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Biomedical subjects

D Lin

Publications and source records attributed to D Lin.

At least 199 records · Page 11Linked to original sources
PubMed1991

Missense mutation serine106----proline causes 17 alpha-hydroxylase deficiency.

Steroid 17 alpha-hydroxylase deficiency is caused by defects in cytochrome P450c17, the single enzyme that has 17-alpha hydroxylase and 17,20-lyase activities. We describe a rapid and efficient polymerase chain reaction tactic for identifying these genetic lesions and identify Ser106----Pro as the cause of 17 alpha-hydroxylase deficiency in two unrelated homozygous patients from Guam. We used site-directed mutagenesis of the normal P450c17 cDNA to construct the Pro106 mutant, and expressed both the normal and mutant sequences in monkey COS-1 cells and in yeast. Expression of the normal sequence permitted the cells to convert pregnenolone to 17-OH pregnenolone, progesterone to 17-OH progesterone, and 17-OH pregnenolone to dehydroepiandrosterone, showing the normal sequence conferred both 17 alpha-hydroxylase and 17,20-lyase activities. Expression of the mutant sequence generated P450c17 mRNA, but conferred none of these activities, proving that the Ser106----Pro mutation abolished the 17 alpha-hydroxylase and 17,20-lyase activities. An HhaI restriction site created by the mutation should permit screening of large populations.

Adolescent↗
PubMed1991

Growth suppression induced by wild-type p53 protein is accompanied by selective down-regulation of proliferating-cell nuclear antigen expression.

The p53 gene is a frequent target of mutation in a wide variety of human cancers. Previously, it was reported that conditional expression of wild-type p53 protein in a cell line (GM47.23) derived from a human glioblastoma multiform tumor had a negative effect on cell proliferation. We have now investigated the effect that induction of wild-type p53 protein in this cell line has on the expression of the proliferating-cell nuclear antigen gene. The proliferating-cell nuclear antigen gene encodes a nuclear protein that is an auxiliary factor of DNA polymerase delta and part of the DNA replication machinery of the cell. We show that inhibition of cell cycle progression into S-phase after induction of wild-type p53 protein is accompanied by selective down-regulation of proliferating-cell nuclear antigen mRNA and protein expression.

Antigens, Neoplasm↗
PubMed1991

Normal genes for the cholesterol side chain cleavage enzyme, P450scc, in congenital lipoid adrenal hyperplasia.

Congenital lipoid adrenal hyperplasia is the most severe form of congenital adrenal hyperplasia. Affected individuals can synthesize no steroid hormones, and hence are all phenotypic females with a severe salt-losing syndrome that is fatal if not treated in early infancy. All previous studies have suggested that the disorder is in the cholesterol side chain cleavage enzyme (P450scc), which converts cholesterol to pregnenolone. A newborn patient was diagnosed by the lack of significant concentrations of adrenal or gonadal steroids either before or after stimulation with corticotropin (ACTH) or gonadotropin (hCG). The P450scc gene in this patient and in a previously described patient were grossly intact, as evidenced by Southern blotting patterns. Enzymatic (polymerase chain reaction) amplification and sequencing of the coding regions of their P450scc genes showed these were identical to the previously cloned human P450scc cDNA and gene sequences. Undetected compound heterozygosity was ruled out in the new patient by sequencing P450scc cDNA enzymatically amplified from gonadal RNA. Northern blots of gonadal RNA from this patient contained normal sized mRNAs for P450scc and also for adrenodoxin reductase, adrenodoxin, sterol carrier protein 2, endozepine, and GRP-78 (the precursor to steroidogenesis activator peptide). These studies show that lipoid CAH is not caused by lesions in the P450scc gene, and suggest that another unidentified factor is required for the conversion of cholesterol to pregnenolone, and is disordered in congenital lipoid adrenal hyperplasia.

Adrenal Hyperplasia, Congenital↗
PubMed1991

Patient controlled analgesia: drug options, infusion schedules, and other considerations.

