Consensus approach to nasal high-flow therapy in neonates.

OBJECTIVE: Nasal high-flow therapy (nHFT) is commonly used for noninvasive respiratory support in the neonatal intensive care unit. Our objective was to determine which aspects of neonatal nHFT have achieved adequate evidence base to support consensus among experienced clinical investigators, and to document areas lacking consensus to promote future investigations. STUDY DESIGN: Prospective, modified Delphi collation of tabular queries related to specific aspects of neonatal nHFT. Seven international nHFT clinical researchers were queried regarding approaches to initiation, escalation, weaning and discontinuing nHFT. Completed tables were reviewed independently by each investigator, results clarified and discussed and areas of consensus determined. RESULTS: Consensus agreement was reached for many aspects of nHFT including: need for adequate heating and humidification, need to prevent nares occlusion, maximum flow rate of 8 l min-1, assessment of fraction of inspired oxygen (FiO2) and work of breathing for either flow escalation or weaning, equivalence of nHFT to nasal continuous positive airway pressure (nCPAP) for noninvasive support of infants of ⩾28 weeks with resolving respiratory distress and use of nHFT for noninvasive support of stable infants on nCPAP. There was general agreement for initial gas flow rates in the range of 4 to 6 l min-1 and for nHFT as primary therapy for mild respiratory distress. There was no consensus on the approach to discontinuing nHFT. CONCLUSIONS: Among an experienced group of nHFT clinical researchers, there was general consensus in the approach to neonatal nHFT. Additional randomized studies are indicated to provide better evidence related to several aspects of nHFT, as well as to identify other clinical conditions where nHFT may provide safe, effective noninvasive support.

A trial of nebulised heparin to limit lung injury following cardiac surgery.

Cardiac surgery with cardiopulmonary bypass triggers an acute inflammatory response in the lungs. This response gives rise to fibrin deposition in the microvasculature and alveoli of the lungs. Fibrin deposition in the microvasculature increases alveolar dead space, while fibrin deposition in alveoli causes shunting. We investigated whether prophylactic nebulised heparin could limit this form of lung injury. We undertook a single-centre double-blind randomised trial. Forty patients undergoing elective cardiac surgery with cardiopulmonary bypass were randomised to prophylactic nebulised heparin (50,000 U) or placebo. The primary endpoint was the change in arterial oxygen levels over the operative period. Secondary endpoints included end-tidal CO2, the alveolar dead space fraction and bleeding complications. We found nebulised heparin did not improve arterial oxygen levels. Nebulised heparin was, however, associated with a lower alveolar dead space fraction (P <0.05) and lower tidal volumes at the end of surgery (P <0.01). Nebulised heparin was not associated with bleeding complications. In conclusion, prophylactic nebulised heparin did not improve oxygenation, but was associated with evidence of better alveolar perfusion and CO2elimination at the end of surgery.

Non-invasive pulsed cavitational ultrasound for fetal tissue ablation: feasibility study in a fetal sheep model.

OBJECTIVES: Currently available fetal intervention techniques rely on invasive procedures that carry inherent risks. A non-invasive technique for fetal intervention could potentially reduce the risk of fetal and obstetric complications. Pulsed cavitational ultrasound therapy (histotripsy) is an ablation technique that mechanically fractionates tissue at the focal region using extracorporeal ultrasound. In this study, we investigated the feasibility of using histotripsy as a non-invasive approach to fetal intervention in a sheep model. METHODS: The experiments involved 11 gravid sheep at 102-129 days of gestation. Fetal kidney, liver, lung and heart were exposed to ultrasound pulses (< 10 µs) delivered by an external 1-MHz focused ultrasound transducer at a 0.2-1-kHz pulse-repetition rate and 10-16 MPa peak negative pressure. Procedures were monitored and guided by real-time ultrasound imaging. Treated organs were examined by gross and histological inspection for location and degree of tissue injury. RESULTS: Hyperechoic, cavitating bubble clouds were successfully generated in 19/31 (61%) treatment attempts in 27 fetal organs beneath up to 8 cm of overlying tissue and fetal bones. Histological assessment confirmed lesion locations and sizes corresponding to regions where cavitation was monitored, with no lesions found when cavitation was absent. Inability to generate cavitation was primarily associated with increased depth to target and obstructing structures such as fetal limbs. CONCLUSION: Extracorporeal histotripsy therapy successfully created targeted lesions in fetal sheep organs without significant damage to overlying structures. With further improvements, histotripsy may evolve into a viable technique for non-invasive fetal intervention procedures.

Initial investigation of acoustic droplet vaporization for occlusion in canine kidney.

