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At least 307 records · Page 17Linked to original sources
PubMed1993

Cell-specific expression in the silkmoth follicle: developmental characterization of a major chorion protein, its mRNA and gene.

Choriogenesis (eggshell formation) within the silkmoth Antheraea polyphemus proceeds in parallel for the two major subpopulations of follicle cells, diverging only during the very late period when aeropyle crown surface structures form in one region but not in the other. Correlated with their appearance is the synthesis of a set of region-specific proteins. In this report, aeropyle crowns are physically isolated and their protein composition is shown to consist of those same region-specific proteins. A cDNA clone, called pcvl 16, has been selected and shown to encode a lamellar-forming, aeropyle crown-specific protein, probably of the previously described C3,4 group. These conclusions are based on hybrid-selected translation, Northern analysis, and sequence analysis. pcvl 16 was used to isolate two distinct cloned copies of the 16 gene. Both 16 genes are closely paired with another region-specific gene but the proximity of the two gene pairs to each other is uncertain. Non-region-specific chorion genes expressed at earlier times in choriogenesis surround the 16 gene pairs, suggesting that cis sequences necessary for regionalized expression may be closely linked to coding sequences. To test this hypothesis, 5'-flanking sequences from eight region-specific genes are compared and shown to share two oligonucleotide sequences. One is a known regulatory element found in virtually all moth and fly chorion genes examined. The other, located just upstream from the TATA box, is not found in non-regionally expressed chorion genes and, thus, is a candidate for specifying regional expression.

Amino Acid Sequence↗
PubMed1992

Response of beta-endorphin and estradiol to resistance exercise in females during energy balance and energy restriction.

The acute effect of weightlifting on beta-endorphin and estradiol was studied in experienced female recreational weightlifters. Five eumenorrheic females completed two months of testing, each with a different sequence of testing conditions (SEQ1 and SEQ2). In SEQ1, a week of weight maintenance diet and prescribed exercise (3 d.wk-1, 3 sets, approximately 85% 1 RM, 10-12 reps, eight lifts) beginning on d 11 of their menstrual cycle was followed by measurement of hormone response to a weightlifting bout during energy balance (EBAL) on d 18. This included blood sampling via a catheter before, just after, and at 15 and 30 min of recovery. The women consumed 500 kcal per day for the next 48 hrs and then repeated the weightlifting test during negative energy balance (NEBAL). SEQ2 was similar except that the 48 hrs of NEBAL preceded the EBAL test condition. Estradiol and beta-endorphin increased from baseline to immediately post exercise under both dietary conditions but was significant only during NEBAL. Estradiol increased 1.6 fold and beta-endorphin 3.7 fold by the end of the resistance exercise bout during NEBAL. Both hormones were also elevated for a longer time during recovery in the NEBAL condition. Since estradiol and beta-endorphin can suppress gonadotropin release, it is possible that repeated elevations in these hormones during weightlifting, especially concurrent with energy restriction, could contribute to disruption of the menstrual cycle.

Adult↗
PubMed1985

MR imaging of the acoustic nerves and small acoustic neuromas at 0.6 T: prospective study.

To evaluate the capability of magnetic resonance (MR) in imaging normal acoustic nerves, 12 volunteers without signs or symptoms of intracranial disease were examined using a 0.6 T superconductive system. Several spin-echo (SE) pulse sequences were tested to identify the optimal sequence for demonstration of the acoustic nerve bundle. Repetition times (TRs) varied from 300 to 2000 msec and echo times (TEs) from 30 to 120 msec. A single-slice technique was used with 5 and 8 mm sections, one or two data acquisitions per projection, and axial and coronal imaging. The normal acoustic nerves were demonstrated readily by MR in axial and/or coronal sections. The distal parts of the nerves and tumors were imaged best with SE 1500/60. The medial extremities of the seventh and eighth nerves tended to be obscured in this sequence by brightening the cerebrospinal fluid signal adjacent to the brainstem, but they were demonstrated clearly with 500 or 800 msec TR and 30 msec TE. Five patients were studied who had hearing loss and evidence of retrocochlear disease. In four patients, MR imaging demonstrated five acoustic nerve tumors ranging in size from purely intracanalicular to a 12 mm cisternal component. In the fifth case, no tumor was identified by MR imaging or gas computed tomographic (CT) cisternography. Contrast-enhanced CT using a Siemens Somatom DR 3 or GE CT/T 8800 scanner failed to provide convincing evidence of tumor in any case, while gas CT cisternography was positive in all five tumors. All five acoustic neuromas were identified readily using the SE sequences that proved optimal for demonstration of normal nerves. This experience revealed that MR imaging can demonstrate the eighth nerve complex well and reliably. Single-slice (5 or 8 mm) technique is adequate, but multislice without tissue gaps (used recently) is more efficient. Small, even intracanalicular, acoustic neuromas are imaged effectively, indicating that the method is capable of superseding contrast CT cisternography, particularly with improving technology.

