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Biomedical subjects

L K Miller

Publications and source records attributed to L K Miller.

At least 145 records · Page 8Linked to original sources
PubMed1988

Baculoviruses for foreign gene expression in insect cells.

Insect baculovirus vector systems are rapid and convenient means of obtaining substantial quantities of many different foreign gene products. The vector systems are proving useful in academic research and in industry. Numerous examples of the successful high-level expression of biologically active vertebrate proteins have been reported. Cleavage of mammalian signal sequences occurred correctly in the cases studied. Vertebrate and invertebrate core glycosylation appears to be similar, but terminal glycosylation differs substantially. Nevertheless, many biologically active glycosylated vertebrate proteins can be expressed in biologically active form using this system. The ease and rapidity of the vector systems are two of their most valuable features. The vectors will be particularly convenient for the rapid analysis of the products of genes subjected to site-specific modifications.

Animals↗
PubMed1988

Influence of young maternal age and parity on term and preterm low birthweight.

Data on 4496 singleton births to young women (19 years or less) are reported by maternal age and parity with birthweight and gestation cross-classified to yield rates of preterm and term low birthweight. After adjustment, the risk of preterm low birthweight was increased with very young maternity (15 years or less); preterm low birthweight and term low birthweight were each increased with young multiparity. These data suggest that the identification of factors associated with preterm birth and their incorporation into the prenatal care regimen may be important in improving pregnancy outcome in young women.

Adolescent↗
PubMed1988

Autoimmune antibody (IgG Kansas) against the fibrin stabilizing factor (factor XIII) system.

Serum from a patient who died from massive hemorrhage within 4 months after onset of an acquired bleeding disorder at age 85 contained a potent inhibitor of fibrin stabilization. Other parameters of coagulation and fibrinolysis and his bleeding time were within normal limits. The inhibitor was shown to be an IgG with kappa light chains (IgG Kansas); its specific target was the factor XIII system itself. Although IgG Kansas combined with the virgin [ab] form of the zymogen, it did not block the thrombin-catalyzed conversion to [a'b]. However, IgG Kansas prevented the subsequent Ca2+-mediated activation of [a'b] to a + b, where a denotes the catalytically competent factor XIIIa species. IgG Kansas, in contrast to a previously studied autoimmune antibody from a similar bleeding disorder (IgG Warsaw), could also inhibit the transamidating activity of the preactivated a enzyme.

Aged↗
PubMed1988

Characterization of an early gene accelerating expression of late genes of the baculovirus Autographa californica nuclear polyhedrosis virus.

The region of the Autographa californica nuclear polyhedrosis virus (AcMNPV) encompassing the EcoRI T fragment (29.0 to 30.1 map units) was characterized by DNA sequencing, transcriptional mapping, and site-directed mutagenesis. The largest transcript from this region, an early 1.7-kilobase (kb) poly(A)+ RNA, encompassed three tandem, nonoverlapping open reading frames (ORFs). The largest of these ORFs, ETL, was proximal to the 5' end of the transcript and had the capacity to encode a 28-kilodalton (kDa) polypeptide. A recombinant virus, vETL beta gal, containing the Escherichia coli beta-galactosidase (beta gal) gene fused to the N-terminal two-thirds of the ETL ORF, produced blue plaques in the presence of a chromogenic indicator of beta gal and wild-type levels of polyhedra in cell culture. This recombinant was also infectious in insect larvae by oral administration of occluded virus. Comparison of vETL beta gal and wild-type viral proteins pulse-labeled at various times postinfection (p.i.) revealed (i) absence of a virus-induced 28-kDa polypeptide, (ii) early expression of a large (approximately 130-kDa) polypeptide which may be the ETL-beta gal fusion protein, (iii) a delay in expression of early 35 and 40-kDa polypeptides, and (iv) a 4- to 6-h delay in the expression of late proteins in vETL beta gal-infected cells. Cycloheximide did not inhibit synthesis of the 1.7-kb RNA but did inhibit its shutoff, which occurs at 12 h p.i. in the absence of inhibitors. Thus, the ETL gene product is apparently an early 28-kDa protein which is necessary, directly or indirectly, for timely expression of many other AcMNPV genes. The promoter-leader regions of the 1.7-kDa transcript showed significant sequence similarities to the leader of the AcMNPV IE-1 gene. The middle ORF within the 1.7-kb transcript, ETM, would encode a hydrophobic polypeptide of 113 amino acid residues. ETS, a small ORF within and proximal to the 3' end of the 1.7-kb transcript, was also transcribed as a set of smaller (approximately 0.5-kb) RNAs initiated heterogeneously in the region between ETL and ETS and persisting throughout infection.

