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Biomedical subjects

S Sebastian

Publications and source records attributed to S Sebastian.

At least 37 records · Page 2Linked to original sources
PubMed1999

Characterization of the rat type III hexokinase gene promoter. A functional octamer 1 motif is critical for basal promoter activity.

A 1532-base pair 5'-flanking region of the gene encoding rat type III hexokinase has been cloned and sequenced. The total sequence includes positions -1548 to -17 (A of the translational start ATG as position +1). Using luciferase reporter constructs transfected into PC12 (rat pheochromocytoma) and L2 (rat lung) cells, basal promoter activity has been associated with sequence between -182 and -89. This includes a single transcriptional start site, an adenine at position -134 identified by primer extension. Together with previously cloned cDNA sequence, this accounts for an mRNA of approximately 3.9 kilobases, found by Northern blotting of RNA from rat lung and kidney. Sequence upstream of the transcriptional start site was devoid of canonical TATA and CAAT elements. An octamer 1 (Oct-1) binding site, located between positions -166 and -159 was shown by deletion analysis and site-directed mutation to be critical for promoter activity. Nuclear extracts from PC12 cells contained a protein (or proteins) specifically binding the octamer sequence, and supershift experiments with anti-Oct-1 indicated involvement of this ubiquitously expressed transcription factor in the complex. Sequence including the Oct-1 site and immediately adjacent regions was protected from DNase I digestion in footprinting experiments with nuclear extracts from PC12 cells. Reverse transcription polymerase chain reaction indicated that levels of type III hexokinase mRNA in rat tissues increased in the order brain < liver < lung approximately kidney; immunoblotting indicated that type III hexokinase protein in these tissues increased in a similar manner.

Animals↗
PubMed1999

Allosteric regulation of type I hexokinase: A site-directed mutational study indicating location of the functional glucose 6-phosphate binding site in the N-terminal half of the enzyme.

The Type I isozyme of mammalian hexokinase has evolved by a gene duplication-fusion mechanism, with resulting internal duplication of sequence and ligand binding sites. However, 1:1 binding stoichiometry indicates that only one of these is available for binding the product inhibitor, Glc-6-P; the location of that site, in the N- or C-terminal half, remains under debate. Recent structural studies (Aleshin et al., Structure 6, 39-50, 1998; Mulichak et al., Nature Struct. Biol. 5, 555-560, 1998) implicated Asp 84 or its analog in the C-terminal half, Asp 532, in binding of Glc-6-P. Zeng et al. (Biochemistry 35, 13157-13164, 1996) demonstrated that mutation of Asp 532 to Lys or Glu did not affect inhibition by the Glc-6-P analog, 1,5-anhydroglucitol-6-P. These same mutations, as well as mutation to Ala, at the Asp 84 position are now shown to result in increased Ki for 1,5-anhydroglucitol-6-P. The ability of Pi to antagonize inhibition by the Glc-6-P analog is severely diminished or abolished by these mutations, suggesting that antagonism is dependent on precise positioning of the inhibitory hexose 6-phosphate. The structure of the enzyme complexed with Glc and Pi has been determined, and shows that Pi occupies the same site as the 6-phosphate group in the complex with Glc-6-P. Thus, antagonism between these ligands results from competition for a common anion binding site in the N-terminal half.

Adenosine Triphosphate↗
PubMed1999

Stimulation of brain hexokinase gene expression by recombinant brain insulin-like growth factor in C6 glial cells.

Glycolysis is essential for cerebral energy generation. Hence, expression and regulation of gene-encoding brain hexokinase (HK I), the exclusive brain glucose phosphorylating enzyme, can be a critical step in this process. The present study demonstrates the ability of recombinant brain insulin-like growth factor (BIGF, a closely related member of insulin superfamily) to stimulate HK I gene expression in a concentration- and time-dependent manner in C6 glial cells. BIGF treatment (10 ng/ml) on quiescent C6 glial cells stimulates transcription and translation of HK I RNA to approximately 2.5-fold within 4 h after the addition of growth factor. In contrast, insulin or epidermal growth factor could not mimic this effect. Coincubation of cycloheximide with BIGF abolished this stimulatory effect, indicating a requirement for prior protein synthesis for this effect. These results suggest that IGF may have a role in regulating hexokinase gene expression in brain and possibly of brain glucose metabolism.

