bluuebunny/crossref_metadata_2025 huggingface dataset:
Growth Hormone Size Variants: Changes in the Pituitary During Development of the Chicken
Authors: Arámburo Carlos , Luna Maricela , Carranza Martha , Reyes Marisa , Martínez‐Coria Hilda , Scanes Colin G.
Published: August 2010

Abstract. There is considerable evidence for the existence of structural variants of growth hormone (GH). The chicken is a useful model for investigating GH heterogeneity as both size and charge immunoreactive‐(ir) variants have been observed in the pituitary and plasma. The present study examined the size distribution of ir‐GH in the pituitary gland of chicken, from late embryogenesis through adulthood. Pituitaries were homogenized in the presence of protease inhibitor, and the GH size variants were separated by SDS‐PAGE, transferred by Western blotting, immunostained with a specific antiserum to chicken GH, and quantitated by chemiluminescence followed by laser densitometry (chemiluminescent assay). Under nonreducing conditions ir‐GH bands of 15, 22, 25, 44, 50, 66, 80, 98, 105 and >110 kDa were observed. Both the relative proportion of the GH size variants and the total pituitary content varied with developmental stage and age. The proportion of the 15‐kDa fragment was greatest in the embryonic stage, and then it decreased. The proportion of the monomeric 22‐kDa form was lowest at 18 days of embryogenesis (dE) and highest at 20 dE. In contrast, the high MW forms (≥66 kDa) were lowest in embryos, and they increased (P < 0.05) after hatching. The 22‐, 44‐, 66‐, and 80‐kDa forms were assayed for activity by radioreceptor assay following isolation by semipreparative SDS‐PAGE. Only the 22‐kDa GH variant showed radioreceptor activity. Under reducing conditions for SDS‐PAGE, ir‐GH bands of 13, 15, 18, 23, 26, 36, 39, 44, 48, 59 and 72 kDa were oberved, but most of the high MW form disappeared. There was a concomitant increase in the proportion of the monomeric band and of several submonomeric forms. The present data indicate that the expression, processing, and/or release of some if not all size variants are under some differential control during growth and development of the chicken.
Pyruvate: Metabolic Protector of Cardiac Performance
Authors: Mallet Robert T.
Published: August 2010

Abstract.Pyruvate, a metabolic product of glycolysis and an oxidizable fuel in myocardium, increases cardiac mechanical performance and energy reserves, especially when supplied at supraphysiological concentrations. The inotropic effects of pyruvate are most impressive in hearts that have been reversibly injured (stunned) by ischemia/reperfusion stress. Glucose appears to be an essential co‐substrate for pyruvate's salutary effects in stunned hearts, but other fuels including lactate, acetate, fatty acids, and ketone bodies produce little or no improvement in postischemic function over glucose alone. In contrast to pharmacological inotropism by catecholamines, metabolic inotropism by pyruvate increases cardiac energy reserves and bolsters the endogenous glutathione antioxidant system. Pyruvate enhancement of cardiac function may result from one or more of the following mechanisms: increased cytosolic ATP phosphorylation potential and Gibbs free energy of ATP hydrolysis, enhanced sarcoplasmic reticular calcium ion uptake and release, decreased cytosolic inorganic phosphate concentration, oxyradical scavengingviadirect neutralization of peroxides and/or enhancement of the intracellular glutathione/NADPH antioxidant system, and/or closure of mitochondrial permeability transition pores. This review aims to summarize evidence for each of these mechanisms and to consider the potential utility of pyruvate as a therapeutic intervention for clinical management of cardiac insufficiency.
Influence of Aging and Caloric Restriction on Activation of Ras/MAPK, Calcineurin, and CaMK‐IV Activities in Rat T Cells
Authors: Pahlavani Mohammad A. , Vargas Daniel M.
Published: August 2010

Abstract. The signaling cascade mediated by Ras (p21ras) and MAPK (mitogen‐activated protein kinase) and calcium/calmodulin regulating enzymes, calcineurin (CaN) and CaMK‐IV, are considered to be essential for T‐cell growth and function. In the present study, the effect of aging and caloric restriction (CR) on the induction of Ras and MAPK activation by concanavalin A (ConA) was studied. Splenic T cells were isolated from young (4–6 months) and old (22–24 months) rats that had free access to food (control group), and from caloric restricted old (22–24 months) rats that beginning at 6 weeks of age were fed 60%(40% caloric restriction) of the diet consumed by the control rats. We found that the induction of Ras activity in T cells isolated from control old rats was lower (P < 0.001) than that in control young rats. However, the levels of Ras activity in T cells isolated from CR old rats were similar to the levels in the age‐matched control rats. The induction of MAPK activity in T cells isolated from control old rats and CR old rats was significantly less than in T cells isolated from control young rats, and caloric restriction significantly (P < 0.05) reduced the age‐related decline in MAPK activation. We also measured the induction of CaN and CaMK‐IV activities by ConA in T cells from control young and old and CR old rats. The induction of both CaN and CaMK‐IV activity decreased with age. Caloric restriction significantly (P < 0.05) reduced the age‐related decline in CaN activity, but had no significant effect on CaMK‐IV activity. The changes in Ras/MAPK activation and in CaN and CaMK‐IV activity with age or with CR were not associated with alterations in their corresponding protein levels. Thus, caloric restriction has a differential effect on the activation of the upstream signaling molecules that are altered with age.
Zinc Deficiency Exacerbates Loss in Blood‐Brain Barrier Integrity Induced by Hyperoxia Measured by Dynamic MRI
Authors: Noseworthy Michael D. , Bray Tammy M.
Published: August 2010

