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S L White

Publications and source records attributed to S L White.

At least 37 records · Page 2Linked to original sources
PubMed2000

Organization of the rat PACAP gene.

The rat PACAP gene has been isolated, cloned, and sequenced. The gene organization is very similar to that of the mouse and human PACAP genes. An analysis of the 5'-UTR in proPACAP mRNAs indicates that multiple transcripts are generated by using different splice sites within exons 1A, 1B, and 2 to form the 5'-UTR. Analysis of PACAP promoter activity in PC12 cells suggests that forskolin responsiveness of the gene is dependent on elements located between -77 and -413 bp from the transcription start site.

5' Untranslated Regions↗
PubMed1999

Mitochondrial DNA mutations at nucleotide 8993 show a lack of tissue- or age-related variation.

Two pathogenic mitochondrial DNA mutations, a T-to-G substitution (8993T > G) and a T-to-C substitution (8993T > C), at nucleotide 8993 have been reported. We describe 13 pedigrees with mitochondrial DNA mutations at nucleotide 8993; 10 pedigrees with the 8993T > G mutation and three with the 8993T > C mutation. Prenatal diagnosis of the nucleotide 8993 mutations is technically possible. However, there are three major concerns: (i) that there is variation in mutant loads among tissues; (ii) that the mutant load in a tissue may change over time; and (iii) that the genotype-phenotype correlation is not clearly understood. We have used the 13 pedigrees to determine specifically the extent of tissue- and age-related variation of the two mutations at nucleotide 8993 in the mitochondrial DNA. The tissue variation was investigated by analysing two or more different tissues from a total of 18 individuals. The age-related variation of the mutation was investigated by comparing the amount of both mutations in blood taken at birth and at a later age. No substantial tissue variation was found, nor was there any substantial change in the proportion of either mutation over periods of 8-23 years in the four individuals studied. In addition, we noted that two features were remarkably common in families with nucleotide 8993 mutations, namely (i) unexplained infant death (8 cases in 13 pedigrees); and (ii) de novo mutations (5 of the 10 8993T > G pedigrees).

Aging↗
PubMed1999

Age dependence of smooth muscle myosin expression by cultured rat aortic smooth muscle cells.

Vascular smooth muscle cells (SMC) in vivo are highly heterogeneous phenotypically, particularly during development and in the adult during periods of remodeling. Much remains to be learned, however, regarding regulation of the SMC phenotype at the gene level. Here, we studied smooth muscle myosin heavy chain (SMMHC) expression at the transcriptional and mRNA levels in SMC cultured from newborn, adult, and old animals, which express different patterns of differentiation markers. We also examined regulation of SMMHC gene expression by TGF-beta, a cytokine known to be involved in the differentiation process. The activity of SMMHC promoter constructs, the expression of which is smooth-muscle-specific, was greatest in SMC from newborn animals and least in cells from old animals. Thus, differences in the degree of differentiation of SMC from these three sources may at least in part be due to transcriptional events. SMC from the three animal sources each contained mRNAs for the SM-1A and SM-2A tail but not those for the SM-1B and SM-2B head isoforms. Total SMMHC mRNA levels reflected similar differences as found at the transcriptional level. SM-2A mRNA as a proportion of total SMMHC mRNA was greatest in SMC from newborn animals, consistent with their higher degree of differentiation. TGF-beta up-regulated both transcription and mRNA levels but did not change the proportions of SMMHC mRNAs. Though the levels of transcriptional activity and mRNA were widely different in untreated cells, the degree of TGF-beta stimulation was approximately the same in all cases.

Age Factors↗
PubMed1999

Genetic counseling and prenatal diagnosis for the mitochondrial DNA mutations at nucleotide 8993.

Mitochondrial genetics is complicated by heteroplasmy, or mutant load, which may be from 1%-99%, and thus may produce a gene dosage-type effect. Limited data are available for genotype/phenotype correlations in disorders caused by mtDNA mutations; therefore, prenatal diagnosis for mtDNA mutations has been hindered by an inability to predict accurately the clinical severity expected from a mutant load measured in fetal tissue. After reviewing 44 published and 12 unpublished pedigrees, we considered the possibility of prenatal diagnosis for two common mtDNA mutations at nucleotide 8993. We related the severity of symptoms to the mutant load and predicted the clinical outcome of a given mutant load. We also used the available data to generate empirical recurrence risks for genetic counseling, which may be used in conjunction with prenatal diagnosis.

