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R Day

Publications and source records attributed to R Day.

At least 91 records · Page 5Linked to original sources
PubMed1998

Identification of mouse CPX-2, a novel member of the metallocarboxypeptidase gene family: cDNA cloning, mRNA distribution, and protein expression and characterization.

A novel member of the metallocarboxypeptidase gene family was identified from its homology with carboxypeptidase E and has been designated CPX-2. The cDNA of 2500 nucleotides encodes a protein of 764 amino acids that contains an N-terminal signal peptide-like sequence, a 158-residue discoidin domain, and a 400-residue carboxypeptidase domain. The 400-residue metallocarboxypeptidase domain has 59% amino acid identity with a protein designated AEBP-1; 44% to 46% identity with carboxypeptidases E, N, and Z; and lower homology with other members of the metallocarboxypeptidase gene family. The discoidin domain of CPX-2 has 22% amino acid identity with the carbohydrate-binding domain of discoideum-I, 29% to 34% identity with the phospholipid-binding domain of human factors V and VIII, and 59% identity with the discoidin-like domain on AEBP-1. CPX-2 is missing several of the predicted active-site residues that are conserved in most other members of the metallocarboxypeptidase gene family and which are thought to be required for enzyme activity. Expression of CPX-2 using the baculovirus system produced several forms of protein, from 80 to 105 kDa, but no detectable activity toward a variety of carboxypeptidase substrates. A shorter 50-kDa form of CPX-2, which contains the carboxypeptidase domain but not the discoidin domain, was also inactive when expressed in the baculovirus system. CPX-2 is able to bind to Sepharose-Arg; this binding is blocked by 10 mM Arg. Northern blot analysis showed CPX-2 mRNA in mouse brain, liver, kidney, and lung. In situ hybridization analysis of brain revealed a broad distribution. Areas that are enriched in CPX-2 include the hippocampus, cerebral cortex, median eminence, and choroid plexus. Taken together, these data suggest a widespread function for CPX-2, possibly as a binding protein rather than an active carboxypeptidase.

Amino Acid Sequence↗
PubMed1998

Daytime CPAP titration: a viable alternative for patients with severe obstructive sleep apnea.

OBJECTIVE: Continuous positive airway pressure (CPAP) is the treatment of choice for patients diagnosed with severe obstructive sleep apnea (OSA). The implementation of CPAP therapy has traditionally been based on full-night titration studies or split-night protocols. This study compared a group of patients who received a regular nocturnal CPAP titration with patients who received a daytime CPAP titration. The objective of the study was to determine if daytime CPAP titration is a viable alternative for the implementation of CPAP treatment in patients with severe OSA. STUDY DESIGN: Fourteen patients (13 men and one woman) received a daytime CPAP titration (day group). The day group was matched to 18 patients (17 men and one woman) who were titrated under a full-night regular nocturnal study (night group). Eligible patients were those with severe OSA (respiratory event index > 40). The groups were matched by age, sex, and body mass index. RESULTS: Daytime and nocturnal CPAP titration studies yielded sufficient amounts of rapid eye movement (REM) and non-REM sleep to help determine CPAP settings. Importantly, the diurnal and nocturnal CPAP titrations resulted in comparable therapeutic pressures as well as comparable resolution of sleep-disordered breathing. After 1 week of treatment, the groups exhibited similar CPAP use and comparable improvements in subjective sleepiness as indicated by their increase in sleep/wake activity inventory scores. CONCLUSIONS: Daytime CPAP titration studies may be a viable alternative for the efficient and expedient implementation of CPAP therapy among some patients with severe OSA.

Adult↗
PubMed1998

Does capsaicin affect physiologic and thermal responses of males during immersion in 22 degrees C?