Animals↗
PubMed1999

Subclinical hyperthyroidism and hyperkinetic behavior in children.

The authors report three children who exhibited developmental learning disabilities (DLDs) associated with behavioral disturbances, such as attention deficit, hyperactivity, and autistic features. The thyroid function tests performed as a part of routine endocrinologic evaluation of children with DLDs revealed a hormonal profile consistent with hyperthyroidism. These children had no systemic signs of hyperthyroidism. Treatment with neomercazole resulted in good control of their hyperkinetic behavior and subsequent improvement in language function attributable to an increased attention span, thereby facilitating speech therapy. Although routine screening of all children with DLDs for thyroid dysfunction may not be cost-effective, selective screening of children with familial attention-deficit hyperactivity disorder and those with attention-deficit and hyperactivity in association with DLDs and pervasive developmental disorders appears to be justified.

Antithyroid Agents↗
PubMed1999

FbpC is not essential for iron acquisition in Neisseria gonorrhoeae.

The fbpABC locus of Neisseria gonorrhoeae has been proposed to encode a periplasmic protein-dependent iron transport system. Although the function of the gonococcal FbpA protein has been well characterized and its role as a periplasmic binding protein is well defined, little is known about the function of the FbpB and FbpC proteins. To define the function of the gonococcal FbpC protein, an N. gonorrhoeae F62 fbpC mutant was constructed by insertional inactivation with the kanamycin gene. The N. gonorrhoeae F62 fbpC mutant was observed to grow with heme, transferrin, or ferric nitrate as the sole exogenous iron source, indicating that the gonococcal FbpC protein is not absolutely required for growth with these iron sources. In previous studies we were unable to detect fbpB- or fbpC-specific transcripts by Northern analysis. Reverse transcription-PCR analysis with RNA obtained from N. gonorrhoeae F62 grown under iron-replete and -depleted conditions detected fbpA and fbpAB transcripts but failed to detect fbpC or fbpBC transcripts. These results indicate that FbpC does not play a pivotal role in iron transport in N. gonorrhoeae and suggest that additional ABC transport systems are functional in the gonococcus for the acquisition of iron.

Bacterial Proteins↗
PubMed1998

Expression of two type II-like tumor hexokinase RNA transcripts in cancer cell lines.

To maintain an elevated glycolytic rate, cancerous or proliferating cells alter the expression pattern of rate limiting glycolytic enzymes. Since glucose phosphorylation is the first step in glycolysis, hexokinase (HK), the first rate limiting glycolytic enzyme, can play a key regulatory role in this process. A low-Km, mitochondrial type II-like tumor HK is described as the predominant form in hepatomas. However, recent identification of a high-Km glucose phosphorylating activity in a range of cancer cells prompted us to characterize glucose phosphorylating enzymes of cancer cells at the molecular level. Highly sensitive reverse-transcription polymerase chain reaction identifies an induction and overexpression of a type II-like tumor HK RNA in a range of cancer cell lines irrespective of tissue origin. In addition, we report here the identification of two RNA transcripts of type II-like tumor HK of approximately 5.5 and approximately 4.0 kb in these cancer cells lines, including muscle-derived L6 myoblast cells. Interestingly, under normal conditions muscle cells express only a approximately 5.5-kb type II HK RNA transcript. A significant amount of type I HK RNA was also found expressed in cancer cell lines. RNA encoding glucokinase (GK), the high-Km HK isozyme, was found only in cancer cells originating from liver and pancreas, which express GK under normal conditions.