Patient controlled analgesia (PCA) has a number of advantages compared with traditional methods of pain management. Some of these advantages include superior pain relief, less sedation due to superior drug titration, increased psychological satisfaction due to patient control of pain management, individualized analgesic dosing, decreased staff time for patient care, and increased patient activity and mobility. Although a few cases of respiratory depression have been reported with the use of PCA, there is a relatively low risk of this complication in most patient populations. Appropriate candidates for PCA include terminally ill-cancer patients, postoperative patients, mentally clear and alert trauma patients, and patients who require massive doses of oral narcotics to control pain but are experiencing intolerable side effects. This article focuses on the principles involved in selecting the optimal analgesic and the therapeutic variables involved in using PCA.

Analgesia, Patient-Controlled↗
PubMed1990

Cloning and sequence of the human adrenodoxin reductase gene.

Adrenodoxin reductase (ferrodoxin:NADP+ oxidoreductase, EC 1.18.1.2) is a flavoprotein mediating electron transport to all mitochondrial forms of cytochrome P450. We cloned the human adrenodoxin reductase gene and characterized it by restriction endonuclease mapping and DNA sequencing. The entire gene is approximately 12 kilobases long and consists of 12 exons. The first exon encodes the first 26 of the 32 amino acids of the signal peptide, and the second exon encodes the remainder of signal peptide and the apparent FAD binding site. The remaining 10 exons are clustered in a region of only 4.3 kilobases, separated from the first two exons by a large intron of about 5.6 kilobases. Two forms of human adrenodoxin reductase mRNA, differing by the presence or absence of 18 bases in the middle of the sequence, arise from alternate splicing at the 5' end of exon 7. This alternately spliced region is directly adjacent to the NADPH binding site, which is entirely contained in exon 6. The immediate 5' flanking region lacks TATA and CAAT boxes; however, this region is rich in G + C and contains six copies of the sequence GGGCGGG, resembling promoter sequences of "housekeeping" genes. RNase protection experiments show that transcription is initiated from multiple sites in the 5' flanking region, located about 21-91 base pairs upstream from the AUG translational initiation codon.

Amino Acid Sequence↗
PubMed1990

Weight cycling in female rats subjected to varying meal patterns.

The effect of weight cycling on energy balance was examined in female rats. Two groups of adult female rats were subjected to three bouts of weight cycling, each bout consisting of 8 days of food restriction (9 g/day or approximately 50% of usual intake) followed by 16 days of refeeding. During refeeding animals were given 22.8 g/day of food so that they were offered, during the 24-day cycle, the same amount of food offered to control rats that were not subjected to weight cycling. One group of weight-cycled rats (gorgers) was given its daily intake in a few large meals (i.e., allowed to gorge). The other weight-cycled group (nibblers) was fed by automated feeders in several small meals during each 24-h period (i.e., prevented from gorging). Neither weight-cycled group displayed an increased food efficiency or an increased body fatness compared with noncycled controls. Weight-cycled rats allowed to gorge did have an increased food efficiency and a greater carcass energy content compared with weight-cycled rats not allowed to gorge. These results suggest a pattern of gorging promoted food efficiency and body energy gain compared with a pattern of nibbling, but gorging during refeeding cannot account for reports of increased food efficiency in weight-cycled rats.

Adipose Tissue↗
PubMed1990

Effect of weight cycling on susceptibility to dietary obesity.

Weight cycling, defined as repeated episodes of weight loss followed by weight regain, has been suggested to make rats more energy efficient and produce a state of energy balance favoring accumulation of excess body fat. In addition, weight cycling may favor accumulation of fat in central vs. peripheral adipose depots. In the present study, we gave two groups of female Wistar rats ad libitum access to an obesity-producing high-fat diet (60% of calories from fat). Both groups had previously eaten a low-fat stock diet, but one group had been subjected to three bouts of weight cycling. Rats that were previously weight cycled gained less body weight and body fat when given the high-fat diet than did controls. The lower rate of weight gain was due to a lesser increase in food intake, since daily energy expenditure was significantly lower in previously cycled rats than in controls. In summary, weight cycling does not appear to predispose rats to becoming obese on a high-calorie diet and apparently produces some effect on food intake that reduces, at least in the short run, weight gain on the high-calorie diet.