BACKGROUND: Capsaicin alters thermoregulation in adult rats by producing a dose-dependent fall in body temperature and metabolism. The present investigation examined the thermal and metabolic responses in males who were fed capsaicin (CAP: 2 mg.kg(-1) body weight) vs. a placebo (PL: a maltodextrin capsule) prior to immersion in cold water. METHODS: Seven Caucasian males aged 20-28 yr were immersed in 22 degrees C twice (PL vs. CAP), for 120 min. The following were examined: metabolism (M; W.m(-2)), rectal temperature (Tre; degrees C), mean skin temperature (Tsk; degrees C), tissue insulation (I; degrees C.m(-2).W(-1) and proportion of energy derived from carbohydrate (%CHO). RESULTS: For M, Tre, I, and Tsk there was no significant differences between treatments (PL vs. CAP) when the variables were pooled jointly over time. However, significant differences across time was detected for Tre (p = 0.0003), Tsk (p = 0.0147), and M (p = 0.0036). Values for %CHO demonstrate a main effect for treatment (p = 0.0210) suggesting the CAP (46.7-/+25.9%) treatment demonstrated a decreased reliance on carbohydrate utilization for energy provision as compared to the PL (75.0+/-21.9%) treatment. Additionally, the treatment x time interaction was significant (p = 0.0096) whereby, PL differed from CAP at 5 min only. CONCLUSION: From these data it appears that while M, Tre, and Tsk differed across time, a CAP (which differentially affected percent of energy derived from carbohydrate) feeding did not differentially affect the thermal and metabolic responses of males during acute cold water immersion.

Adult↗
PubMed1997

Human and rat testis express two mRNA species encoding variants of NRD convertase, a metalloendopeptidase of the insulinase family.

Rat testis NRD convertase (EC 3.4.24.61) is a Zn2+-dependent endopeptidase that cleaves, in vitro, peptide substrates at the N-terminus of Arg residues in dibasic sites. This putative processing enzyme of the insulinase family of metallopeptidases exhibits a significant degree of similarity to insulinase and two yeast processing enzymes, Axl1 and Ste23. We report the cloning of two human testis cDNA species encoding isoforms of NRD convertase, hNRD1 and hNRD2. Whereas the hNRD1 transcript (3.7 kb) is equivalent to the previously characterized rat cDNA (rNRD1), hNRD2 and rNRD2 are 3.9 kb novel forms containing a nucleotide insertion encoding a 68-residue segment. This motif, which is inserted N-terminal of the Zn2+-binding site, HXXEH, is contained within the most conserved region among the insulinase family members. Analysis of the deduced primary sequences revealed 92% identity between rat and human orthologues. The human gene encoding NRD convertase was localized to chromosome 1p32.1-p32.2. Whereas NRD convertase is mostly expressed in testis and in 24 cell lines, low mRNA levels were detected in most of the 27 other tissues tested.

Amino Acid Sequence↗
PubMed1997

Adjuvant radiation for vulvar carcinoma: improved local control.

PURPOSE: Local recurrence is a significant problem following primary surgery for advanced vulva carcinoma. The objectives of this study were to evaluate the impact of adjuvant vulvar radiation on local control in high risk patients and the impact of local recurrence on overall survival. METHODS AND MATERIALS: From 1980-1994, 62 patients with invasive vulva carcinoma and either positive or close (less 8 mm) margins of excision were retrospectively studied. Thirty-one patients were treated with adjuvant radiation therapy to the vulva and 31 patients were observed after surgery. Kaplan-Meier estimates and the Cox proportional hazard regression model were used to evaluate the effect of adjuvant radiation therapy on local recurrence and overall survival. Independent prognostic factors for local recurrence and survival were also assessed. RESULTS: Local recurrence occurred in 58% of observed patients and 16% in patients treated with adjuvant radiation therapy. Adjuvant radiation therapy significantly reduced local recurrence rates in both the close margin and positive margin groups (p = 0.036, p = 0.0048). On both univariate and multivariate analysis adjuvant radiation and margins of excision were significant prognostic predictors for local control. Significant determinants of actuarial survival included International Federation of Gynecologists and Obstetricians (FIGO) stage, percentage of pathologically positive inguinal nodes and margins of excision. The positive margin observed group had a significantly poorer actuarial 5 year survival than the other groups (p = 0.0016) and adjuvant radiation significantly improved survival for this group. The 2 year actuarial survival after developing local recurrence was 25%. Local recurrence was a significant predictor for death from vulva carcinoma (risk ratio 3.54). CONCLUSION: Local recurrence is a common occurrence in high risk patients. In this study adjuvant radiation therapy significantly reduced local recurrence rates and may improve overall survival in certain subgroups. As salvage rates after developing local recurrence are poor adjuvant vulvar radiation should be considered for patients at risk after primary surgery.