Abstract. Using dynamic Magnetic Resonance Imaging (dMRI), blood‐brain barrier (BBB) permeability (kPSρ) and tissue interstitial leakage space (ve) were evaluated in zinc‐deficient (ZnDF) male weanling Wistar rats following 3 days exposure to hyperoxia (85% O2). Temporal monitoring of T1‐weighted MR image changes, following a bolus intravenous injection of gadolinium‐DTPA, allowed estimation of BBB integrity. Three‐day exposure of hyperoxia caused a marginal loss of BBB integrity, reflected in a slight increase in kPSρ and ve, observed in both the animals fed adequate zinc (ZnAL) and pair‐fed controls (ZnPF). However, zinc deficiency resulted in a significant increase in both kPSρ and ve, indicating a severely disturbed BBB. In addition MR‐visible free water was elevated in ZnDF brains following hyperoxia treatment indicating that a loss of BBB integrity may be associated with neuronal edema. The diminished BBB integrity may be free‐radical mediated as the ratio of oxidized to reduced glutathione (GSSG:GSH) was significantly elevated.
Proarrhythmia of Azimilide and Other Class III Antiarrhythmic Agents in the Adrenergically Stimulated Rabbit
Authors: Brooks Robert R. , Drexler Anne P. , Maynard Anne E. , Al‐Khalidi Hussein , Kostreva David R.
Published: August 2010

Abstract.The ventricular proarrhythmic actions of five class III antiarrhythmic agents were compared in the Carlsson rabbit model. In adrenergically stimulated anesthetized rabbits, azimilide, clofilium, dofetilide, sematilide, andd,l‐sotalol caused premature ventricular contractions and nonsustained and sustained ventricular tachyarrhythmias (NSVT and SVT) at pharmacologically equivalent intravenous doses that increased QTc intervals 20% (ED20). There were no significant differences between agents in the percentage of rabbits with serious arrhyhthmias at the ED20doses of 5.2, 0.033, 0.015, 0.66, and 2.8 mg/kg iv, respectively. Proarrhythmia was dose‐dependent. Linear regression analysis of arrhythmia score versus log dose estimated the NSVT doses as 6.2, 0.055, 0.0089, 1.5, and 5.7, respectively. Analysis of arrhythmia states during a 10‐min window after infusion when QTc prolongation was 20% showed that the compounds differed significantly in the proportion of time treated rabbits spent in SVT and combined NSVT and SVT. Rabbits treated with azimilide spent significantly less time in SVT and combined NSVT and SVT, followed in order of increasing time byd,l‐sotalol, sematilide, clofilium, and dofetilide.
Feeding Reduces Activity of Growth Hormone‐Releasing Hormone and Somatostatin Neurons
Authors: McMahon C. D. , Chapin L. T. , Lookingland K. J. , Radcliff R. P. , Tucker H. A.
Published: August 2010

Abstract.Secretion of growth hormone (GH) is synchronized among castrate male cattle (steers) around feeding when access to feed is restricted to a 2‐hr period each day. Typically, concentrations of GH increase before and decrease after feeding. Our objectives were to determine whether i) concentrations of GH decrease in blood after start of feeding; ii) activity of immunoreactive growth hormone‐releasing hormone (GHRH‐ir) neurons decreases in the arcuate nucleus (ARC) after feeding; iii) activity of immunoreactive somatostatin (SS‐ir) neurons in the periventricular nucleus (PeVN) and ARC increase after feeding; and iv) GHRH stimulates release of GH to a similar magnitude at 0900 and at 1300 hr, in steers fed between 1000 and 1200 hr. Blood samples were collected at 20‐min intervals from 0700 to 1300 hr. Groups of steers were euthanized at 0700, 0900, 1100, and 1300 hr (n= 5 per group). Dual‐label immunohistochemistry was performed on free‐floating sections of hypothalami using antibodies directed against Fos and Fos‐related antigens (Fos/FRA) as a marker of neuronal activity in immunoreactive GHRH and SS neurons. Concentrations of GH were high before and decreased after feeding. The percentage of SS‐ir neurons containing Fos/FRA‐ir in the PeVN was 50% lower (P< 0.01) at 1100 hr and 36% lower (P< 0.05) at 1300 hr than at 0900 hr. There was no change in percentage of SS‐ir neurons containing Fos/FRA‐ir in the ARC. The percentage of GHRH‐ir neurons containing Fos/FRA‐ir in the ARC was 66% lower (P< 0.05) at 1100 hr and 65% lower (P< 0.05) at 1300 hr than at 0700 hr. In contrast, the number of GHRH‐ir neurons increased from 0700 to 1300 hr. GHRH‐induced release of GH was suppressed at 1300 hr compared with 0900 hr. In conclusion, reduced basal and GHRH‐induced secretion of GH after feeding was associated with decreased activity of GHRH neurons in the ARC and decreased activity of SS neurons in the PeVN.
Anticancer Agents Suppressive for Adult Parasites of Filariasis in Mongolian Jirds
Authors: Kinnamon Kenneth E. , Engle Robert R. , Poon Bing T. , Ellis William Y. , McCall John W. , Dzimianski Michael T.
Published: August 2010

Abstract. Eight chemical structures not previously reported to possess antifilarial activity have been identified. A total of 79 compounds with anticancer properties were evaluated for possible macrofilaricidal activity against Brugia pahangi and Acanthocheilonema viteae transplanted into male Mongolian jirds (Meriones unguiculatus). All eight active compounds were suppressive for the onchocerciasis type (Acanthocheilonema viteae) of the disease. None was macrofilaricidal for the lymphatic form (Brugia pahangi). These new structures may represent a nucleus around which effective drugs can be synthesized.
Intraluteal Administration of a Nitric Oxide Synthase Blocker Stimulates Progesterone and Oxytocin Secretion and Prolongs the Life Span of the Bovine Corpus Luteum
Authors: Jaroszewski Jerzy Jan , Hansel William
Published: August 2010