Adipose Tissue↗
PubMed1990

Metabolic differences between obesity-prone and obesity-resistant rats.

We compared, across several physiological variables, rats most and least susceptible to develop obesity when given a high-fat diet. After 4 wk of eating a high-fat diet (60% of calories from fat), rats in the upper (obesity prone, OP) and lower (obesity resistant, OR) quartiles for weight gain were further studied. OP rats ate significantly more than OR rats, but this did not completely explain differences in their susceptibility to dietary obesity. No differences in 24-h energy expenditure were found between groups. OR rats had a significantly lower 24-h respiratory quotient, indicative of a greater relative proportion of fat oxidation and lower plasma levels of free fatty acids (FFA) than OP rats. Thus the ability to avoid dietary obesity produced by a high-fat diet may depend on an ability to increase fat oxidation in response to increased fat intake. Insulin sensitivity, measured by a euglycemic insulin clamp, was significantly higher in OR than OP rats. We cannot determine from these data whether insulin resistance developed as a consequence of elevated FFA levels or whether the ability to oxidize FFA declined as a result of development of insulin resistance. In summary, we propose that rats able to resist becoming obese on a high-fat diet have the ability to adjust the composition of fuel oxidized to the fuel composition of the diet with a minimum increase in body fat. The specific mechanisms by which this occurs are unknown but may be related to effects of diet on insulin sensitivity.

Animals↗
PubMed1990

Molecular biology of mammalian glucose transporters.

The oxidation of glucose represents a major source of metabolic energy for mammalian cells. However, because the plasma membrane is impermeable to polar molecules such as glucose, the cellular uptake of this important nutrient is accomplished by membrane-associated carrier proteins that bind and transfer it across the lipid bilayer. Two classes of glucose carriers have been described in mammalian cells: the Na(+)-glucose cotransporter and the facilitative glucose transporter. The Na(+)-glucose cotransporter transports glucose against its concentration gradient by coupling its uptake with the uptake of Na+ that is being transported down its concentration gradient. Facilitative glucose carriers accelerate the transport of glucose down its concentration gradient by facilitative diffusion, a form of passive transport. cDNAs have been isolated from human tissues encoding a Na(+)-glucose-cotransporter protein and five functional facilitative glucose-transporter isoforms. The Na(+)-glucose cotransporter is expressed by absorptive epithelial cells of the small intestine and is involved in the dietary uptake of glucose. The same or a related protein may be responsible for the reabsorption of glucose by the kidney. Facilitative glucose carriers are expressed by most if not all cells. The facilitative glucose-transporter isoforms have distinct tissue distributions and biochemical properties and contribute to the precise disposal of glucose under varying physiological conditions. The GLUT1 (erythrocyte) and GLUT3 (brain) facilitative glucose-transporter isoforms may be responsible for basal or constitutive glucose uptake. The GLUT2 (liver) isoform mediates the bidirectional transport of glucose by the hepatocyte and is responsible, at least in part, for the movement of glucose out of absorptive epithelial cells into the circulation in the small intestine and kidney. This isoform may also comprise part of the glucose-sensing mechanism of the insulin-producing beta-cell. The subcellular localization of the GLUT4 (muscle/fat) isoform changes in response to insulin, and this isoform is responsible for most of the insulin-stimulated uptake of glucose that occurs in muscle and adipose tissue. The GLUT5 (small intestine) facilitative glucose-transporter isoform is expressed at highest levels in the small intestine and may be involved in the transcellular transport of glucose by absorptive epithelial cells. The exon-intron organizations of the human GLUT1, GLUT2, and GLUT4 genes have been determined. In addition, the chromosomal locations of the genes encoding the Na(+)-dependent and facilitative glucose carriers have been determined. Restriction-fragment-length polymorphisms have also been identified at several of these loci.(ABSTRACT TRUNCATED AT 400 WORDS)

Amino Acid Sequence↗
PubMed1988

Gene expression in mineralizing chick epiphyseal cartilage.