Atrophy↗
PubMed2024

Association analysis of mitochondrial DNA heteroplasmic variants: Methods and application.

We rigorously assessed a comprehensive association testing framework for heteroplasmy, employing both simulated and real-world data. This framework employed a variant allele fraction (VAF) threshold and harnessed multiple gene-based tests for robust identification and association testing of heteroplasmy. Our simulation studies demonstrated that gene-based tests maintained an appropriate type I error rate at &#x3b1;&#x202f;=&#x202f;0.001. Notably, when 5&#x202f;% or more heteroplasmic variants within a target region were linked to an outcome, burden-extension tests (including the adaptive burden test, variable threshold burden test, and z-score weighting burden test) outperformed the sequence kernel association test (SKAT) and the original burden test. Applying this framework, we conducted association analyses on whole-blood derived heteroplasmy in 17,507 individuals of African and European ancestries (31&#x202f;% of African Ancestry, mean age of 62, with 58&#x202f;% women) with whole genome sequencing data. We performed both cohort- and ancestry-specific association analyses, followed by meta-analysis on both pooled samples and within each ancestry group. Our results suggest that mtDNA-encoded genes/regions are likely to exhibit varying rates in somatic aging, with the notably strong associations observed between heteroplasmy in the RNR1 and RNR2 genes (p&#x202f;<&#x202f;0.001) and advance aging by the Original Burden test. In contrast, SKAT identified significant associations (p&#x202f;<&#x202f;0.001) between diabetes and the aggregated effects of heteroplasmy in several protein-coding genes. Further research is warranted to validate these findings. In summary, our proposed statistical framework represents a valuable tool for facilitating association testing of heteroplasmy with disease traits in large human populations.

Humans↗
PubMedDate not supplied

In vivo measurement of water self diffusion in the human brain by magnetic resonance imaging.

A new pulse sequence for in vivo diffusion measurements by magnetic resonance imaging (MRI) is introduced. The pulse sequence was tested on phantoms to evaluate the accuracy, reproducibility and inplane variations. The sensitivity of the sequence was tested by measuring the self diffusion coefficient of water with different temperatures. This phantom study showed that the water self diffusion could be measured accurately and that the inplane deviation was less than +/- 10 per cent. Seven healthy volunteers were studied with a 10 mm thick slice through the lateral ventricles, clear differences between grey and white matter as well as regional differences within the white matter were seen. In two patients with infarction, alternations in water self diffusion were seen in the region of the infarct. Likewise, pronounced changes in brain water self diffusion were observed in a patient with benign intracranial hypertension. The results indicate that brain water self diffusion can be measured in vivo with reasonable accuracy. The clinical examples suggest that diffusion measurements may be clinically useful adding further information about in vivo MR tissue characterization.

Adolescent↗
PubMed1993

The genes of major lysosomal membrane glycoproteins, lamp-1 and lamp-2. 5'-flanking sequence of lamp-2 gene and comparison of exon organization in two genes.