Abstract.To test the role of nitric oxide (NO) in secretory functions of bovine corpora lutea (CL), two groups of four Holstein heifers each were treated as follows: Group 1, Nω‐Nitro‐L‐Arginine Methyl Ester (L‐NAME), an inhibitor of nitric oxide synthase (NOS), on Day 11 or 12 of the cycle and Group 2, L‐NAME on Days 17 and 18 of the cycle. All treatments were administered by an intraluteal microdialysis system (MDS). Drugs were infused for 4‐hr periods on the designated days, and the treatment periods were preceded and followed by 4‐hr control periods. Perfusate and jugular blood samples were collected at half‐hour intervals. Perfusate samples were analyzed for progesterone (P4), oxytocin (OT), prostaglandin F2α(PGF2α), and leukotriene C4(LTC4); jugular plasma samples were analyzed for P4, OT, and LH. Perfusion of L‐NAME on Day 11 or 12 consistently increased P4concentration in the perfusate, but had no effect on the life span of the CL. Perfusion of L‐NAME on Days 17–18 also elevated P4levels in the perfusate, and in addition, maintained P4levels in the plasma of three of the four treated animals through Day 25 of the cycle. L‐NAME perfusion also increased OT release concomitant with P4into the perfusate at both the mid‐ and late‐luteal phase treatments. For the most part, concentrations of LH, OT, and P4in the jugular plasma samples collected during the perfusions were unaffected by treatments. L‐NAME perfusion caused small, but significant (P< 0.05) increases in perfusate PGF2αand LTC4at Days 17 and 18 and in LTC4on Day 11 or 12. These data indicate that NO plays a direct luteolytic role in regression of the bovine CL.
Water‐Soluble Hexasulfobutyl[60]fullerene Inhibit Low‐Density Lipoprotein Oxidation in Aqueous and Lipophilic Phases
Authors: Lee Yuan‐Teh , Chiang Long‐Yong , Chen Wei‐Jao , Hsu Hsiu‐Ching
Published: August 2010

Abstract.Oxidative modification of low‐density lipoprotein (LDL) plays a pivotal role in the pathogenesis of atherosclerosis. Increasing the resistance of LDL to oxidation may therefore mitigate, or even prevent, atherosclerosis. A new water‐soluble C60derivative, hexasulfobutyl[60]fullerene [C60− (CH2CH2CH2CH2‐SO3Na)6; FC4S], consisting of 6 sulfobutyl moieties covalently bound onto the C60cage is a potent free radical scavenger. This study explored the antioxidative effect of sulfobutylated fullerene derivatives (FC4S) on LDL oxidation. FC4S was found to be effective in protecting LDL against oxidation induced by either Cu2+or azo peroxyl radicals generated initially in the aqueous or lipophilic phase, respectively. Levels of the oxidative products, conjugated diene and thiobarbituric acid‐reactive substances, and the relative electrophoresis mobility of the LDL were decreased. The addition of 20 μMFC4S at the early stage of oxidation increased the kinetic lag time from 69 ± 11 to 14 ± 10 min (P< 0.05) and decreased the propagation rate from 17.1 ± 2.6 to 6.3 ± 1.0 mOD/min (P< 0.005). Persistent suppression of peroxidation reaction was observed upon further addition of FC4S after full consumption of all endogenous antioxidants during the propagation period. Intravenous injection of hypercholesterolemic rabbits with FC4S (1 mg/kg/day) efficiently decreased atheroma formation. Data substantiate the use of FC4S as an excellent hydrophilic antioxidant in protecting atheroma formation,viaremoving free radicals, in either aqueous or lipophilic phase.
Antisperm Antibodies Associated with Infertility: Properties and Encoding Genes of Target Antigens
Authors: Koide Samuel S. , Wang Linfang , Kamada Masaharu
Published: August 2010

Abstract.Infertility among couples of reproductive age is a perplexing condition when the cause is indeterminate. These cases are classified as unexplained infertility. In a subset of subjects, antisperm antibodies with sperm agglutinating and/or immobilizing activities have been detected in the blood or fluids of the reproductive tract. These cases are designated as immunologic infertility although a cause and effect relationship of the antibodies to infertility has not been established. In this review, seven target sperm antigens to antibodies associated with infertility and their encoding genes are described. The antisperm antibodies (ASAs) examined were obtained from infertile women or were monoclonal antibodies (mAb) raised against human sperm proteins. All the ASAs studied possessed potent sperm agglutinating and/or immobilizing activities. The target antigens were isolated from human and other mammalian sperm, and the encoding genes identified. The seven antigens are YWK‐II, BE‐20, rSMP‐B, BS‐63 (nucleoporin‐related), BS‐17 (calpastatin), HED‐2 (zyxin), and 75‐ kDa. Each antigen is a distinct and separate entity and is produced by different cells of the reproductive tract, (e.g., germ cells, epididymal epithelial cells, and Sertoli cells). No single predominant target component has been found to interact with the ASAs. It is proposed that immunologic infertility is the consequence of the combined actions of multiple ASAs in immobilizing and/or agglutinating spermatozoa, blocking spermegg interaction, preventing implantation, and/or arresting embryo development.
Differential Effects of Nicotine and Aging on Splenocyte Proliferation and the Production of Th1‐ Versus Th2‐Type Cytokines
Authors: Hallquist Nora , Hakki Amal , Wecker Lynn , Friedman Herman , Pross Susan
Published: August 2010