Amino Acid Sequence↗
PubMed1988

Expression and complex formation of simian virus 40 large T antigen and mouse p53 in insect cells.

Recombinant baculoviruses were constructed which express simian virus 40 large T antigen (SVT-Ag) or murine p53 to high levels in infected insect cells. Characterization of the expressed proteins revealed that they display many properties of the corresponding mammalian-derived proteins. Both proteins are of wild-type size, localize to the nucleus, are recognized by several SVT-Ag- or p53-specific monoclonal antibodies, and are phosphorylated in this system. Complexes are formed between baculovirus-derived SVT-Ag and p53 after coinfection of insect cells with both recombinant viruses. After infection of insect cells with either virus individually, each protein can self-associate to form a variety of oligomeric species. Pulse-chase experiments indicated that both SVT-Ag and p53 are highly stable in insect cells, even in the absence of complex formation.

Animals↗
PubMed1988

Effect of massive weight loss on hypothalamic-pituitary-gonadal function in obese men.

To study the ability of weight loss to reverse the hyperestrogenemia-induced hypogonadotropic hypogonadism that occurs in obese men, we measured the 24-h mean plasma free and total estradiol (E2), total estrone, FSH, LH, and free and total testosterone concentrations in 11 healthy obese men (100-305% above desirable body weight) and again 5-39 months later after weight loss of 26-129 kg and restabilization at the new weight. Weight loss produced significant increases in mean plasma total testosterone [240 +/- 116 (+/- SD, 8.5 +/- 4.0) to 377 +/- 113 ng/dL (13.0 +/- 4.0 nmol/L); P less than 0.01], free testosterone [9.5 +/- 5.0 (329 +/- 173) to 13.4 +/- 4.3 ng/dL (464 +/- 149 pmol/L); P less than 0.025], and FSH (6.5 +/- 4.7 to 10.9 +/- 8.5 IU/L; P less than 0.025). Plasma LH was lower than levels in normal men before and after weight loss and did not change significantly (10.3 +/- 4.8 and 10.8 +/- 6.8 IU/L, respectively). There was no change in plasma total E2 [54 +/- 26 (196 +/- 94) to 50 +/- 13 pg/mL (180 +/- 50 pmol/L)], free E2 [1.48 +/- 0.7 (5.37 +/- 2.54) to 1.33 +/- 0.42 pg/mL (4.83 +/- 1.45 pmol/L)], or total estrone [75 +/- 38 (280 +/- 140) to 82 +/- 24 (300 +/- 90) pmol/L], and sex hormone-binding globulin rose from 9.2 +/- 3.2 to 12.9 +/- 5.4 nmol/L (P less than 0.005). The increases in plasma free and total testosterone and sex hormone-binding globulin were proportional to the degree of weight loss. Thus, the hypogonadotropic hypogonadism was largely reversed by the weight loss without any decrease in hyperestrogenemia, its presumed cause. We postulate a change in hypothalamic-pituitary function with weight loss, such that GnRH-gonadotropin secretion becomes less sensitive to suppression by a given amount of estrogen.

Adult↗
PubMed1988

Dual carcinoid tumors of Meckel's diverticulum presenting as metastasis in an inguinal hernia sac: case report with literature review.