Human lysosomal membrane glycoproteins lamp-1 and lamp-2 are the major sialoglycoproteins present in lysosomal membranes. The expression of lamp-2 molecules is uniquely regulated, whereas lamp-1 is constitutively synthesized. In order to investigate the unique expression of lamp-2, and the gene evolution of lamp-1 and lamp-2, we isolated genomic phage clones encoding these glycoproteins. Comparison of the genomic and cDNA sequences revealed that the lamp-2 gene consists of nine exons. The transcriptional start site of the lamp-2 gene was determined by primer extension analysis. In order to locate the transcriptional regulatory region of this gene, various regions of 5'-sequences were tested for promoter activity using chloramphenicol acetyltransferase as a reporter molecule. The results revealed that the 5'-flanking sequence from -172 to -20 base pairs has strong promoter activity. In this sequence, potential SP1 and AP-1 binding sites and CAAT boxes are found. Most notably, the promoter activity is suppressed if the 5' farther upstream KpnI repeat sequence is included in the tested 5'-flanking sequence, thus suggesting that the KpnI repeat sequence may have some regulatory function in the lamp-2 gene expression. Comparison of the exon organization of human lamp-2 and lamp-1 genes, or chicken lamp-1 gene, reveals that these two proteins utilize the same exon phase in corresponding introns. Furthermore, each exon encodes almost identical portions of the proteins. On the other hand, the amino acid sequence of human lamp-1 is more homologous to lamp-1 of other species than it is to human lamp-2. These results indicate that lamp-1 and lamp-2 genes were most likely produced by duplication of a primordial gene, which took place early in evolution.

Amino Acid Sequence↗
PubMed1996

Decision-making: a comparison of referral practice and primary care.

BACKGROUND: Should care by subspecialist physicians be more costly than care by primary care physicians? This article addresses diagnostic testing, one element of the answer to this question. METHODS: A theoretical analysis was conducted of the sequences of testing, treatment, or watchful waiting in patients with low, intermediate, or high probabilities of disease. This was followed by a reanalysis of data from a previously published study of patients with chest pain from two referral populations and two primary care populations. The study used a chest pain score as a summary measure of the number of suggestive findings. RESULTS: The analysis of sequences of testing, treatment, and watchful waiting suggests that patients with intermediate probabilities of disease are most likely to be referred. The study of patients with chest pain shows that the probability of disease for a given chest pain history score is higher in referred patients than it is in primary care patients, as is the proportion of patients with intermediate and high chest pain scores. This result is direct evidence that referral physicians get more patients with suspect but often uncertain histories. In general, the probability of disease given a particular history will be lower in primary care patients, and hence testing will be less fruitful. CONCLUSIONS: Subspecialists are more likely to see patients who represent a diagnostic puzzle and have intermediate probabilities of disease. Since patients with intermediate probabilities of disease are most likely to benefit from testing, a per capita rate of testing that is higher than in a primary care practice might be appropriate in a subspecialist's practice.

Adult↗
PubMed1986

Frequent occurrence of short complementary sequences in nucleic acids.

The hypothesis, that nucleic acids which code specifically interacting receptor and ligand proteins contain complementary sequences was tested. Human insulin mRNA (HSINSU) contained 16 sequences which were 23.8 +/- 1.4 nucleotides long and were complementary to the insulin receptor mRNA (HSIRPR, 74.8 +/- 1.9% complementary matches, p less than 0.001 compared to randomly occurring matches). However, when examining 10 different nucleic acids (coding proteins not interacting with the insulin receptor), 81 additional sequences were found which were also complementary to HSIRPR. Although the finding of short complementary sequences was statistically highly significant, we concluded that this is not specific for nucleic acids coding specifically interacting proteins.

Base Sequence↗
PubMed2026

Prospective clinical validation of targeted long-read sequencing for preimplantation genetic testing of &#x3b1;-thalassaemia.