Adenosine Triphosphatases↗
PubMed1999

Smooth-muscle myosin heavy-chain SM-B isoform expression in developing and adult rat lung.

The smooth-muscle cells composing the vasculature and airways of the lung display a variety of contractile protein phenotypes. To date, however, it has remained unclear how these phenotypes might contribute differentially to contractile activity. To address this issue, we made monospecific rabbit polyclonal antibodies against the difference peptide for the SM-B smooth-muscle myosin heavy chain (SMMHC) and used these to investigate the distribution of the SM-B isoform in lung. SM-B has a seven-amino acid insert in the head region that is known to result in a higher actin-activated adenosine triphosphatase activity and in vitro motility. During development, reactivity is first seen in the trachea and bronchi of saccular lung at the time of birth, when other SMMHC isoforms also are present. Immunoreactivity spreads distally through the airways as development proceeds, reaching the level of alveolar septae in the adult. Although the smaller vessels of the pulmonary vasculature react strongly with the SM-B antibody, reactivity is infrequently observed in large pulmonary vessels. Adult tracheal smooth muscle is highly and more uniformly reactive, commensurate with its relatively high maximal velocity of shortening. The differential expression of the SM-B isoform in vascular and airway smooth muscles demonstrated in this study may provide the molecular basis for functional differences between these smooth-muscle cell types and may provide one mechanism for adapting contractility in response to physiologic stresses in the lung.

Aging↗
PubMed1999

Two cases of prenatal analysis for the pathogenic T to G substitution at nucleotide 8993 in mitochondrial DNA.

We report the outcome of two prenatal analyses for the T to G mutation at nucleotide 8993 in the mitochondrial DNA. This mutation is associated with neurogenic muscle weakness, ataxia and retinitis pigmentosa (NARP) and the neurodegenerative condition, Leigh syndrome. One prospective mother was the sister of a severely affected individual, and had previously had an unaffected child and a stillborn child. The second prospective mother had two unaffected children and two affected children. The mutation was not detected in the chorionic villus sample from one fetus nor in the amniocytes from the other fetus. Both pregnancies were continued, and the resulting children were healthy at two years and five years of age. Prenatal diagnosis of this mitochondrial DNA mutation is an option likely to be acceptable to some families to prevent the birth of a child at high risk for neurological disease.

Adult↗
PubMed1998

Smooth muscle-specific expression of the smooth muscle myosin heavy chain gene in transgenic mice requires 5'-flanking and first intronic DNA sequence.

The smooth muscle myosin heavy chain (SM-MHC) gene encodes a major contractile protein whose expression exclusively marks the smooth muscle cell (SMC) lineage. To better understand smooth muscle differentiation at the transcriptional level, we have initiated studies to identify those DNA sequences critical for expression of the SM-MHC gene. Here we report the identification of an SM-MHC promoter-intronic DNA fragment that directs smooth muscle-specific expression in transgenic mice. Transgenic mice harboring an SM-MHC-lacZ reporter construct containing approximately 16 kb of the SM-MHC genomic region from -4.2 to + 11.6 kb (within the first intron) expressed the lacZ transgene in all smooth muscle tissue types. The inclusion of the intronic sequence was required for transgene expression, since 4.2 kb of the 5'-flanking region alone was not sufficient for expression. In the adult mouse, transgene expression was observed in both arterial and venous smooth muscle, in airway smooth muscle of the trachea and bronchi, and in the smooth muscle layers of all abdominal organs, including the stomach, intestine, ureters, and bladder. During development, transgene expression was first detected in airway SMCs at embryonic day 12.5 and in vascular and visceral SMC tissues by embryonic day 14.5. Of interest, expression of the SM-MHC transgene was markedly reduced or absent in some SMC tissues, including the pulmonary circulation. Moreover, the transgene exhibited a heterogeneous pattern between individual SMCs within a given tissue, suggesting the possibility of the existence of different SM-MHC gene regulatory programs between SMC subpopulations and/or of episodic rather than continuous expression of the SM-MHC gene. To our knowledge, results of these studies are the first to identify a promoter region that confers complete SMC specificity in vivo, thus providing a system with which to define SMC-specific transcriptional regulatory mechanisms and to design vectors for SMC-specific gene targeting.

Animals↗
PubMed1998

Fluid, electrolyte, and acid-base balances in three-day, combined-training horses.