Adult↗
PubMed1997

TGFbeta1 regulates gene expression of its own converting enzyme furin.

TGFbeta1 is known for its potent and diverse biological effects, including immune regulation, and cell growth and differentiation. We have recently shown that TGFbeta1 precursor is processed by human furin COOH-terminal to the R-H-R-R278 cleavage site to generate authentic mature TGFbeta1. In the present study, we demonstrate that steady-state furin mRNA levels are increased in rat synovial cells by 2 and 20 ng/ml TGFbeta1. Stimulation with TGFbeta1 results in a significant increase in furin mRNA levels, starting at 3 h with the peak effect observed at 12 h (2.5-fold increase +/-0.4). TGFbeta1 did not increase furin mRNA stability, and treatment of synovial cells with actinomycin D, before TGFbeta1 addition prevented the increase in fur gene expression, suggesting that the observed regulation occurs at the level of gene transcription. Treatment of synovial and NRK-49F fibroblastic cells with exogenous TGFbeta1 (5 ng/ml) or TGFbeta2 (10 ng/ml) translates into an increase in pro-TGFbeta1 processing as evidenced by the appearance of a 40-kD immunoreactive band corresponding to the TGFbeta1 NH2-terminal pro-region. Furin processing activity stimulated by TGFbeta2 correlates with significant increase in extracellular mature and heat-activable TGFbeta1 as determined by an isoform-specific ELISA assay. Taken together, these results demonstrate for the first time that TGFbeta1 upregulates gene expression of its own converting enzyme, and that this expression is translated into augmented processing of the TGFbeta1 precursor form. Such adaptive responsiveness of the TGFbeta1 convertase may represent an important aspect of TGFbeta1 bioavailibility in TGFbeta1-related processes and pathological conditions.

Amino Acid Sequence↗
PubMed1997

Cellular localization of the prohormone convertases in the hypothalamic paraventricular and supraoptic nuclei: selective regulation of PC1 in corticotrophin-releasing hormone parvocellular neurons mediated by glucocorticoids.

The prohormone convertases (PCs) are processing enzymes that activate proproteins via cleavage at specific single or pairs of basic residues. The hypothalamic paraventricular nucleus (PVN) and supraoptic nucleus (SON) are primary sites of biosynthesis of several neuroendocrine hormone precursors, including provasopressin (pro-AVP), pro-oxytocin (pro-OT), and procorticotrophin-releasing hormone (pro-CRH), which require post-translational processing to yield active products. Using in situ hybridization, we observed PC1 and PC5 mRNAs in PVN and SON magnocellular neurons, while PC2 mRNA was observed in both magnocellular and parvocellular PVN neurons as well as magnocellular SON neurons. Similar to furin, PC7 mRNA was expressed throughout the PVN and SON, whereas PACE4 mRNA levels were undetectable. Both immunohistochemical and Western blot studies were performed to demonstrate the presence of PC proteins and forms in the PVN and SON. Using double-labeling in situ hybridization, we examined the cellular colocalization of each PC mRNA with pro-AVP, pro-OT, and pro-CRH mRNAs in PVN and SON. PC1 mRNA was colocalized with both AVP and OT mRNA in PVN and SON magnocellular neurons. All AVP, OT, and CRH neurons expressed PC2. In contrast, PC5 mRNA was colocalized only with OT mRNA. We examined the effects of adrenalectomy (ADX) on PVN PC mRNA levels. PC1 mRNA levels were increased selectively within CRH/AVP parvocellular neurons but were unchanged in PVN magnocellular AVP or OT neurons. These results established the anatomical organization of each convertase and proneuropeptide substrates in the PVN and SON and suggested potential roles for each enzyme under resting and stimulated conditions.