Abstract.Nicotine has a multitude of biological actions in the central and peripheral nervous systems where nicotinic acetylcholine receptors are found. Nicotinic acetylcholine receptors have also been identified on immune cells, but the effects of nicotine on immune responses are not well characterized. These studies tested the hypotheses that nicotine has an effect on both T‐lymphocyte proliferation and the production of cytokines by activated T cells, processes that are necessary for effective T‐cell–mediated immune responses. In addition, the effects of nicotine on these immune responses in aging animals and the effects of nicotine exposure prior to immunostimulation were investigated. Murine splenocytes were exposed to nicotine and stimulated with concanavalin A (ConA). The highest concentration of nicotine (128 μg/ml) significantly depressed proliferation of T cells both when nicotine and ConA were added concurrently and when nicotine was added 3 hr prior to ConA. Nicotine, added concurrently with ConA at concentrations between 0.25 and 64 μg/ml, significantly inhibited the production of IL‐10 by splenocytes from young adult mice, whereas the inhibition of production of IL‐10 by splenocytes from old mice was significantly inhibited, but the response was more variable, depending on the nicotine concentration. In contrast, the production of IFN‐γ by splenocytes from either young adult or old mice was not affected when nicotine (0.016–64 μg/ml) was added concurrently with ConA. Pre‐exposure to 1 μg/ml of nicotine for 3 hr significantly enhanced the production of IFN‐γ by splenocytes from young adult mice, whereas pre‐exposure to 0.016 μg/ml of nicotine tended to but did not significantly enhance IFN‐γ production. Nicotine is now being used as an over‐the‐counter drug by people who differ in age and general immunocompetence. Therefore, the effects of nicotine on immune responses, independent from the effects of the other chemicals found in tobacco, need to be investigated.
Advances in Livestock Nuclear Transfer
Authors: Kühholzer B. , Prather R. S.
Published: August 2010

Abstract. Cloning and transgenic animal production have been greatly enhanced by the development of nuclear transfer technology. In the past, genetic modification in domestic animals was not tightly controlled. With the nuclear transfer technology one can now create some domestic animals with specific genetic modifications. An ever‐expanding variety of cell types have been successfully used as donors to create the clones. Both cell fusion and microinjection are successfully being used to create these animals. However, it is still not clear which stage(s) of the cell cycle for donor and recipient cells yield the greatest degree of development. While for the most part gene expression is reprogrammed in nuclear transfer embryos, all structural changes may not be corrected as evidenced by the length of the telomeres in sheep resulting from nuclear transfer. Even after these animals are created the question of “are they really clones?” arises due to mitochondrial inheritance from the donor cell versus the recipient oocyte. This review discusses these issues as they relate to livestock.
Thiamine Intestinal Transport and Related Issues: Recent Aspects
Authors: Rindi Gianguido , Laforenza Umberto
Published: August 2010

Abstract.In the intestinal lumen thiamine is in free form and very low concentrations. Absorption takes place primarily in the proximal part of the small intestine by means of a dual mechanism, which is saturable at low (physiological) concentrations and diffusive at higher. Thiamine undergoes intracellular phosphorylation mainly to thiamine pyrophosphate, while at the serosal side only free thiamine is present. Thiamine uptake is enhanced by thiamine deficiency, and reduced by thyroid hormone and diabetes. The entry of thiamine into the enterocyte, as evaluated in brush border membrane vesicles of rat small intestine in the absence of H+gradient, is Na+‐ and biotransformation‐independent, completely inhibited by thiamine analogs and reduced by ethanol administration and aging. The transport involves a saturable mechanism at low concentrations of vitamin and simple diffusion at higher. Outwardly oriented H+gradients enhance thiamine transport, whose saturable component is a Na+‐independent electroneutral uphill process utilizing energy supplied by the H+gradient, and involving a thiamine/ H+1:1 stoichiometric exchange. The exit of thiamine from the enterocyte, as evaluated in basolateral membrane vesicles, is Na+‐dependent, directly coupled to ATP hydrolysis by Na+‐K+‐ATPase, and inhibited by thiamine analogs. Transport of thiamine by renal brush border membrane vesicles is similar to the intestinal as far as both H+gradient influence and specificity are concerned. In the erythrocyte thiamine transport is a Na+‐independent, electroneutral process yet with two components: saturable, prevailing at low thiamine concentrations, and diffusive at higher. The saturable (specific) component is missing in patients of the rare disease known as thiamine‐responsive megaloblastic anaemia (TRMA), producing a general disturbance of thiamine transport up to thiamine deficiency. The TRMA gene is located in chromosome 1q23.3. Recently, the thiamine transporter has been cloned: it is a protein of 497 aminoacid residues with high homology with the reduced‐folate transporter.
Characterization of Serum Dehydroepiandrosterone Secretion in Golden Hamsters
Authors: Pieper David R. , Lobocki Catherine A.
Published: August 2010

Abstract. Dehydroepiandrosterone (DHEA) is an adrenal androgen whose function is poorly understood. Although DHEA and DHEA sulfate (DHEAS) are secreted in relatively high quantities by the human adrenal, the laboratory rat secretes very little, thus hindering experimental studies of the hormone. In this paper, we measured the changes in serum DHEA and DHEAS under various physiological conditions in golden hamsters. Evening serum DHEAS fell from 6.30 ± 0.78 μg/dl (mean ± SE) before surgery to 3.03 ± 0.23 μg/dl 12 days after bilateral adrenalectomy. Hamsters had higher levels of DHEA and DHEAS in the evening than in the morning, but removal of the gonads did not consistently decrease serum DHEA or DHEAS in males or females. Evening levels of DHEA and DHEAS reached a peak around 7 weeks of age and then gradually decreased to about one‐third of these levels by one year of age. These results suggest that DHEA and DHEAS are secreted at least in part from the hamster adrenal, that they do not originate from the gonads, and that there is a daily rhythm with peak levels at a time of day just preceding the active phase. In addition, the levels of these hormones decrease with aging.
Antiproliferative and Apoptotic Effects of Tocopherols and Tocotrienols on Preneoplastic and Neoplastic Mouse Mammary Epithelial Cells
Authors: McIntyre Barry S. , Briski Karen P. , Gapor Abdul , Sylvester Paul W.
Published: August 2010