Acoustic droplet vaporization (ADV) shows promise for spatially and temporally targeted tissue occlusion. In this study, substantial tissue occlusion was achieved in operatively exposed and transcutaneous canine kidneys by generating ADV gas bubbles in the renal arteries or segmental arteries. Fifteen canines were anesthetized, among which 10 underwent laparotomy to externalize the left kidney and five were undisturbed for transcutaneous ADV. The microbubbles were generated by phase conversion of perfluoropentane droplets encapsulated in albumin or lipid shells in the blood. A 3.5-MHz single-element therapy transducer was aligned with an imaging array in a water tank with direct access to the renal artery or a segmental artery. In vivo color flow and spectral Doppler imaging were used to identify the target arteries. Tone bursts of 1 kHz pulse repetition frequency with 0.25% duty cycle vaporized the droplets during bolus passage. Both intracardiac (IC) and intravenous (IV) injections repeatedly produced ADV in chosen arteries in externalized kidneys, as seen by B-mode imaging. Concurrent with this in two cases was the detection by pulse-wave Doppler of blood flow reversal, along with a narrowing of the waveform. Localized cortex occlusion was achieved with 87% regional flow reduction in one case using IC injections. Vaporization from IV injections resulted in a substantial echogenicity increase with an average half-life of 8 min per droplet dose. Gas bubbles sufficient to produce some shadowing were generated by transcutaneous vaporization of intrarenal artery or IV-administered droplets, with a tissue path up to 5.5 cm.

Features of carnitine palmitoyltransferase type I deficiency.

Carnitine palmitoyltransferase type I (CPT I) is unique among long-chain fatty acid oxidation enzymes in that there are two tissue-specific isoforms, 'hepatic' and 'muscle', which are encoded by two separate genes. The 'hepatic' isoform is expressed in liver, kidney and fibroblasts and at low levels in the heart, while the other isoform occurs in skeletal muscle and is the predominant form in heart. Reported patients with CPT I deficiency lack activity of the hepatic isoform and present before 30 months of age with hypoketotic hypoglycaemia, hepatomegaly with raised transaminases, seizures and coma. We discuss four new cases in three families showing, variously, renal tubular acidosis, transient hyperlipidaemia and, paradoxically, myopathy with elevated creatinine kinase or cardiac involvement in the neonatal period as additional features that deserve wider recognition.

Enhancing smoking cessation of low-income smokers in managed care.

BACKGROUND: Although office-based and telephone support services enhance the rate of smoking cessation in managed care systems, it is not clear whether such services are effective for very low-income smokers. We evaluated the comparative effectiveness of usual care (physician-delivered advice and follow-up) and usual care enhanced by 6 computer-assisted telephonic-counseling sessions by office nurses and telephone counselors for smoking cessation in very low-income smokers in Medicaid managed care. METHODS: A randomized clinical trial comparing the 2 approaches was conducted in 3 Michigan community health centers. All clinicians and center staff received standard training in usual care. Selected nurses and telephone counselors received special training in a computer-assisted counseling program focusing on relapse prevention. RESULTS: The majority of the study population (233 adult smokers with telephones) were white (64%) women (70%) with annual incomes of less than $10,000 (79%) and with prescriptions of nicotine replacement therapy (>90%). At 3 months, quit rates (smoke-free status verified by carbon monoxide monitors) were 8.1% in the usual-care group and 21% in the telephonic-counseling group (P=.009) by intention-to-treat analysis. Special tracking methods were successful in maintaining participants in treatment. CONCLUSIONS: Smoking cessation rates are enhanced in a population of very low-income smokers if individualized telephonic-counseling is provided. State and Medicaid managed care plans should consider investing in both office-based nurse and centralized telephonic-counseling services for low-income smokers.

Complementary mechanisms of enhanced oxytocin-stimulated prostaglandin E2 synthesis in rabbit amnion at the end of gestation.

The up-regulation of oxytocin (OT) receptors in rabbit amnion at the end of gestation is associated with a large increase in the ability of OT to stimulate PGE2 synthesis. The purpose of these investigations was to determine what other factors contribute to this increase. OT enhanced PGE2 synthesis at several levels. The concentrations of cytosolic phospholipase A2, which generates arachidonic acid for PGE2 synthesis, and PGH endoperoxide synthases (types 1 and 2), which catalyze the conversion of arachidonic acid to prostanoids, rose substantially in rabbit amnion at term. OT stimulated translocation of cytosolic phospholipase A2 to the cell particulate fraction, presumably by a Ca2+-mediated process, and phosphorylation of cytosolic phospholipase A2 via the extracellular regulated protein kinase 2/1-mediated pathway. OT-stimulated increases in intracellular Ca2+ concentrations and extracellular regulated protein kinase 2/1 phosphorylation were both mediated by G(q/11) activation. OT also increased the expression of PGH endoperoxide synthase-2 after treatment of amnion cells in culture for 2 h; however, PGE2 release in response to OT was virtually immediate. These findings show that the rapid stimulation of PGE2 synthesis by OT occurs through cytosolic phospholipase A2 activation and PGH endoperoxide synthase-1 activity, both of which, along with OT receptor concentrations, are considerably up-regulated in the amnion at the end of gestation.

