lipopolysaccharide (LPS; 100 μg) in mice resulted in the disappearance of almost all proteose peptone-induced polymorphonuclear neutrophils (PMNs) with high-level fluorescence for the cell surface marker Gr-1 (Gr-1high) at 15 min postinjection, followed by doubling of their proportion at 30 min postinjection. High staining levels of 3′-acetyl-2′-carboxyl-6′,7′-(dihyropyran-2′-one)-5 or 6-carboxyfluorescein diacethoxylmethyl ester-labeled PMNs injected into the peritoneal cavity were detected in mesenteric lymph nodes 15 min postinjection of LPS. Therefore, the time of decrease of Gr-1highPMNs coincided with that of the increase in cell accumulation in mesenteric lymph nodes. Since milk fat globule-EGF factor 8 (MFG-E8), which is secreted by macrophages, bound many PMNs exhibiting Gr-1highand Gr-1mediumat 30 min postinjection of LPS, the staining level of annexin V on those cells was very low because its binding site is the same as the receptor for MFG-E8. At 60 min postinjection of LPS, the proportion of Gr-1highPMNs decreased, and almost all Gr-1mediumPMNs tended to shift to the right compared with those at 30 min postinjection. The geomeans of Toll-like receptor 4 (TLR4) expression on PMNs at 15, 30, and 60 min postinjection of LPS were 63, 66, and 24%, respectively, compared with that on normal PMNs, indicating that the expression of TLR4 decreases in response to exposure to LPS. Our results suggest that LPS induced PMN death and that many PMNs expressing Gr-1highundergo apoptosis 180 min postinjection of LPS.
Escherichia coliandStaphylococcus aureusElicit Differential Innate Immune Responses following Intramammary Infection
Authors: Bannerman Douglas D. , Paape Max J. , Lee Jai-Wei , Zhao Xin , Hope Jayne C. , Rainard Pascal
Published: May 2004

Staphylococcus aureusandEscherichia coliare among the most prevalent species of gram-positive and gram-negative bacteria, respectively, that induce clinical mastitis. The innate immune system comprises the immediate host defense mechanisms to protect against infection and contributes to the initial detection of and proinflammatory response to infectious pathogens. The objective of the present study was to characterize the different innate immune responses to experimental intramammary infection withE. coliandS. aureusduring clinical mastitis. The cytokine response and changes in the levels of soluble CD14 (sCD14) and lipopolysaccharide-binding protein (LBP), two proteins that contribute to host recognition of bacterial cell wall products, were studied. Intramammary infection with eitherE. coliorS. aureuselicited systemic changes, including decreased milk output, a febrile response, and induction of the acute-phase synthesis of LBP. Infection with either bacterium resulted in increased levels of interleukin 1β (IL-1β), gamma interferon, IL-12, sCD14, and LBP in milk. High levels of the complement cleavage product C5a and the anti-inflammatory cytokine IL-10 were detected at several time points followingE. coliinfection, whereasS. aureusinfection elicited a slight but detectable increase in these mediators at a single time point. Increases in IL-8 and tumor necrosis factor alpha were observed only in quarters infected withE. coli. Together, these data demonstrate the variability of the host innate immune response toE. coliandS. aureusand suggest that the limited cytokine response toS. aureusmay contribute to the well-known ability of the bacterium to establish chronic intramammary infection.