Horses competing in 3-day, combined-training events develop a metabolic acidosis that is partially compensated for by a respiratory alkalosis immediately after phases B and D. By the end of phase C and 30 minutes to 2 hours after phase D, the acidosis is resolved by the oxidation of lactate, and a metabolic alkalosis prevails. A reduction in TBW and cation content occurs, which often is not replenished 12 to 24 hours after the event, even though the serum or plasma concentration of various constituents may be within normal limits. Hypochloremia and hypocalcemia, however, may persist 12 or more hours after the speed and endurance test. All of the data cited in this article are from horses that successfully completed their respective tests. Nevertheless, some horses developed substantial fluid and cation losses. In horses that are not well conditioned or in competitions in which terrain, footing, or hot environments increase the thermal load or decrease heat loss, greater losses of fluids and electrolytes can be expected. Body weight losses exceeding 5% and cation losses exceeding 4000 mEq/L occur in endurance horses suffering from exhaustion and synchronous diaphragmatic flutter. In one study, two thirds of the Na+ lost during exercise-induced sweating in cool, dry conditions was replenished from salt supplements added to a balanced forage and concentrated diet. Consequently, horses in regular training and competition may benefit from salt supplementation. The composition of the salt supplement and the amount fed should be based on the composition of the horse's diet, degree of work, and environmental conditions. Horses competing in a 3-day, combined-training event may be expected to have persistent losses of weight and cations, particularly if conditions result in heavy sweating. Many horses in the field studies had minimal changes in weight and cation balance compared with pre-event values. The diet and electrolyte supplementation of the horses in the majority of these studies were unknown; consequently, no firm conclusions may be made regarding the degree of weight, water, and cationic losses. It is likely that the horse's large intestine serves as a reservoir for both fluid and electrolytes so that performance is not diminished, despite losses demonstrated consistently in treadmill studies. Horses that are inadequately trained or have other systemic or musculoskeletal problems or those that are competing in hot conditions may develop a variety of metabolic problems. It is hoped that knowledge of the fluid and electrolyte losses may help in treating horses with metabolic problems and, more importantly, in improving management of horses in training and competition to prevent any loss of performance.

Acid-Base Equilibrium↗
PubMed1998

Lung smooth muscle differentiation.

The vascular and visceral smooth muscle tissues of the lung perform a number of tasks that are critical to pulmonary function. Smooth muscle function often is compromised as a result of lung disease. Though a great deal is known about regulation of smooth muscle cell replication and cell and tissue contractility, much less is understood regarding the phenotype of the contractile protein machinery of lung smooth muscle cells. This review focuses on the expression of cytoskeletal and contractile proteins of lung vascular and airway smooth muscle cells during development, in the adult and during vascular and airway remodeling. Emphasis is placed on the expression of the heavy chain of smooth muscle myosin, as well as the regulation of its gene. Important areas for future research are discussed.

Amino Acid Sequence↗
PubMed1998

Myosin heavy chain isoform expression in rat smooth muscle development.

Smooth muscle myosin heavy chains (MHCs), the motor proteins that power smooth muscle contraction, are produced by alternative splicing from a single gene. The smooth muscle MHC gene is capable of producing four isoforms by utilizing alternative splice sites located at the regions encoding the carboxy terminus and the junction of the 25- and 50-kDa tryptic peptides. These four isoforms, SM1A, SM1B, SM2A, and SM2B, are a combination of one of two heavy chains containing different carboxy-terminal tails (1 or 2) without (A) or with (B) an additional motif in the myosin head. In the present study, using RNA analysis and isoform-specific antibodies, we demonstrate the expression patterns of MHC isoforms during development in rat smooth muscle tissues. RNase protection analysis indicates that the mRNAs for SMA and SMB isoforms, which differ by a 21-nucleotide insertion in the region encoding the S1 head region of the myosin molecule, are differentially expressed during development in a highly tissue-specific manner. Smooth muscle MHC transcripts are first detectable in developing rat smooth muscle tissues at 17 days of fetal development. The SMB mRNA is shown to be expressed in smooth muscle from fetal bladder, intestine, and stomach and from neonatal aorta; however, it is not expressed in cultured smooth muscle cells from rat aorta. The SMA mRNA is also present at all stages of development in the smooth muscles examined; however, it is much less abundant than SMB mRNA in most fetal smooth muscles. We show here that the SMB isoform, which contains a unique seven-amino acid insertion at the junction of the 25- and 50-kDa tryptic peptides, is present in conjunction with SM1 and SM2 tails on immunoblots of smooth muscle from stomach, intestine, bladder, and uterus and is expressed during development in a pattern distinct from that of the SM1 and SM2 tail isoforms.