Abstract. Studies were conducted to determine the comparative effects of tocopherols and tocotrienols on preneoplastic (CL‐S1), neoplastic (‐SA), and highly malignant (+SA) mouse mammary epithelial cell growth and viability in vitro. Over a 5‐day culture period, treatment with 0–120 μMα‐ and γ‐tocopherol had no effect on cell proliferation, whereas growth was inhibited 50% (IC50) as compared with controls by treatment with the following: 13, 7, and 6 μM tocotrienol‐rich‐fraction of palm oil (TRF); 55, 47, and 23 μMδ‐tocopherol; 12, 7, and 5 μMα‐tocotrienol; 8, 5, and 4 μMγ‐tocotrienol; or 7, 4, and 3 μMδ‐tocotrienol in CL‐S1, ‐SA and +SA cells, respectively. Acute 24‐hr exposure to 0–250 μMα‐ or γ‐tocopherol (CL‐S1, ‐SA, and +SA) or 0–250 μMδ‐tocopherol (CL‐S1) had no effect on cell viability, whereas cell viability was reduced 50% (LD50) as compared with controls by treatment with 166 or 125 μMδ‐tocopherol in ‐SA and +SA cells, respectively. Additional LD50 doses were determined as the following: 50, 43, and 38 μM TRF; 27, 28, and 23 μMα‐tocotrienol; 19, 17, and 14 μMγ‐tocotrienol; or 16, 15, or 12 μMδ‐tocotrienol in CL‐S1, ‐SA, and +SA cells, respectively. Treatment‐induced cell death resulted from activation of apoptosis, as indicated by DNA fragmentation. Results also showed that CL‐S1, ‐SA, and +SA cells preferentially accumulate tocotrienols as compared with tocopherols, and this may partially explain why tocotrienols display greater biopotency than tocopherols. These data also showed that highly malignant +SA cells were the most sensitive, whereas the preneoplastic CL‐S1 cells were the least sensitive to the antiproliferative and apoptotic effects of tocotrienols, and suggest that tocotrienols may have potential health benefits in preventing and/or reducing the risk of breast cancer in women.
Inhibition of Ethanol‐Induced Liver Disease in the Intragastric Feeding Rat Model by Chlormethiazole
Authors: Gouillon Z.‐Q. , Lucas D. , Li J. , Hagbjork A. L. , French B. A. , Fu P. , Fang C. , Ingelman‐Sundberg M. , Donohue T. M. , French S. W.
Published: August 2010

Abstract.The purpose of this investigation was to assess the effect of chlormethiazole treatment on liver damage in the experimental rat intragastric ethanol‐feeding model of alcoholic liver disease. Chlormethiazole has been used in the treatment of alcoholic withdrawal and has been shown to inhibit cytochrome P4502E1. Since treatment of experimental alcoholic liver disease with CYP2E1 inhibitors had an ameliorating effect on liver injury in the rat, chlormethiazole was used to see if it had a similar effect. Rats fed ethanol for 2 months had significantly less liver injury when chlormethiazole was added to the diet, fed intragastrically. The CYP2E1 apoprotein levels, which were increased by ethanol feeding, were also increased when chlormethiazole was fed with ethanol. Chlormethiazole inhibited the increase in the ethanol‐induced CYP2E1 activityin vivo,as measured by chlorzoxazone 6‐hydroxylation, but did not affect the level of CYP2E1 apoprotein. Likewise, the reduction in proteasome proteolytic enzyme activity produced by ethanol feeding was blunted in chlormethiazole‐fed rats. These results support the conclusion that chlormethiazole treatment partially protects the liver from injury by inhibiting CYP2E1 activityin vivo.
The Cannabinoid System and Cytokine Network
Authors: Klein Thomas W. , Lane Brian , Newton Catherine A. , Friedman Herman
Published: August 2010

Abstract.Many advances have been made in the last few years concerning our understanding of the receptors and ligands composing the cannabinoid system. Likewise, the science surrounding cytokine biology has advanced enabling us to measure these proteins more precisely as well as understand and interpret the meaning of changes in their levels. Scientists wishing to study the health consequences of smoking marijuana as well as understand the possible role of endogenous cannabimimetic ligands in immune regulation have continued to study the influence of these substances on the regulation and development of the cytokine network. Research has shown that two major cannabinoid receptor subtypes exist and that subtype 1 (CB1) is expressed primarily in the brain whereas subtype 2 (CB2) is expressed primarily in the periphery. A variety of ligands for these receptors based on the cannabinoid structure have been synthesized and studied as well as low affinity compounds, noncannabinoid ligands, and endogenous ligands derived from fatty acid eicosanoids. Highly selective receptor antagonists have also been introduced and studied. Synthetic, low affinity ligands such as (+)‐HU‐211 and DMH‐11C have been shown to cause anti‐inflammatory effects possibly through inhibiting the production and action of TNF‐α and other acute phase cytokines. In addition, suppression of TNF and other cytokines such as GM‐CSF, IL‐6, IFNγ, and IL‐12 has also been seen following exposure to high affinity and psychoactive ligands such as marijuana and THC. However, some of these ligands have also been shown to increase rather than decrease interleukins such as IL‐1, IL‐4, IL‐10, and IL‐6, cytokines such as TNF‐α, and chemokines such as IL‐8, MIP‐1, and RANTES. The endogenous ligand, anandamide, has been shown in culture to either suppress the proliferation response to prolactin or enhance the response to cytokines such as IL‐3 and IL‐6. This eicosanoid has also been shown to increase the production of interleukins and other cytokines. Cannabinoid receptors have been shown to be involved in some but not all of these effects. It is clear that psychoactive and nonpsychoactive compounds have demonstrated effectsin vivoandin vitroon the production and function of a variety of cytokines. Depending upon the model system, these effects are often conflicting, and the involvement of cannabinoid receptors is unclear. However, enough evidence exists to suggest that the cannabinoid system significantly impacts the functioning of the cytokine network, and this association may provide clues to the mechanisms of certain immune diseases and form the basis for new immunotherapies.
Ion Transport in an Immortalized Rat Submandibular Cell Line SMG‐C6
Authors: Castro Robert , Barlow‐Walden Lornell , Woodson Trudi , Kerecman Jay D. , Zhang Guo H. , Martinez J. Ricardo
Published: August 2010