Animals↗
PubMed1997

Insulin-like growth factor I induces tumor hexokinase RNA expression in cancer cells.

Increased glycolysis is a characteristic of cancer cells. Though less efficient in energy production, it ensures continuous supply of energy and phosphometabolites for biosynthesis enabling metastatic and less vascularized cancer cells to proliferate even under hypoxic conditions. Since hexokinase is the first rate limiting enzyme in the glycolytic pathway, elevated levels of Type II like hexokinase in tumors are of great significance in this context. Under normal conditions insulin regulates expression of hexokinase Type II isoenzyme, which is predominantly expressed in muscle. On the other hand cancer cells overexpress insulin-like growth factors and their receptors which mimic many activities of insulin. This prompted us to examine a hypothesis that insulin-like growth factors may be responsible for overexpression of tumor hexokinase. Our experiments demonstrate that insulin-like growth factor I indeed induces hexokinase gene expression in a concentration and time dependent manner in two cancer cell lines we studied.

Blotting, Northern↗
PubMed1997

Chromosomes with high gene density are preferentially repaired in human cells.

It is known that DNA repair is heterogeneous in human cells since open chromatin, active genes and their transcribed strands are preferentially repaired. It is thus expected that DNA repair is clustered in chromosomes with high gene density. We have employed a DNA repair inhibitor, cytosine arabinoside (Ara-C), to convert ethyl methane sulfonate (EMS)-induced excision repairable lesions to chromosomal breaks, to check for the existence of heterogeneity of repair at the chromosome level. Chromosome staining by fluorescence in situ hybridization (FISH) was used to analyze breakage in chromosomes with diverse gene densities. These chromosomes were identified by means of the CpG island distribution after FISH with a CpG island-rich probe isolated from total human genomic DNA. Thus, three chromosomes with very high gene density (numbers 1, 19 and 20) were compared with two chromosomes with very low gene density (numbers 4 and 18) for clastogenicity and sensitivity to co-treatment with Ara-C and EMS. Our data indicate that those chromosome with higher gene density are more sensitive to a combination treatment with Ara-C and EMS, indicating that the level of excision repair synthesis is higher in those chromosome. It is therefore concluded that DNA excision repair is preferentially directed to chromosomes with high gene density. The implications of this finding in human biomonitoring using FISH techniques are discussed.

Antimetabolites, Antineoplastic↗
PubMed1997

Apparent digestibility of protein and amino acids in broiler chickens fed a corn-soybean diet supplemented with microbial phytase.

The effect of microbial phytase supplementation on CP and amino acid (AA) digestibility was investigated in a 28-d trial using 360 sexed, day-old broiler chickens fed corn-soybean meal diets. The experimental design was a completely randomized one with a 3 x 2 factorial arrangement of treatments. The variables included P and Ca levels and phytase: P and Ca levels were: normal P-normal Ca [0.45% available P (Pa), 1.0% Ca], low P-normal Ca (0.35% Pa, 1.0% Ca), and low P-low Ca (0.35 Pa and 0.6% Ca); and phytase at 0 and 600 U/kg diet. Phytase supplementation increased body weight gain (P < 0.014) and feed intake (P < 0.004) at 19 d in male chickens; in females, phytase increased (P < 0.012) only body weight gain at 19 d. The low P-normal Ca diet reduced (P < 0.05) feed intake and body weight gain in both sexes at 7, 14, and 19 d, compared to the normal P-normal Ca diet; the reduction of Ca in the low P diet prevented the above depression, resulting in body weight gain and feed intake to a level comparable to that of the normal P-normal Ca diet. Microbial phytase supplementation had no effect (P < 0.065) on the apparent ileal digestibility (AID) of CP or any AA except Met and Phe in male broiler chickens. In females, adding phytase increased the AID of all AA except Lys, Met, Phe, and Pro. The low P-normal Ca diet reduced (P < 0.05) the AID of Phe, Asp, and Ser in male chickens and reduced the AID of all the AA except Met and Pro in females compared to the normal P-normal Ca diet. The reduction of Ca in the low P diet prevented the depression of the AID of the AA caused by the low P-normal Ca diet, resulting in AID of AA having a level comparable to that of the normal P-normal Ca diet in both sexes. Phytase supplementation did not have any effect (P > 0.05) on apparent "fecal" digestibility (AFD) of CP or any of the AA in male chickens; however, in female chickens it increased the AFD of Thr, Asp, Glu, and Ser. In summary, phytase supplementation increased growth performance in both sexes; increased AID and AFD of most of the AA, particularly in female chickens. The optimum growth performance and AA digestibilities were obtained with the lowest input of resources, in the low P-low Ca diet supplemented with microbial phytase.