Aging↗
PubMed1997

Expression of the smooth muscle myosin heavy chain gene is regulated by a negative-acting GC-rich element located between two positive-acting serum response factor-binding elements.

To identify cis- and trans-acting factors that regulate smooth muscle-specific gene expression, we studied the smooth muscle myosin heavy chain gene, a rigorous marker of differentiated smooth muscle. A comparison of smooth muscle myosin heavy chain promoter sequences from multiple species revealed the presence of a highly conserved 227-base pair domain (nucleotides -1321 to -1095 in rat). Results of a deletion analysis of a 4.3-kilobase pair segment of the rat promoter (nucleotides -4220 to +88) demonstrated that this domain was necessary for maximal transcriptional activity in smooth muscle cells. Gel-shift analysis and site-directed mutagenesis demonstrated that one true CArG and another CArG-like element contained within this domain were both recognized by the serum response factor and were both required for the positive activity attributable to this domain. Additional studies demonstrated that mutation of a GC-rich sequence within the 227-base pair conserved domain resulted in a nearly 100% increase in transcriptional activity. Gel-shift analysis showed that this GC-rich repressor element was recognized by both Sp1 and Sp3. These data demonstrate that transcriptional control of the smooth muscle myosin heavy chain gene is highly complex, involving both negative and positive regulatory elements, including CArG sequences found in the promoters of multiple smooth muscle differentiation marker genes.

Animals↗
PubMed1996

Identification of promoter elements involved in cell-specific regulation of rat smooth muscle myosin heavy chain gene transcription.

In order to identify cis-acting regulatory elements involved in smooth muscle cell-specific gene regulation, we have cloned a 4. 7-kilobase pair fragment of the promoter for the rat smooth muscle myosin heavy chain, a protein expressed in differentiated smooth muscle cells. Sequence analysis of a 1.7-kilobase pair portion of this clone reveals potential binding sites for known transcription factors. A comparison of the primary sequence between the rat and rabbit smooth muscle myosin heavy chain promoters reveals numerous conserved consensus binding sites. Transient transfection analysis of promoter deletion constructs in rat aorta and tracheal smooth muscle cells, L8 myoblast cells, and rat pulmonary aorta endothelial cells suggests that a region of the promoter located between -1,249 and -1,317 base pairs is important for the restriction of gene expression to smooth muscle cells. Electrophoretic mobility shift analysis of a highly conserved region located between -1,317 and -1, 085 base pairs reveals specific DNA-protein complexes formed in smooth muscle cell extracts, which can be competed with an oligonucleotide containing a nuclear factor 1 binding site.

Animals↗
PubMed1996

Differential effects of CTLA-4 substitutions on the binding of human CD80 (B7-1) and CD86 (B7-2).

CTLA-4 expressed on activated T cells binds to CD80 (B7-1) and CD86 (B7-2) molecules present on APC with high avidity and appears to deliver a negative regulatory signal to the T cell. We have investigated the kinetics of CTLA-4 binding to CD80 and CD86, together with the effects of selected CTLA-4 mutations on binding activity. The dissociation constants (Kd) for binding of CTLA-4-Ig to CD80 and CD86 transfectants were 8.1 and 6.7 nM, respectively. Surface plasmon resonance was used to determine kinetic parameters of CTLA-4-Ig binding to CD80-Ig and CD86-Ig fusion proteins and revealed enhanced association (ka) and dissociation (kd) rate constants for CD86-Ig compared with CD80-Ig. Furthermore, CD80-Ig and CD86-Ig fusion molecules demonstrated variable abilities to cross-compete for binding to several modified forms of CTLA-4-Ig. Differential binding of CD80 and CD86 to CTLA-4 was further revealed by analysis of 10 discrete CTLA-4 mutants. Five single amino acid substitutions within the CTLA-4 MYPPPY domain exerted modest effects on CD80 binding, but each of these substitutions completely abrogated CD86 binding. In addition, substitutions just N-terminal of the MYPPPY region, and within the CDR1-like region of CTLA-4, eliminated both CD80 and CD86 binding. Hence, CD80 and CD86 bind with different association/dissociation kinetics to similar, but distinct, sites on CTLA-4.

Abatacept↗
PubMed1996

Biochemical changes in three-day-event horses at the beginning, middle and end of Phase C and after Phase D.