Abstract. The immortalized rat submandibular epithelial cell line, SMG‐C6, cultured on porous tissue culture supports, forms polarized, tight‐junction epithelia facilitating bioelectric characterization in Ussing chambers. The SMG‐C6 epithelia generated transepithelial resistances of 956±84Ω.cm2 and potential differences (PD) of −16.9 ± 1.5mV (apical surface negative) with a basal short‐circuit current (Isc) of 23.9 ± 1.7 μA/cm2 (n = 69). P2 nucleotide receptor agonists, ATP or UTP, applied apically or basolaterally induced a transient increase in Isc, followed by a sustained decreased below baseline value. The peak ΔIsc increase was partly sensitive to Cl− and K+ channel inhibitors, DPC, glibenclamide, and tetraethylammonium (TEA) and was completely abolished following Ca2+ chelation with BAPTA or bilateral substitution of gluconate for Cl−. The major component of basal Isc was sensitive to apical Na+ replacement or amiloride (half‐maximal inhibitory concentration 392 nM). Following pretreatment with amiloride, ATP induced a significantly greater Isc; however, the poststimulatory decline was abolished, suggesting an ATP‐induced inhibition of amiloride‐sensitive Na+ transport. Consistent with the ion transport properties found in Ussing chambers, SMG‐C6 cells express the rat epithelial Na+ channel α‐subunit (α‐rENaC). Thus, cultured SMG‐C6 cells produce tight polarized epithelia on permeable support with stimulated Cl− secretory conductance and an inward Isc accounted for by amiloride‐sensitive Na+ absorption.
Hormonal Regulation of Sodium/Sulfate Co‐Transport in Renal Epithelial Cells
Authors: Lee Hwa Jeong , Sagawa Kazuko , Shi Wei , Murer Heini , Morris Marilyn E.
Published: August 2010

Abstract.Serum sulfate concentrations are elevated in infants, young children, and pregnant women due, at least in part, to increased renal sulfate reabsorption. Little is known about the effects of hormones, particularly those involved in growth, development, and pregnancy, on renal sulfate reabsorption. The objective of this investigation was to examine the effects of growth hormone (GH), insulin‐like growth factor 1 (IGF‐1), progesterone (PG), and 17β‐estradiol (EST) on renal sodium/sulfate co‐transport.35S‐sulfate uptake was determined in Madin‐Darby canine kidney (MDCK)/NaSi‐1 cells (MDCK cells that have been stably transfected with rat sodium/sulfate co‐transporter (NaSi‐1) cDNA) and in opossum kidney (OK) cells. NaSi‐1 mRNA was determined by RT‐PCR and protein levels by ELISA. GH (0.1 nM) significantly increased the sodium/sulfate co‐transport in MDCK/NaSi‐1 cells up to 35%. IGF‐1 induced a concentration‐related stimulation of the sodium/sulfate co‐transport with a maximal response observed at 1000 nM(59% increase). Sodium‐dependent sulfate uptake was significantly increased when cells were preincubated with 10 nMPG, 10 nMEST, or 10 nMPG/10 nMEST up to 41%, 46%, or 39%, respectively. OK cells exhibited endogenous sodium‐dependent sulfate transport; significantly increased sodium/sulfate co‐transport was also observed in OK cells that were preincubated with GH, IGF‐1, and PG/EST, although not with EST alone. The NaSi‐1 mRNA and NaSi‐1 protein levels were significantly increased in MDCK/NaSi‐1 cells treated with 0.1 nMGH, 100 nMIGF‐1, 10 nMPG, and/or 10 nMEST compared with control. These results suggest that the increased renal sulfate reabsorption that occurs in neonates, young and pregnant humans, and animals could be mediated by the increased steady‐state levels of NaSi‐1 mRNA produced by the higher plasma concentrations of GH, IGF‐1, or PG/EST.
Promotive Effects of a Silk Film on Epidermal Recovery from Full‐Thickness Skin Wounds
Authors: Sugihara Akira , Sugiura Kikuya , Morita Haruo , Ninagawa Takashi , Tubouchi Kouzou , Tobe Ryuukichi , Izumiya Mamoru , Horio Takeshi , Abraham Nader G. , Ikehara Susumu
Published: August 2010

Abstract. We examined the effects of the transparent fibroin film (silk film) on full‐thickness skin wounds. Full‐thickness dermatotomies (15 mm × 9 mm) were prepared on the dorsal wall of CRJ:CD‐1 nu/nu (ICR nu/nu) mice. The area of the wounds dressed with silk film was reduced to 10% of that made by the dermatotomy 14 days after the dermatotomy and were covered with regenerated epidermis 21 days after the dermatotomy. In contrast, less recovery and epidermal regeneration were found 14 days after dermatotomy in the wounds dressed with a conventional hydrocolloid dressing (Duro Active). Furthermore, only partial incomplete epidemal growth was obtained 21 days after dermatotomy. Most importantly, the healing time of wounds dressed with silk film was 7 days shorter than those dressed with DuoActive dressing. The silk film showed an almost similar or slightly better promotive effect as the lyophilized porcine dermis (Alloask D), which is used as a dressing for burns, ulcers, and decubitis. Histologic findings revealed that there was greater collagen regeneration and less inflammation and neutrophil‐lymphocyte infiltration of the wounds dressed with silk film than with DuoActive dressing. It is clear that regeneration of the epidermis and dermis of the wound beds covered with silk film was faster than with DuoActive dressing. Finally, silk film is easily obtainable, sterilizable, and transparent, and it allows easy observation of tissue recovery. Therefore, silk film offers advantages over other dressings and may be clinically useful for wound treatment.
Acetaldehyde‐Stimulated PKC Activity in Airway Epithelial Cells Treated with Smoke Extract from Normal and Smokeless Cigarettes
Authors: Wyatt T. A. , Schmidt S. C. , Rennard S. I. , Tuma D. J. , Sisson J. H.
Published: August 2010