Adrenalectomy↗
PubMed1997

A phase II trial of all-trans-retinoic acid in hormone-refractory prostate cancer: a clinical trial with detailed pharmacokinetic analysis.

Retinoids have been shown to have substantial anticancer activity in a number of preclinical and clinical situations. There are considerable epidemiologic, in vitro and in vivo data which indicate that retinoids may have a role in the prevention and therapy of human prostate cancer. Based on anecdotal evidence of response in one patient with hormone-refractory prostate cancer (HRPC), we conducted a phase II trial in HRPC during which we also examined changes in pharmacokinetics of all-trans-retinoic acid (ATRA) which occurred during therapy. Enrolled in the study were 17 patients with HRPC who received 50 mg/m2 ATRA three times daily orally on days 1-14, repeated every 22 days. The pharmacokinetics of ATRA were assessed with the first dose on day 1, again on day 14 and after a 7-day interruption in ATRA therapy on day 22. Patients were evaluable for response if they completed two 14-day courses of ATRA; among 13 such patients no responses were seen. Four patients were considered unevaluable for response owing to rapid disease progression in three and intercurrent illness in one. Apparent clearance of ATRA changed substantially during therapy: day 1 3779 +/- 4215 ml/min, day 14 7179 +/- 3197 ml/min, day 22 3213 +/- 2357 ml/min. Area under the curve was proportionately diminished on day 14 compared with day 1 and had returned to baseline by day 22. We conclude that ATRA is not active in HRPC. Failure of this agent in HRPC may be related to failure of drug delivery associated with enhanced mechanisms of ATRA clearance which occur within a few days of beginning ATRA treatment.

Adult↗
PubMed1997

Differences in traffic judgements between young and old adult pedestrians.

Older pedestrians have been shown to be over-involved in casualty crashes, compared to younger pedestrians, in recent reports. This study set out to investigate whether older pedestrians' road crossing behaviour might render them more vulnerable to crashes because of declines in their physical, sensory, perceptual or cognitive abilities. An initial 'blackspot' accident analysis highlighted the types of crashes in which older (and younger) adult pedestrians were involved and likely crossing actions. Road crossing behaviour was then systematically measured from unobtrusive video recordings of individual road crossings for a sample of younger and older pedestrians at several urban locations. On two-way undivided roads, older pedestrians crossed more frequently when there was closer moving traffic and generally adopted less safe road crossing strategies than their younger counterparts. On one-way divided roads, their crossing behaviour was considerably more safe and similar to that of younger pedestrians. The findings suggest that age-related perceptual and cognitive deficits may play a substantial role in many of the crashes involving older pedestrians.

Accidents, Traffic↗
PubMed1997

Immunohistochemical localization of the pro-peptide processing enzymes PC1/PC3 and PC2 in the human anal canal.

The distribution of prohormone/pro-peptide convertases PC1/PC3 and PC2 was investigated in the human anal canal by immunohistochemistry. Both prohormone convertases exhibited region-specific distribution patterns and were observed in neural and neuroendocrine cells and in nonneuroendocrine cellular elements. PC1/PC3 immunoreactivity was present in enteric neurons, subsets of nerve fibers, and neuroendocrine cells, and also in epithelial cells like intestinal stem cells, and a subpopulation of squamous cells. Enteric neurons were PC2 immunoreactive, whereas PC2 immunostaining in nerve fibers was slightly above background levels. Few neuroendocrine cells contained PC2 immunoreactivity, which were located predominantly in the anal transitional zone. In the squamous epithelium, the basal cell layer stained for PC2. The tissue-specific distribution of PC1/PC3 and PC2 indicates region-specific processing of peptides with regulatory functions in the anal canal and further supports the hypothesis that neuropeptides are important regulators of anal functions.