BACKGROUND: Preimplantation genetic testing for monogenic disorders (PGT-M) can prevent transmission of severe &#x3b1;-thalassaemia, but conventional workflows remain limited by family-specific assay design for direct variant detection, dependence on additional family samples for haplotype construction, and labour-intensive multi-step procedures across several platforms. Targeted long-read sequencing-based PGT-M for &#x3b1;-thalassaemia (tlrPGT-&#x3b1;-thal) integrates direct variant detection and haplotype linkage analysis within a single assay, but prospective clinical validation is lacking. METHODS: This prospective clinical study enrolled 103 families at high risk of transmitting &#x3b1;-thalassaemia at a reproductive medicine centre between August 2024 and March 2025. All families underwent blinded parallel analysis using both conventional NGS-based PGT-M (comparator) and tlrPGT-&#x3b1;-thal. RESULTS: In the primary concordance analysis, tlrPGT-&#x3b1;-thal was fully concordant with conventional NGS-based PGT-M (507/507, 100.0%; exact 95% CI, 99.3-100.0). Direct variant detection was successful in 501/507 embryos (98.82%; 95% CI, 97.4-99.6), haplotype linkage was established in 505/507 embryos (99.61%; 95% CI, 98.6-100.0), and one meiotic recombination event was identified. Among 93 families proceeding to embryo transfer, 57 pregnancies underwent invasive prenatal diagnosis, and all were concordant with the corresponding tlrPGT-&#x3b1;-thal results. Of the 26 comparator-inconclusive embryos, tlrPGT-&#x3b1;-thal resolved 6 complex cases, including cases with incomplete pedigrees or insufficient informative SNPs. Among the remaining 20 embryos with HBA-region aneuploidies, genotype and parental origin could be determined in 12. CONCLUSIONS: The findings show that tlrPGT-&#x3b1;-thal enables direct detection of diverse &#x3b1;-thalassaemia-causing variants together with efficient haplotype linkage analysis within a single workflow, without requiring family-specific assay design or additional family samples. The method demonstrated high diagnostic accuracy while providing added value in complex scenarios. Taken together, tlrPGT-&#x3b1;-thal represents a simplified and broadly applicable strategy for &#x3b1;-thalassaemia PGT-M.

Humans↗
PubMed1994

A small test of a sequence-based typing method: definition of the B*1520 allele.

Santamaria et al. (Human Immunology 1993 37: 39-50) describe a method of sequence-based typing (SBT) for HLA-A, B and C alleles said to give "unambiguous typing of any sample, heterozygous or homozygous, without requiring additional typing information". From SBT analysis, which involves determination of partial sequences of mixed alleles, these investigators reported that cell lines KT17 (HLA-B35,62) and OLGA (HLA-B62) from the reference panel of the 10th International Histocompatibility Workshop express novel variants of HLA-B15 (B1501-MN6) and HLA-B35 (B3501-MN7) respectively. To study further the novel alleles, we cloned and sequenced full-length HLA-B cDNA clones isolated from the KT17 and OLGA cell lines. We find that KT17 expresses B*3501, as assigned by SBT, and B*1501, the common allele encoding the B62 antigen. We were unable to confirm that KT17 expresses the novel B1501-MN6 variant identified by SBT. For OLGA our analysis confirms the partial sequences obtained by SBT. Thus OLGA expresses B*1501 and a novel HLA-B allele. The complete sequence of the latter shows it is a hybrid having exons 1 and 2 in common with B*1501 and other B15 subtypes and exons 3-7 in common with B*3501 and related molecules including B*5301 and B*5801. The novel allele has been designated B*1520 because of its sequence similarity with the B15 group; furthermore, serological analysis shows that the B*1520 product does not express epitopes in common with either B35, B53 or B58. The B*1520 heavy chain has a similar isoelectric point to A*3101; B*1520 was undetected by previous applications of isoelectric focusing because B*1520 and A31 are both expressed by OLGA. In conclusion, HLA-B typing of two cell lines by cDNA cloning and sequencing gives concordant results with SBT for three of the four alleles. The cause of the discrepancy for the fourth allele is unknown, however, this finding indicates that the novel HLA-A, B and C sequences emerging from SBT studies need independent verification.

Alleles↗
PubMed1991

Correlation between amount of virus with altered nucleotide sequence and the monkey test for acceptability of oral poliovirus vaccine.

Production of live attenuated oral poliomyelitis vaccine (OPV) requires rigorous neurovirulence safety testing of each vaccine lot, currently carried out in monkeys. It has been reported that a change from 472-U to 472-C in the type 3 OPV RNA is associated with an increased histologic lesion score produced upon intraspinal inoculation of the mutant virus in monkeys. We have developed a method, based on polymerase chain reaction, for measuring the relative abundance of these mutant sequences directly in vaccine preparations and used this method to evaluate the proportion of 472-C in 40 different lots of type 3 OPV. Six vaccine lots that had failed the intraspinal monkey neurovirulence test contained a higher proportion of 472-C than all other lots that had passed this test. OPV type 3 virus containing 472-C was rapidly selected during serial passages in African green monkey kidney cells that are used for manufacturing of the vaccine. We have also found that the wild-type poliovirus type 3 strain Leon/37, from which the vaccine strain was originally derived, contained a mixture of 472-U and 472-C sequences. No other mutations in OPV type 3 RNA have been detected by similar assays at position 2034, also associated with attenuation, or at several other positions reported to be altered in some vaccine preparations. Our results suggest that molecular diagnostics may provide a supplement or a potential alternative to animal testing of live attenuated vaccines.