Abstract. Previously, we have found that acetaldehyde, a volatile component of cigarette smoke, stimulates the protein kinase C (PKC) pathway and inhibits ciliary motility. A “smokeless” cigarette (Eclipse) now exists in which most of the tobacco is not burned, reducing the pyrolyzed components in the extract. We hypothesized that acetaldehyde is a component of cigarette smoke that activates PKC in the airway epithelial cell, and therefore the Eclipse cigarette would not activate epithelial cell PKC. In this study, bovine bronchial epithelial cells (BBEC) were incubated with cigarette smoke extract (CSE) or Eclipse smoke extract (ESE). We found that PKC activity was significantly higher in cells exposed to 5% CSE than cells exposed to 5% ESE or media. When acetaldehyde levels of both extracts were measured by gas chromatography, CSE was found to have 15–20 times greater concentration (μM) of acetaldehyde than ESE. When BBEC were treated with 5% CSE, ciliary beating was further decreased from baseline levels. This decrease in ciliary beating was not observed in cells treated with ESE, suggesting that acetaldehyde contained in CSE slows cilia. These results suggest that volatile components such as acetaldehyde in cigarette smoke may inhibit ciliary motility via a PKC‐dependent mechanism.
Role of Sertoli Cells in Injury‐Associated Testicular Germ Cell Apoptosis
Authors: Boekelheide Kim , Fleming Shawna L. , Johnson Kamin J. , Patel Sutchin R. , Schoenfeld Heidi A.
Published: August 2010

Abstract. This review examines experimental models of Sertoli cell injury resulting in germ cell apoptosis. Since germ cells exist in an environment created by Sertoli cells, paracrine signaling between these intimately associated cells must regulate the process of germ cell death. Germ cell apoptosis may be signaled by a decrease in Sertoli cell pro‐survival factors, an increase in Sertoli cell pro‐apoptotic factors, or both. The different models of Sertoli cell injury indicate that spermatogenesis is susceptible to disruption, and that targeting critical Sertoli cell functions can lead to rapid and massive germ cell death.
Sugar Transport Regulation: Comparative Characterization of the Effect of NADH CoQ Reductase Deficiency in Two Cell Culture Systems
Authors: Germinario Ralph J. , Continelli Lia , Pratt Sue
Published: August 2010

Abstract. In this report, we have characterized the upregulation of glucose transport in two different respiration‐deficient fibroblast cell cultures. We have demonstrated that glucose transport increases in respiration‐deficient cells as measured by 2 deoxy D‐glucose transport and is readily observed in both the WG750 human and G14 Chinese hamster fibroblast respiration‐deficient cell lines when compared with the MCH55 normal human and V79 parental Chinese hamster cell lines, respectively. Using subcellular fractionation techniques, the GLUT 1 glucose transporter was found located predominantly in the plasma membrane–enriched fraction of the human and hamster cell lines. In human cells, the expression of the GLUT 1 glucose transporter was elevated three‐fold in the plasma membrane–enriched fraction of the WG750 respiration‐deficient mutant cells. In the Chinese hamster cell lines, the respiration‐deficient G14 cells exhibited no such GLUT 1 glucose transporter elevation in the plasma membrane–enriched fraction, yet expressed a >2‐fold increase in glucose transport. Furthermore, the G14 cells had a similar content of GLUT 1 glucose transporter in the plasma membrane fraction when compared with the V79 parental cell line. Using Western blot analysis, the GLUT 1 glucose transporter in G14 cells exhibited a different mobility on a polyacrylamide gel when compared with the mobility of the GLUT 1 glucose transporter of the V79 cell line. This differential mobility of the glucose transporters in the hamster cells appeared to be related to glycosylation differences of the glucose transporters. Although normal human and hamster cell lines exhibited significant increases in insulin‐stimulated sugar transport (P < 0.05), the two respective respiration‐deficient cell lines exhibited no significant increases in insulin‐stimulated sugar transport (P > 0.05). Additionally, the expression of the GLUT 1 mRNA in the human WG750 mutant cells was elevated when compared with GLUT 1 mRNA in normal cells. Insulin exposure significantly increased GLUT 1 mRNA in human cells (P < 0.05). No differences in the GLUT 1 mRNA were observed between both hamster cell lines. Thus, both respiration‐deficient cell lines are insulin resistant (i.e., regarding their insulin‐stimulated sugar transport). The respiration‐deficient mutation results in an increased sugar transport in the human and hamster cells; however, the human cells adapt to the mutation by increasing their levels of GLUT 1 mRNA and eventually membrane‐located glucose transporters. On the other hand, the hamster cells adapt by apparently modifying their glucose transporters' intrinsic activity via glycosylation. We feel that these cell systems can be effective models to study the multiple factors involved in sugar transport regulation in vertebrate cells.
Characterization of the Calcium Signaling System in the Submandibular Cell Line SMG‐C6
Authors: Liu Xiao‐Bing , Sun Xiuhua , Mörk Ann‐Christin , Dodds Michael W. J. , Martinez J. Ricardo , Zhang Guo H.
Published: August 2010