Blood samples were collected 12-16 h before the Speed and Endurance test, immediately after steeplechase, midway through Phase C (4 km marker), at the end of Phase C and immediately after cross-country from 36 horses that completed a 3-day-event at the CCI* (n = 19) or CCI** (n = 17) level. Packed cell volume (PCV), plasma concentrations of sodium, potassium, chloride, ionized calcium, pH and lactate; and serum concentrations of total protein (TP), albumin, total calcium, alkaline phosphatase (AP), gamma glutamyl transferase (GGT), creatine kinase (CK), aspartate aminotransferase (AST), glucose and creatinine were measured. No differences were noted in any biochemical measurements between the CCI* and CCI** horses at any sampling time, despite differences in speed and length of various phases of the Speed and Endurance test. For all horses (n = 36), biochemical parameters changed significantly over time (P < 0.01). The PCV, calcium, potassium, lactate, total protein and albumin concentrations were significantly increased over pre-exercise concentrations immediately after Phase B. During Phase C, these parameters decreased towards pre-exercise concentrations, thereby supporting the concept that Phase C is a recovery phase. In contrast, chloride concentration decreased, and the creatinine concentration and CK increased compared to pre-exercise concentrations from the end of Phase B to the end of Phase C. These changes probably resulted from fluid and electrolyte losses in sweat, reduced renal blood flow and continued muscular activity. Many parameters did not change significantly between the 4 km marker (midway point on Phase C) and the end of Phase C. However, there were a few notable exceptions: potassium, chloride, lactate and glucose concentrations decreased, whereas pH and creatinine concentrations increased significantly from the 4 km marker to the end of Phase C. The most significant benefit of the extra distance from the 4 km marker to the end of Phase C was that it facilitated further dissipation of lactate concentrations prior to the start of Phase D.

Alkaline Phosphatase↗
PubMed1996

In vitro and in vivo ultrastructural changes induced by macrolide antibiotic LY281389.

High doses of LY281389 (9-N-(n-propyl)-erythromycylamine) cause cytoplasmic vacuolar changes in striated and smooth muscle characteristic of drug-induced phospholipidosis. This study characterized phospholipidosis in striated and smooth muscle of rats and dogs, compared in vivo observations with those in a cultured rat myoblast model, and attempted to confirm the lysosomal origin of the drug-induced vacuoles. Standard transmission electron microscopy and acid phosphatase cytochemistry techniques were used to evaluate ultrastructural changes in vivo and in vitro. Rats and dogs exposed to LY281389 had a time- and dose-related increase in number and size of vacuoles containing concentric lamellar figures in cardiac and skeletal muscle. Cytochemical staining of dog stomach smooth muscle for acid phosphatase, a lysosomal enzyme, stained the periphery of vacuoles that contained concentric lamellar figures. Cultured rat L6 myoblast cells were exposed to 0.25 mg LY281389/ml for 2.5, 5, 10, 20, 30, or 90 min and 2, 6, 12, 24, or 48 hr. Cell cultures exposed for 2 hr had several predominantly large, clear, membrane-bound vacuoles, and at 6 and 12 hr there were greater numbers of large vacuoles that contained increased amounts of membranous figures. Following 24- or 48-hr exposures, vacuoles occupied most of the cytoplasmic volume, and were engorged predominantly with amorphous or granular material. These findings indicate that LY281389 can induce similar phospholipidosis-like vacuolar changes in rat and dog muscle and in a cultured rat muscle cell line. Further, positive acid phosphatase staining of drug-induced vacuolar structures, in conjunction with standard transmission electron microscopy techniques, strongly suggests that vacuoles seen in vitro and in vivo are lysosomal in origin.

Acid Phosphatase↗
PubMed1995

CNS development of health maintenance programs: quality improvement and cost reduction.

Several outpatient health maintenance programs were developed by CNSs that resulted in quality improvement and cost savings. Common implementation strategies, barriers to implementation, marketing issues and techniques, and revenue generation issues were found. An outpatient consultation model was used to expand enterostomal therapy and diabetic teaching services. Education and support programs include a cardiac support group and community education classes on aging. Preadmission programs include physical therapy consultation for patients scheduled for elective orthopedic surgery. Outpatient programs facilitate continuity of care in a managed care format, promote cost savings, and provide unique services that encourage patients to return to our hospital, thereby increasing market share. Key barriers encountered include inconsistent or ambiguous administrative support, budgetary constraints, lack of collaboration, communication problems, facility limitations, funding considerations, resource allocation, and territoriality. The programs are consistent with the change in focus of health care from treatment to prevention.

Ambulatory Care↗