Animal Testing Alternatives↗
PubMed1995

The vitamin D3 hydroxylase-associated protein is a propionamide-metabolizing amidase enzyme.

Previously we isolated a novel protein that coimmunoprecipitates with the 1,25-dihydroxyvitamin D3-24R-hydroxylase and 25-hydroxyvitamin D3-1 alpha-hydroxylase. This kidney-specific protein found in the inner membrane of mitochondria is named the vitamin D3 hydroxylase-associated protein (VDHAP). To determine a putative function for this protein, an extensive computer search of the deduced amino acid sequence of VDHAP was performed. A BLAST homology search identified amino acid residues 133 through 321 in acetamidase from Aspergillus nidulans that exhibit 38% amino acid identify and 65% amino acid similarity to VDHAP. A protein consensus sequence dictionary, MOTIFS, identified an amidase consensus sequence in VDHAP. This sequence, G-G-S-S-G-G-E-G-A-L-I-A-G-G-G-S-L-L-G-I-G-S-D-V-A-G-S-I-R-L-P-S, in VDHAP is located between amino acids 223 and 254. Propionamide, acetamide, and acrylamide were identified as substrates for an amidase activity in soluble chicken kidney mitochondria. Propionamide is the best substrate with a Vmax of 16.7 nmol NH4+/min/mg protein and an apparent Km of 7.9 mM in soluble chicken kidney mitochondria. A VDHAP monoclonal antibody, IVC2G8, immunoprecipitates 78% of the total propionamidase activity in soluble chicken kidney mitochondria. These results suggest that VDHAP is a propionamidase enzyme in soluble chicken kidney mitochondria and a member of the amidase signature gene family.

Amides↗
PubMed1995

Characterization of a soybean cDNA clone encoding the mitochondrial isozyme of aspartate aminotransferase, AAT4.

A soybean leaf cDNA clone, pSAT2, was isolated by hybridization to a carrot aspartate aminotransferase (EC 2.6.1.1.; AAT) cDNA clone at low stringency. pSAT2 contained an open reading frame encoding a 47640 Da protein. The protein encoded by pSAT2 showed significant sequence similarity to AAT proteins from both plants and animals. It was most similar to two Panicum mitochondrial AATs, 81.5% and 82.0% identity. Alignment of the pSAT2-encoded protein with other mature AAT enzymes revealed a 25 amino acid N-terminal extension with characteristics of a mitochondrial transit peptide. A plasmid, pEXAT2, was constructed to encode the mature pSAT2 protein lacking the putative mitochondrial transit peptide. Escherichia coli containing the plasmid expressed a functional AAT isozyme which comigrated with the soybean AAT4 isozyme during agarose gel electrophoresis. Equilibrium sucrose gradient sedimentation of soybean extracts demonstrated that AAT4 specifically cofractionated with mitochondria. Antibodies raised against the pEXAT2-encoded AAT protein reacted with AAT4 of soybean and not with other AAT isozymes detected in soybean tissues, providing further evidence that clone pSAT2 encodes the soybean mitochondrial isozyme AAT4.

Amino Acid Sequence↗
PubMed1991

Magnetic resonance imaging for localizing the nonpalpable undescended testis.

A prospective evaluation of the value of magnetic resonance imaging (MRI) for localizing the nonpalpable undescended testis was performed in 17 boys aged 1 year to 5 years 4 months who had a clinical diagnosis of nonpalpable testes. The results were compared between the MRI, ultrasonographic, clinical, and definitive surgical diagnosis in all patients. Nine nonpalpable undescended testes were identified in the inguinal canal or just proximal to the inguinal ring by MRI, and these were confirmed by surgical findings. Six nonpalpable testes were not observed by MRI, and the absence of these testes was surgically confirmed. Like scrotal testes, all the undescended testes showed a low intensity on sequences with a short repetition time (TR) and echo time (TE), and a high intensity on long TR/TE sequences. Undescended testes were observed better on coronal images than on axial images. MRI promises to become an important diagnostic tool in the detection of nonpalpable undescended testes.