Neoplasms arising in Meckel's diverticulum or involving hernia sacs are rare. The authors present a unique case of two asymptomatic carcinoid tumors arising in a Meckel's diverticulum, which were discovered because of microscopic metastasis in a grossly unremarkable inguinal hernia sac. This article describes the clinical and morphologic features of this unusual case, reviews the topic of neoplasms involving hernia sacs and Meckel's diverticula, and includes a brief review of immunocytochemical findings in carcinoid tumors.

Aged↗
PubMed1988

The location, sequence, transcription, and regulation of a baculovirus DNA polymerase gene.

The Autographa californica nuclear polyhedrosis virus (AcMNPV) DNA polymerase gene was identified with the aid of an oligonucleotide probe corresponding to an amino acid sequence conserved among viral DNA polymerases of other virus families. A 3.6-kb pair region of the AcMNPV DNA, from 39.5 to 42.5 map units (m.u.), was sequenced and an open reading frame (ORF) of 2994 bp was observed. From the first ATG of this ORF, a translation product of 984 amino acids (Mr 114,310) was predicted. Amino acid sequence similarities to other viral DNA polymerases were found. Transcription was analyzed by Northern RNA blot analysis and nuclease protection studies of RNA:DNA hybrids. The ORF is transcribed in the counterclockwise direction as a 3-kb RNA. Transcripts appear to initiate at two differently regulated sites (ca. -120 and -212 bp) upstream of the initiating ATG (+1,+2,+3) and to be polyadenylated at a single site near a signal (A2UA3) which overlaps the translational termination signal (UAA) at +2952. Transcripts were observed only during a narrow window between 2 to 8 hr postinfection (p.i.) with maximum expression between 4 and 6 hr p.i. Polymerase gene transcripts were observed in the presence of the protein synthesis inhibitor cycloheximide which also blocked the shut-off of these early transcripts. Aphidicolin, an inhibitor of both viral and host DNA polymerases, inhibited polymerase gene transcription suggesting a unique regulation involving DNA replication.

Amino Acid Sequence↗
PubMed1988

Partial reversal of the hypogonadotropic hypogonadism of obese men by administration of corticosuppressive doses of dexamethasone.

Obese men have hyperestrogenemia-induced hypogonadotropic hypogonadism (HHG), due, we believe, to increased rarmatization of adrenal androgens by the increased bulk of aromatase-containing adipose tissue. We studied the effects of corticosuppressive doses of dexamethasone (D) on 24-h mean plasma total and free estradiol (E2), estrone (E1), LH, FSH, total and free testosterone, delta 4-androstenedione (delta 4), and sex-hormone-binding globulin (SHBG) in nine obese men and five normal-weight controls. In the obese men, the following hormones fell: E2 [59 +/- 19 to 39 +/- 11 pg/ml (P less than 0.01)], E1 [93 +/- 41 to 50 +/- 25 pg/ml; (P less than 0.01)], delta 4-androstenedione [120 +/- 80 to 55 +/- 27 ng/dl; (P less than 0.02)]; free E2 [1.6 +/- 0.4 to 1.1 +/- 0.2 pg/ml; (P less than 0.01)], SHBG [12.8 +/- 5.3 to 8.2 +/- 3 nM/l; (P less than 0.04)]. FSH rose from 4.8 +/- 3.2 to 7.6 +/- 4.2 miu/ml (P less than 0.01). LH, total and free testosterone showed no significant change. In the nonobese men, there were decreases in total E2 [(34 +/- 6.8 to 25 +/- 10 pg/ml; P less than 0.04)], SHBG [16.8 +/- 7.5 to 10.4 +/- 2.0 nM/l: P less than .05.], free E2 [0.9 +/- 0.2 to 0.7 +/- 0.3 pg/ml: P less than 0.05], delta 4 [91.4 +/- 3.6 to 33.4 +/- 16.7 ng/dl; P less than .01] and total T [492 +/- 44 to 393 +/- 121 ng/dl; P less than 0.04]. There was no significant change in E1, FSH, LH or free T.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1987

Differential binding of dexamethasone to ammonium sulfate precipitates of human adipose tissue cytosols.