6-Phytase↗
PubMed1996

Comparative assessment of bacterial inoculation and propionic acid treatment of aerobic stability and microbial populations of ensiled high-moisture ear corn.

High-moisture ear corn (HMEC) was untreated, treated with propionic acid (PA), or inoculated with a mixture of Lactobacillus plantarum and Enterococcus faecium and allowed to ensile in laboratory silos for 0, 7, 21, 42, 138, or 202 d. The silages were evaluated for fermentation quality, microbial populations, and aerobic stability. In all treatments, silage pH declined rapidly within 7 d, but the rate of decline seemed greatest with the inoculum. The lactic acid content of inoculated HMEC was higher (P < .05) than that of control of PA-treated HMEC. Regardless of treatment, the population of lactic acid bacteria (LAB) increased (P < .1) up to 7 to 21 d of fermentation then declined; LAB counts decreased (P < .05) up to 42 d in control and PA-treated silage but continued to decline until 138 d for inoculated silage. Yeast and mold counts tended to decrease up to 42 d of ensiling then decreased (P < .05) as fermentation progressed. Between 138 and 202 d of ensiling, the control silage showed a marked increase (P < .10) in pH and yeast and mold populations, providing evidence of secondary fermentation; PA treatment and bacterial inoculation prevented secondary fermentation. Inoculation tended to reduce estimates of sample temperature for silage stored for 138 d and exposed to air, but not for the corresponding silage stored for 202 d. Treatment with PA prevented the loss (P > .05) of acetic acid and the rise (P > .05) in pH during air exposure of the 138-d silage; both control and PA-treated silage showed an increase (P < .05) in yeast and mold populations, but the increments were 38% and 23%, respectively. Compared with PA, the relative efficacy of inoculation in improving aerobic spoilage of HMEC depended on the period of silo storage and the criterion used to assess aerobic stability.

Acetates↗
PubMed1996

The effects of supplemental microbial phytase on the performance and utilization of dietary calcium, phosphorus, copper, and zinc in broiler chickens fed corn-soybean diets.

A 3-wk feeding trial with 180 sexed day-old broiler chickens was conducted to study the efficacy of microbial phytase (Natuphos 1000) on growth performance, relative retention of P, Ca, Cu, and Zn, and mineral contents of plasma and bone. Treatments involved a normal P level corn-soybean diet, a low-P diet, and a low-P plus phytase (600 phytase units/kg) diet. Phytase supplementation increased (P < or = 0.05) body weight in male and female chickens by 13.2 and 5.8%, respectively, at 21 d. The improvements yielded body weights comparable to those obtained on the normal P diet. Phytase supplementation overcame (P < or = 0.05) the depression of feed intake observed on the low-P diet. Treatments had no effect on feed:gain ratio. Phytase supplementation of the low-P diet increased (P < or = 0.05) the relative retention of total P, Ca, Cu, and Zn by 12.5, 12.2, 19.3, and 62.3 percentage units, respectively, in male chickens. Microbial phytase increased the plasma P by 15.7% and reduced (P < or = 0.05) the Ca concentration by 34.1%, but had no effect on plasma concentrations of Cu or Zn. Phytase supplementation increased the percentage ash in both head and shaft portions of dry, fat-free tibia bone to a level comparable to that of the normal-P diet. Phytase supplementation had no effect on the concentration of any of the minerals measured in whole tibia ash but did increase (P < or = 0.05) the DM percentage of P and Ca min tibia head of male chickens by 0.65 and 1.4 percentage units, respectively. These results show that microbial phytase supplementation of a low-P diet increased growth and relative retention of total P, Ca, Cu, and Zn and improved bone mineralization in broiler chickens.