To map transcriptional events associated with mineralization in developing long bones, we have established protocols for preparing RNA from regions of chick epiphyseal cartilage. Using these RNA preparations, we have probed for appearance of mRNA coding for type I, II, and X collagen, as well as osteonectin and calmodulin. Type II collagen mRNA was found in proliferating cartilage and, in lower amounts, in hypertrophic/calcifying cartilage. Type X mRNA was absent from proliferating cartilage and present in hypertrophic/calcifying cartilage at steady state levels slightly lower than that of type II mRNA. Type I mRNA was the major collagen mRNA species in endochondral bone; however, significant amounts of type X mRNA were also found. Examination of type X/type II ratios suggest that the cells producing type X mRNA in bone are different from those in the hypertrophic/calcifying cartilage region. Osteonectin mRNA was present in endochondral bone; however, significant amounts were also detected in precalcified cartilage. Indeed, the level of osteonectin mRNA was significantly higher in the resting/proliferating region than in the hypertrophic/calcifying region of the cartilage. No correlation was observed between calmodulin mRNA and the development of mineralization; levels of this message were slightly lower in endochondral bone, embryonic sterna, and calvaria than they were in chick liver and considerably lower than the calmodulin mRNA levels in chick brain.

Animals↗
PubMed1988

Yeast ARS function and nuclear matrix association coincide in a short sequence from the human HPRT locus.

A sequence that supports extrachromosomal replication of plasmids in yeast has been identified within the first intron of the human hypoxanthine-guanine phosphoribosyltransferase (HPRT) gene. This represents the first isolation of such an autonomously replicating sequence (ARS) from an exactly known position in the human genome. This ARS shares similarities of imparted yeast phenotype and DNA sequence with other heterologous ARSs. In addition, this sequence is found to be a matrix association region (MAR) on the basis of specific binding to nuclear matrices prepared from several mammalian cell types. It also exhibits anomalous electrophoretic behavior, characteristic of bent DNA, on polyacrylamide gels. The coincidence of these properties supports the possibility that this region may play a role in DNA replication within its normal chromosomal context.

Base Sequence↗
PubMed1988

Comparative study of DNase I sensitivity at the X-linked human HPRT locus.

To examine the association between chromatin structure and gene expression at the human hypoxanthine phosphoribosyltransferase (HPRT) locus, DNase I sensitivity of active and inactive genes was analyzed. In a set of human-hamster hybrid lines containing either an active or an inactive human X chromosome, or a derivative of the latter in which the HPRT gene was reactivated by 5-azacytidine treatment, only the promoter region of the gene was found to contain a hypersensitive domain, and its presence was strictly correlated with gene activity. An S1 nuclease-sensitive site was mapped upstream from the DNase I hypersensitive domain using supercoiled plasmids. The overall level of DNase I sensitivity in the interior of the HPRT gene was also assessed by comparing the degradation of polymorphic restriction fragments on active and inactive alleles in both polyclonal and monoclonal lines of female human cells. In these internally controlled experiments, the active X chromosome was found to be approximately twofold more susceptible to DNase I digestion than the inactive X chromosome.

Animals↗
PubMed1988

Effects of meal frequency on energy utilization in rats.

The effects of differences in meal frequency on body weight, body composition, and energy expenditure were studied in mildly food-restricted male rats. Two groups were fed approximately 80% of usual food intake (as periodically determined in a group of ad libitum fed controls) for 131 days. One group received all of its food in 2 meals/day and the other received all of its food in 10-12 meals/day. The two groups did not differ in food intake, body weight, body composition, food efficiency (carcass energy gain per amount of food eaten), or energy expenditure at any time during the study. Both food-restricted groups had a lower food intake, body weight gain, and energy expenditure than a group of ad libitum-fed controls. In conclusion, these results suggest that amount of food eaten, but not the pattern with which it is ingested, has a major influence on energy balance during mild food restriction.

Animals↗