Anal Canal↗
PubMed1997

Pilocarpine-induced seizures are accompanied by a transient elevation in the messenger RNA expression of the prohormone convertase PC1 in rat hippocampus: comparison with nerve growth factor and brain-derived neurotrophic factor expression.

Several prohormone convertases that are involved in the posttranslational processing of precursor proteins, including neuropetides, hormones and neurotrophic factors, are produced in the central nervous system. These include enzymes named furin, PC1, PC2, PC5 and PACE4. To understand better the potential role played by prohormone convertases in the central nervous system we studied the expression of their messenger RNAs in the hippocampus of rats with pilocarpine-induced seizures. Moreover, we compared their expression patterns with those of neurotrophins such as nerve growth factor and brain-derived neurotrophic factor, which are up-regulated in the hippocampus during seizures. Pilocarpine (380 mg/kg, i.p.) induced seizure activity that appeared within the first hour and persisted for approximately 8 h. In situ hybridization showed transient increases in messenger RNA for nerve growth factor and brain-derived neurotrophic factor that peaked at 120 min in the hippocampus. Among the convertases studied, only PC1 messenger RNA displayed up-regulation, with temporal and topographic features comparable to those of nerve growth factor and brain-derived neurotrophic factor messenger RNA. The expression of furin, PC2 and PC5 messenger RNA changed little, while PACE4 was not expressed at all, both before and after pilocarpine administration. The highest increase in PC1 messenger RNA expression was found in granule cells of the dentate gyrus and, to a lesser extent, in the pyramidal layer of CA1 and CA3 subfields. Thus, in the rat hippocampus, the epileptiform activity induced by pilocarpine mediates a co-ordinated expression of messenger RNAs for PC1, nerve growth factor and brain-derived neurotrophic factor. Our findings suggest the involvement of PC1 in the processing of precursor proteins during seizure activity.

Animals↗
PubMed1997

Prorenin activation and prohormone convertases in the mouse As4.1 cell line.

The precise identification of prorenin-processing enzymes has been hampered by the very low abundance of juxtaglomerular cells in the kidney. Recently, an immortalized renin-producing renal tumor cell line (As4.1) has been proposed as a model to carry out such studies. Despite the fact that they contain secretory granules, we found no evidence (on the basis of enzymatic assays of renin activity in the supernatant of the cells and of immunoprecipitations experiments) that the As4.1 cells can secrete active renin through the regulated pathway. As4.1 cells produce only renin-1, as they derive from a strain of mice expressing only one renin gene. However, stable transfection of these cells with a renin-2 expression plasmid increased the capacity of this cell line to secrete active renin in the regulated pathway. Northern blot and reverse transcriptase-polymerase chain reaction amplification (RT-PCR) assays revealed that furin, PACE4 and PC5 were the only members of the proprotein convertase (PC) family to be present in these cells. As PC5 is the only such enzyme with the demonstrated ability to process mouse prorenin 2, it may constitute a candidate enzyme for the processing of prorenin-2 in mouse juxtaglomerular cells. However, it is not likely to be involved in the processing of mouse prorenin 1.

Animals↗
PubMed1997

Convertase PC2 and the neuroendocrine polypeptide 7B2 are co-induced and processed during neuronal differentiation of P19 embryonal carcinoma cells.

Convertases of the subtilisin/kexin family are responsible for the biological activation of a variety of pro-proteins, pro-hormones, and pro-trophic factors, and thus can modulate various aspects of embryonic development. We investigated the expression of each convertase by Northern hybridization during cell differentiation in vitro, using the mouse embryonal carcinoma cell line P19 as a model. The neuroendocrine convertase PC2 and 7B2, its specific binding protein, are co-induced during neuronal differentiation of P19 cells with retinoic acid, whereas the other convertases are not or follow different patterns of temporal expression. The mature forms of PC2 and 7B2 proteins are detected together by immunoblotting following induction of mRNA expression, indicating that these proteins are processed early during brain development. These results demonstrate that PC2 and 7B2 gene expression and protein processing are in a close temporal association during neuronal differentiation and point to the value of the P19 cell model to study the significance and the regulation of this relationship in mammalian brain development.