Saturation analysis of the binding of [3H]dexamethasone [( 3H]DEX) to ammonium sulfate precipitates (ASPs) confirmed the presence of a limited-capacity, high-affinity binder in human adipose tissue cytosols. Various non-radioactive steroids competed with [3H]DEX for binding to the ASPs in the following sequence: dexamethasone (DEX) approximately equal to triamcinolone acetonide (TA) greater than progesterone (P) much greater than estradiol (E2). The steroid specificity of the binder precipitated by AS was consistent with the specificities reported for glucocorticoid receptors in a number of systems. In order to investigate possible regional differences, glucocorticoid binding to ASPs derived from adipose tissues removed from two different sites in the same subject was quantitated. ASPs of human omental adipose tissue bound significantly more [3H]DEX than did similar preparations of subcutaneous adipose tissue from the abdominal wall (116 +/- 32 vs. 50 +/- 22 fmol/mg protein; mean +/- SD; p less than 0.02). The findings are consistent with reports from other laboratories suggesting that intra-abdominal fat is more responsive to glucocorticoids than is subcutaneous adipose tissue.

Adipose Tissue↗
PubMed1987

Effects of serial passage of Autographa californica nuclear polyhedrosis virus in cell culture.

A study of the major genomic alterations occurring during serial passage of Autographa californica nuclear polyhedrosis virus (AcNPV) in a Trichoplusia ni cell line was conducted. Progeny viruses from 24 independent passages were randomly selected and analyzed with restriction endonucleases. Specific deletion mutations were generated repeatedly in the PstI-G (7.6 to 13.1%) and the PstI-I (14.4-17.9%) regions; these mutations became predominant in the serially passaged stocks in which they arose. The deletions in the PstI-G region and two different insertions in this region were mapped to a 1 Kb PvuII-Bg/II fragment (9.85-10.70%) reflecting a high degree of sequence specificity in the initiation or selection of genomic alterations in this region. Insertional mutations were observed frequently and repeatedly within the PstI-E/HindIII-I region (33.6-37.2%) of the AcNPV genome. Individual examples of insertional mutations were observed in several other regions of the genome.

Animals↗
PubMed1987

Baculovirus interaction with nontarget organisms: a virus-borne reporter gene is not expressed in two mammalian cell lines.

The safety of baculoviruses with respect to mammalian species was studied by using a genetically engineered recombinant of Autographa californica nuclear polyhedrosis virus. This recombinant contains the chloramphenicol acetyltransferase (CAT) gene under the control of a mammalian-active promoter and expresses substantial levels of CAT activity on infection of permissive and nonpermissive insect cells (L.F. Carbonell, M.J. Klowden, and L.K. Miller, J. Virol. 56:153-160, 1985). Extremely low levels of CAT activity were detected in mouse and human cell lines that were continuously exposed to the A. californica nuclear polyhedrosis virus recombinant. The appearance of CAT was not inhibited by cycloheximide. Isopycnic centrifugation of purified inoculum showed that a low level of CAT activity was associated with the insect-derived viral particles. Thus, the observed CAT activity is carried into the cells with the virus inoculum, and active expression of the baculovirus-borne CAT gene is not observed in either cell line. The inability of the CAT gene to be expressed in these cell lines with this model system provides additional assurance of the safety of insect baculoviruses with respect to mammalian species.

Acetyltransferases↗
PubMed1987

Location, transcription, and sequence of a baculovirus gene encoding a small arginine-rich polypeptide.