Abstract.Establishment of salivary cell lines retaining normal morphological and physiological characteristics is important in the investigation of salivary cell function. A submandibular gland cell line, SMG‐C6, has recently been established. In the present study, we characterized the phosphoinositide (PI)‐Ca2+signaling system in this cell line. Inositol 1,4,5‐trisphosphate(1,4,5‐IP3) formation, as well as Ca2+storage, release, and influx in response to muscarinic, α1‐adrenergic, P2Y‐nucleotide, and cytokine receptor agonists were determined. Ca2+release from intracellular stores was strongly stimulated by acetylcholine (ACh) and ATP, but not by norepinephrine (NA), epidermal growth factor (EGF), interleukin‐6 (IL‐6), and tumor necrosis factor‐α (TNFα). Consistently, 1,4,5‐IP3formation was dramatically stimulated by ACh and ATP. ACh‐stimulated cytosolic free Ca2+concentration [Ca2+]iincrease was inhibited by ryanodine, suggesting that the Ca2+‐induced Ca2+release mechanism is involved in the ACh‐elicited Ca2+release process. Furthermore, ACh and ATP partially discharged the IP3‐sensitive Ca2+store, and a subsequent exposure to thapsigargin (TG) induced further [Ca2+]iincrease. However, exposure to TG depleted the store and a subsequent stimulation with ACh or ATP did not induce further [Ca2+]iincrease, suggesting that ACh and ATP discharge the same storage site sensitive to TG. As in freshly isolated submandibular acinar cells, exposure to ionomycin and monensin following ACh or TG induced further [Ca2+]iincrease, suggesting that IP3‐insensitive stores exist in SMG‐C6 cells. Ca2+influx was activated by ACh, ATP, or TG, and was significantly inhibited by La3+, suggesting the involvement of store‐operated Ca2+entry (SOCE) pathway. These results indicate that in SMG‐C6 cells: (i) Ca2+release is triggered by muscarinic and P2Y‐nucleotide receptor agonists through formation of IP3; (ii) both the IP3‐sensitive and ‐insensitive Ca2+stores are present; and (iii) Ca2+influx is mediated by the store‐operated Ca2+entry pathway. We conclude that Ca2+regulation in SMG‐C6 cells is similar to that in freshly isolated SMG acinar cells; therefore, this cell line represents an excellent SMG cell model in terms of intracellular Ca2+signaling.
Molecular paleoecology: using gene regulatory analysis to address the origins of complex life cycles in the late Precambrian
Authors: Dunn Ewan F. , Moy Vanessa N. , Angerer Lynne M. , Angerer Robert C. , Morris Robert L. , Peterson Kevin J.
Published: August 2010

SUMMARY
Molecular paleoecology is the application of molecular data to test hypotheses made by paleoecological scenarios. Here, we use gene regulatory analysis to test between two competing paleoecological scenarios put forth to explain the evolution of complex life cycles. The first posits that early bilaterians were holobenthic, and the evolution of macrophagous grazing drove the exploitation of the pelagos by metazoan eggs and embryos, and eventually larvae. The alternative hypothesis predicts that early bilaterians were holopelagic, and new adult stages were added on when these holopelagic forms began to feed on the benthos. The former hypothesis predicts that the larvae of protostomes and deuterostomes are not homologous, with the implication that larval‐specific structures, including the apical organ, are the products of convergent evolution, whereas the latter hypothesis predicts homology of larvae, specifically homology of the apical organ. We show that in the sea urchin, Strongylocentrotus purpuratus, the transcription factors NK2.1 and HNF6 are necessary for the correct spatial expression profiles of five different cilia genes. All of these genes are expressed exclusively in the apical plate after the mesenchyme‐blastula stage in cells that also express NK2.1 and HNF6. In addition, abrogation of SpNK2.1 results in embryos that lack the apical tuft. However, in the red abalone, Haliotis rufescens, NK2.1 and HNF6 are not expressed in any cells that also express these same five cilia genes. Nonetheless, like the sea urchin, the gastropod expresses both NK2.1 and FoxA around the stomodeum and foregut, and FoxA around the proctodeum. As we detected no similarity in the development of the apical tuft between the sea urchin and the abalone, these molecular data are consistent with the hypothesis that the evolution of mobile, macrophagous metazoans drove the evolution of complex life cycles multiple times independently in the late Precambrian.
Delayed onset of midline netrin expression in Artemia franciscana coincides with commissural axon growth and provides evidence for homology of midline cells in distantly related arthropods
Authors: Duman‐Scheel Molly , Clark Stephanie M. , Grunow Eric T. , Hasley Andrew O. , Hill Brandon L. , Simanton Wendy L.
Published: August 2010

SUMMARY
Although many similarities in arthropod central nervous systems (CNS) development exist, differences in midline cell formation and ventral nerve cord axonogenesis have been noted in arthropods. It is possible that changes in the expression of axon guidance molecules such as Netrin, which functions during commissural axon guidance in Drosophila and many other organisms, may parallel these differences. In this investigation, we analyze this hypothesis by examining Netrin accumulation during development of the brine shrimp Artemia franciscana, a branchiopod crustacean. An Artemia franciscana netrin (afrnet) orthologue was cloned. An antibody to the afrNet protein was generated and used to examine the pattern of afrNet accumulation during Artemia development. Despite differences between Drosophila and Artemia nerve cord development, examination of afrNet accumulation suggests that this protein functions to regulate commissure formation during Artemia CNS development. However, detection of afrNet at the midline and on commissural axons occurs at a relatively later time point in Artemia as compared with Drosophila. Detection of afrNet in a subset of midline cells that closely resemble Netrin‐expressing cells at the Drosophila midline provides evidence for homology of midline cells in arthropods. Expression of Netrins in many other tissues is comparable, suggesting that Netrin proteins may play many conserved roles during arthropod development.