6-Phytase↗
PubMed1996

Efficacy of supplemental microbial phytase at different dietary calcium levels on growth performance and mineral utilization of broiler chickens.

A 3-wk feeding trial with 240 sexed, day-old broiler chickens was conducted to determine the efficacy of microbial phytase at different levels of dietary Ca on performance and utilization of minerals in broiler chickens fed a low-P corn-soybean diet. The experimental design was a 3 x 2 factorial arrangement of treatments; Ca at 0.6, 1.0, and 1.25% and phytase at 0 and 600 phytase U/kg diet. Phytase supplementation, regardless of Ca level, increased (P < or = 0.005) feed intake, (P < or = 0.0001) body weight, and (P < or = 0.025) feed efficiency at 21 d; the optimum levels of body weight, feed intake, and feed efficiency were obtained with low (0.6%) dietary Ca plus phytase. Retentions of P, Ca, and N were increased (P < or = 0.05) by phytase supplementation. Although maximum retentions of P and N were obtained at the 1.0 and 1.25% Ca levels, respectively, they were not significantly different from the values obtained at 0.6% Ca. The increasing level of dietary Ca decreased plasma P ( P < or = 0.05) and Cu (P < or = 0.06). Phytase supplementation had the opposite effect; i.e., increased plasma P (P < or = 0.03) and Cu (P < or = 0.02). The maximum level of plasma P was obtained with phytase at the 1.0% Ca level, but this value was not significantly different from the value obtained with phytase at the 0.6% Ca level. Phytase supplementation increased (P < 0.04) the ash content of both tibia head and shaft but had no effect on mineral contents in the ash. The optimum level of ash content was observed with the 0.06% Ca diet plus phytase. The results show that microbial phytase supplementation to a low P diet improved growth performance and mineral utilization in broiler chickens. Dietary Ca levels had a significant effect on the response to phytase; the optimum growth performance and mineral utilization were achieved at the low (0.6%) level of dietary Ca supplemented with phytase.

6-Phytase↗
PubMed1992

Special wounds. Nail bed, plantar puncture, and cartilage.

Traumatic wounds are one of the most common problems encountered in the practice of emergency medicine. The literature is replete with articles concerning the evaluation and care of life threatening wounds, but many common non-life threatening injuries have received very little attention. This article focuses on nail bed injuries, plantar puncture wounds, and cartilaginous wounds. Although not immediately life threatening, these injuries may be associated with significant long-term morbidity and, rarely, life threatening sequelae.

Cartilage↗
PubMed1990

Fatal pulmonary edema due to nitric acid fume inhalation in three pulp-mill workers.

Three young men died of rapidly progressive pulmonary edema of delayed onset after inhalation of fumes from an accidental nitric acid explosion. Electron microscopy revealed altered neutrophils and necrotic endothelial cells in alveolar capillaries. Immunohistochemistry showed small and large serum proteins, including immunoglobulin M, in the edema fluid and hyaline membranes. Increased permeability is a consequence of direct microvascular injury by inhaled nitrogen dioxide. However, our findings, implicating neutrophils and serum-derived mediators in the pathogenesis of the pulmonary edema, are consistent with recent proposals on their roles in the maintenance and/or progression of edema initiated by toxic inhalations.

Accidents, Occupational↗