Child, Preschool↗
PubMed1975

Structural genes adjacent to interspersed repetitive DNA sequences.

The observation that repetitive and single copy sequences are interspersed in animal DNAs has suggested that repetitive sequences are adjacent to single copy structural gene sequences. To test this concept, single copy DNA sequences contiguous to interspersed repetitive sequences were prepared from sea urchin DNA by hydroxyapatite fractionation (repeat-contiguous DNA fraction). These single copy sequences included about one third of the total nonrepetitive sequence in the genome as determined by the amounts recovered during the hydroxyapatite fractionation and by reassociation kinetics. 3H-labeled mRNA from sea urchin gastrula was prepared by puromycin release from polysomes and used in DNA-driven hybridization reactions. The kinetics of mRNA hybridization reactions with excess whole DNA were carefully measured, and the rate of hybridization was found to be 3-5 times slower than the corresponding single copy DNA driver reassociation rate. The mRNA hybridized with excess repeat-contiguous DNA with similar kinetics relative to the driver DNA. At completion 80 percent of that mRNA hybridizable with whole DNA (approximately 65 percent) had reacted with the repeat-contiguous DNA fraction (50 percent). This result shows that 80-100 percent of the mRNA molecules present in sea urchin embryos are transcribed from single copy DNA sequences adjacent to interspersed repetitive sequences in the genome.

Animals↗
PubMed1990

The amino acid sequence of the outer coat protein VP2 of neutralizing monoclonal antibody-resistant, virulent and attenuated bluetongue viruses.

Monoclonal antibodies which reacted with four different epitopes were used to select neutralization-resistant variants of Australian bluetongue virus serotype 1 (BTV1AUS; isolate CS156). Nucleotide sequencing of the VP2 outer coat protein gene of these variants showed that two of them contained alterations within the previously defined neutralization site at amino acids 328 to 335 (Gould et al., 1988). Comparison of VP2 sequences of several BTV serotypes, in addition to nucleotide sequence changes in a number of variants, suggested that this neutralization site was larger and contained 19 amino acids, the conformation of which could be affected by other regions of the VP2 protein. Nucleotide sequencing of neutralization-resistant variants revealed a total of four other regions of VP2 affecting the ability of monoclonal antibodies to neutralize the virus and these results support the notion that the neutralization site in VP2 was conformation dependent. The complete nucleotide sequence of the VP2 gene of virulent BTV1AUS (C5156) was determined directly from viral nucleic acid isolated from the blood of a sheep suffering clinical bluetongue disease. Comparison of the VP2 sequence of this virulent virus with that previously published for an avirulent, laboratory strain (Gould, 1988), indicated that the passage of virulent virus approximately 20 times in tissue culture over the last decade, not only led to attenuation but resulted in the appearance of ten nucleotide changes in the VP2 gene. Six of these nucleotide changes were silent, two resulted in conservative amino acid substitutions and two generated radical amino acid changes. However, in a separate experiment, a single passage of the virulent virus in tissue culture while leading to attenuation did not result in a nucleotide change in the VP2 outer coat protein gene.

Amino Acid Sequence↗
PubMed1991

Purification and characterization of a heat-stable enterotoxin of Vibrio mimicus.

A heat-stable enterotoxin produced by Vibrio mimicus (VM-ST) was studied. VM-ST was purified from a culture supernatant of V. mimicus strain AQ-0915 by ammonium sulfate fractionation, hydroxyapatite treatment, ethanol extraction, column chromatography on both SP-Sephadex C-50 and DEAE-Sephadex A-25, and HPLC, and the recovery rate was about 15%. Purified VM-ST was heat-stable. VM-ST activity was cross-neutralized by anti-STh antiserum. The amino acid composition of the purified VM-ST was determined 17 amino acid residues in the following sequence: Ile-Asp-Cys-Cys-Glu-Ile-Cys-Cys-Asn-Pro-Ala-Cys-Phe-Gly-Cys-Leu-Asn. This composition and sequence were identical to those of V. cholerae non-O1-ST. These results clearly demonstrate the production of a characteristic VM-ST by V. mimicus.

Amino Acid Sequence↗