The nucleotide sequence of the cDNA of an abundant late 0.5-kilobase transcript of Autographa californica nuclear polyhedrosis virus revealed a small open reading frame encoding an arginine-rich 6.9-kilodalton protein. The predicted amino acid composition of the 6.9-kilodalton protein was essentially identical to that of the core protein of viral nucleocapsids. The precise location of the 5' and 3' ends of the transcript were confirmed by S1 nuclease and primer extension analyses. Multiple overlapping transcripts through this region include three early and three abundant late RNAs which are transcribed counterclockwise and one transient RNA which is transcribed clockwise with respect to the physical map of the virus.

Arginine↗
PubMed1987

Expression of polyomavirus large T antigen by using a baculovirus vector.

A gene encoding the large T antigen of polyomavirus was inserted into the baculovirus Autographa californica nuclear polyhedrosis virus so that gene expression was under the control of the strong, very late polyhedrin gene promoter. Significantly more large T antigen was produced in recombinant virus-infected insect cells than was observed in polyomavirus-transformed mouse cells. The insect-derived T antigen exhibited polyomavirus origin-specific DNA binding. The baculovirus expression system provides a convenient source of T antigen for in vitro studies.

Animals↗
PubMed1987

Divergent transcription of early 35- and 94-kilodalton protein genes encoded by the HindIII K genome fragment of the baculovirus Autographa californica nuclear polyhedrosis virus.

The organization of viral genes within the 3.7-kilobase-pair HindIII-K/EcoRI-S region of the Autographa californica nuclear polyhedrosis virus genome (85 to 88 map units) was determined by using a combination of nucleotide sequencing, transcriptional mapping, and in vitro translation of hybrid selected RNA. Two nonoverlapping genes, extending in opposite directions and encoding polypeptides with molecular weights of 35,000 and 94,000 (35K and 94K polypeptides), were identified. Unspliced, messenger-active RNAs were transcribed from both genes early (2 h) after infection. Indicative of immediate-early genes, transcription of the divergent RNAs was unaffected by the protein synthesis inhibitor, cycloheximide. Late in infection, abundant RNAs were transcribed from promoters located at least 2.5 kilobase pairs upstream from the gene encoding the 35K polypeptide. These transcripts completely overlapped both the 35K and 94K polypeptide genes but apparently lacked protein-coding potential, suggesting that the transcripts may play a role in the suppression of early viral gene expression.

Amino Acid Sequence↗
PubMed1987

Tuberculosis risk among migrant farm workers on the Delmarva peninsula.

A survey was conducted to ascertain the risk of tuberculosis (TB) among migrant farm workers on the Delmarva peninsula. Relevant histories were obtained from 842 migrants; a total of 709 skin tests were completed, and 239 sputum specimens were examined for acid-fast bacilli (AFB) and culture. No cases of infectious tuberculosis were ascertained by history or AFB examination. One sputum culture was positive for M. tuberculosis and 13 were positive for various species of nontuberculous mycobacteria. Thirty-seven per cent of migrants tested had significant skin test reactions of 10 mm or more. Reaction rates for men were 41 per cent and for women 25 per cent. Age specific rates ranged from 14 per cent in children aged 5-14 to 54 per cent for ages 45-54. Rates for the principal national/ethnic groups were Haitians 55 per cent, Mexicans 36 per cent, US Blacks 29 per cent and US-born Latinos 20 per cent. Based on these results and other information currently available, it is recommended that current Centers for Disease Control (CDC) recommendations for TB prophylaxis continue to be applied for migrant workers, but that mass screening by skin testing in camp populations not be emphasized. Other recommendations focus on: case finding of active disease, improving continuity and follow-up, increasing coordination among involved agencies, and actively supporting improved economic and living conditions for migrant farm workers.

Adolescent↗
PubMed1987

Developmentally delayed musical savant's sensitivity to tonal structure.

A developmentally delayed, musically gifted child with no formal musical training was asked to repeat passages on the piano. Analysis of responses to melodies in each of the 24 major and minor keys indicated sensitivity to aspects of diatonic structure exhibited by mature listeners.

Child, Preschool↗