Functional oxytocin receptors in a human endometrial cell line.

OBJECTIVE: Our goal was to demonstrate expression and functionality of oxytocin receptors in a human endometrial cell line. This cell line could then be used for further investigation of the role of oxytocin in reproductive function at the cellular level. STUDY DESIGN: Oxytocin receptor messenger ribonucleic acid expression was determined by reverse transcriptase-polymerase chain reaction deoxyribonucleic acid amplification with ribonucleic acid from confluent Ishikawa cells. Ligand binding to whole cells was evaluated by nonlinear regression analysis with an iodinated oxytocin antagonist. The coupling of the oxytocin receptor to signaling pathways was evaluated by measuring oxytocin-stimulated increases in intracellular calcium concentration, phosphorylation of ERK2 (extracellular-regulated protein kinase 2) mitogen-activated protein kinase, and prostaglandin E(2) release. RESULTS: Polyacrylamide gel electrophoresis of the reverse transcriptase-polymerase chain reaction products demonstrated the presence of oxytocin receptor messenger ribonucleic acid in Ishikawa cells. Ligand-binding analysis of these cells demonstrated a single class of noninteracting sites, with a B(max) (maximal number of binding sites) of 77.7 fmol/mg deoxyribonucleic acid and an apparent dissociation constant of 8.3 x 10(-11) mol/L. Stimulation with 100-nmol/L oxytocin caused a rapid transient increase in intracellular free calcium concentration, which was blocked by 1-micromol/L oxytocin antagonist. Treatment of cells with oxytocin for 10 minutes resulted in a marked increase in the phosphorylation of ERK2, as determined by Western blot analysis, and a 5-fold increase in prostaglandin E(2) release. CONCLUSION: This study is the first to demonstrate functional oxytocin receptors in an established human endometrial cell line. This cell line will be useful in elucidating the mechanisms of action of oxytocin in the reproductive tract at the molecular level.

The rational clinical examination. Does this patient have strep throat?

CONTEXT: Sore throat is a common complaint, and identifying patients with group A beta-hemolytic streptococcal pharyngitis (strep throat) is an important task for clinicians. Previous reviews have not systematically reviewed and synthesized the evidence. OBJECTIVE: To review the precision and accuracy of the clinical examination in diagnosing strep throat. DATA SOURCE: MEDLINE search for articles about diagnosis of strep throat using history-taking and physical examination. STUDY SELECTION: Large blinded, prospective studies (having > or =300 patients with sore throat) reporting history and physical examination data and using throat culture as the reference standard were included. Of 917 articles identified by the search, 9 met all inclusion criteria. DATA EXTRACTION: Pairs of authors independently reviewed each article and used consensus to resolve discrepancies. DATA SYNTHESIS: The most useful findings for evaluating the likelihood of strep throat are presence of tonsillar exudate, pharyngeal exudate, or exposure to strep throat infection in the previous 2 weeks (positive likelihood ratios, 3.4, 2.1, and 1.9, respectively) and the absence of tender anterior cervical nodes, tonsillar enlargement, or exudate (negative likelihood ratios, 0.60, 0.63, and 0.74, respectively). No individual element of history-taking or physical examination is accurate enough by itself to rule in or rule out strep throat. Three validated clinical prediction rules are described for adult and pediatric populations. CONCLUSIONS: While no single element of history-taking or physical examination is sufficiently accurate to exclude or diagnose strep throat, a well-validated clinical prediction rule can be useful and can help physicians make more informed use of rapid antigen tests and throat cultures.

The proximal portion of the COOH terminus of the oxytocin receptor is required for coupling to g(q), but not g(i). Independent mechanisms for elevating intracellular calcium concentrations from intracellular stores.