Animals↗
PubMed1997

Cloning and sequence analysis of cDNA encoding rat carboxypeptidase D.

Carboxypeptidase D (CPD) is a recently described 180-kD enzyme with carboxypeptidase E-like enzymatic properties. CPD has been proposed to be present in the secretory pathway and to contribute to peptide hormone processing in the Cpe(fat)/Cpe(fat) mouse, which lacks functional CPE. Sequence analysis of cDNA clones encoding rat CPD show the protein to contain an amino-terminal signal peptide, three carboxypeptidase-like domains, a putative transmembrane domain, and a 60-amino-acid cytoplasmic tail. Whereas active site, substrate-binding, and metal-binding residues of other metallocarboxypeptidases are conserved in the first two domains of CPD, several of the critical residues are not conserved in the third domain; this third domain is not predicted to form an active carboxypeptidase. The overall homology between rat CPD and the duck homolog gp180 is high, with 75% amino acid identity. The three carboxypeptidase domains show 66%, 83%, and 82% amino acid identity between rat CPD and duck gp180. Homology is also high in the transmembrane domain (86%) and in the cytoplasmic tail (97%). The mouse Cpd gene maps to the medial portion of chromosome 11, approximately 45.5 cM distal to the centromere. Northern blot analysis of CPD mRNA shows major bands of approximately 8 and 4 kb in many rat tissues, and additional species ranging from 1.4 to 5 kb that are expressed in some tissues or cell lines. CPD mRNA is detectable in most tissues examined, and is most abundant in hippocampus, spinal cord, atrium of the heart, colon, testis, and ovaries. In situ hybridization of CPD mRNA shows a distribution in many cells in rat brain and other tissues, with high levels in hippocampus, olfactory bulb, and the intermediate pituitary. The broad distribution is consistent with a role for CPD in the processing of many peptides and proteins that transit the secretory pathway.

Amino Acid Sequence↗
PubMed1997

Enhanced expression of endothelin-1 gene in resistance arteries in severe human essential hypertension.

BACKGROUND: Endothelins are potent vasoconstrictors, and may also act as mitogens and hypertrophic agents. Expression of a member of this family of peptides, endothelin-1, is enhanced in the endothelium of blood vessels of rats with severe forms of hypertension, even in the absence of elevated plasma endothelin levels. In some of these hypertensive models enhanced endothelin-1 gene expression may contribute to vascular hypertrophy of small arteries and to elevation of blood pressure. OBJECTIVE: To establish whether endothelin-1 may play a role in essential hypertension in humans, in whom plasma levels are known to be usually within normal limits, by examining the expression of the endothelin-1 gene in resistance-size arteries of normotensive subjects, and in humans with mild and severe hypertension. METHODS: Using in-situ hybridization, the abundance of endothelin-1 messenger RNA transcripts was evaluated in small arteries of subcutaneous gluteal fat obtained by biopsy in normotensive and hypertensive patients. RESULTS: Vessels from five normotensive subjects and four untreated mild essential hypertensive patients did not exhibit topographically localized specific labeling with the antisense human endothelin-1 probe. Biopsies from four untreated hypertensive patients with moderate-to-severe blood pressure elevation, in contrast, showed a heavy density of grains on endothelial cells of small arteries of gluteal subcutaneous fat, corresponding to hybridization of the antisense human endothelin-1 complementary RNA probe with endothelin-1 messenger RNA. CONCLUSION: Some patients with moderate-to-severe essential hypertension, similar to some experimental rat models with severe blood pressure elevation, exhibit enhanced endothelial expression of the endothelin-1 gene. This is the first demonstration that overexpression of the endothelin-1 gene may occur in the vascular wall in a small sample of this subset of hypertensive patients. This pathophysiologic phenomenon could play a role in blood pressure elevation and perhaps in the pathogenesis of vascular hypertrophy. Treatment with endothelin receptor antagonists may offer a novel therapy for these moderate-to-severe hypertensive patients.

Adipose Tissue↗