As the oxytocin receptor plays a key role in parturition and lactation, there is considerable interest in defining its structure/functional relationships. We previously showed that the rat oxytocin receptor transfected into Chinese hamster ovary cells was coupled to both G(q/11) and G(i/o), and that oxytocin stimulated ERK-2 phosphorylation and prostaglandin E(2) synthesis via protein kinase C activity. In this study, we show that deletion of 51 amino acid residues from the carboxyl terminus resulted in reduced affinity for oxytocin and a corresponding rightward shift in the dose-response curve for oxytocin-stimulated [Ca(2+)](i). However, oxytocin-stimulated ERK-2 phosphorylation and prostaglandin E(2) synthesis did not occur in cells expressing the truncated receptor. Oxytocin also failed to increase phospholipase A activity or activate protein kinase C, indicating that the mutant receptor is uncoupled from G(q)-mediated pathways. The Delta51 receptor is coupled to G(i), as oxytocin-stimulated Ca(2+) transients were inhibited by pertussis toxin, and a Gbetagamma sequestrant. Preincubation of Delta51 cells with the tyrosine kinase inhibitor, genistein, also blocked the oxytocin effect. A Delta39 mutant had all the activities of the wild type oxytocin receptor. These results show that the portion between 39 and 51 residues from the COOH terminus of the rat oxytocin receptor is required for interaction with G(q/11), but not G(i/o). Furthermore, an increase in intracellular calcium was generated via a G(i)betagamma-tyrosine kinase pathway from intracellular stores that are distinct from G(q)-mediated inositol trisphosphate-regulated stores.

Enhancing smoking cessation rates in primary care.

BACKGROUND: The Agency for Health Care Policy and Research (AHCPR) guidelines on smoking cessation recommend that primary care physicians provide both brief advice against smoking and follow-up care for all smokers. Surveys show that although physicians understand the importance of smoking cessation, the actual implementation of these guidelines is limited. The main objective of our study was to evaluate the comparative effectiveness of 2 different approaches to smoking cessation counseling: practice-based and community-based. METHODS: Both smoking cessation approaches consisted of 1 recruitment session and 6 computer-assisted counseling sessions. In the practice-based approach, counseling was provided by office nurses and telephone counselors; in the community-based approach, the counseling was given by telephone counselors only. Four practices in 3 mid-Michigan communities participated, including 120 physicians and 487 patients who were smokers. The physicians were trained to provide brief advice for smoking cessation consistent with the AHCPR guidelines; the nurses and telephone counselors were trained in relapse prevention, computer skills, and individual case management. Sixty-two percent of the participants obtained free nicotine replacement therapy. RESULTS: At 6 months, quit rates (7-day smoke-free status) were 35% in the practice-based group and 36% in the community-based group. Participants who completed at least 4 sessions showed higher quit rates than those who did not. CONCLUSIONS: Nurses in primary care practices and counselors can be trained to deliver effective relapse-prevention counseling during office visits and by telephone. Our study showed an increase in the reported rates of smoking cessation by using these counseling methods.

Functional oxytocin receptors discovered in human osteoblasts.

Undifferentiated or differentiated human trabecular bone cells with osteogenic capacity in primary culture express oxytocin receptors (OTRs). OTR expression then persists upon differentiation to an osteoblast phenotype. A human epithelial osteosarcoma cell line, Saos-2, also expresses OTRs. Expression was determined both at mRNA and protein levels. Functional OTRs are evidenced by an increase in intracellular calcium concentration, [Ca2+]i, in response to 10 nM oxytocin (OT). An oxytocin antagonist (OTA) blocked this effect, demonstrating specificity for OT. OT also stimulated prostaglandin E2 (PGE2) synthesis in both confluent undifferentiated and differentiated human trabecular bone cells. This is the first report of OTR mRNA and protein expression and of prescribed OT signal pathways in osteoblastic cells. Since PGE2 has been shown to increase bone turnover in favor of bone formation, OT may be a new class of a bone anabolic agent.

Multiple protein kinase pathways are involved in gastrin-releasing peptide receptor-regulated secretion.

Gastrin-releasing peptide (GRP) and its amphibian homolog, bombesin, are potent secretogogues in mammals. We determined the roles of intracellular free Ca(2+) ([Ca(2+)](i)), protein kinase C (PKC), and mitogen-activated protein kinases (MAPK) in GRP receptor (GRP-R)-regulated secretion. Bombesin induced either [Ca(2+)](i) oscillations or a biphasic elevation in [Ca(2+)](i). The biphasic response was associated with peptide secretion. Receptor-activated secretion was blocked by removal of extracellular Ca(2+), by chelation of [Ca(2+)](i), and by treatment with inhibitors of phospholipase C, conventional PKC isozymes, and MAPK kinase (MEK). Agonist-induced increases in [Ca(2+)](i) were also inhibited by dominant negative MEK-1 and the MEK inhibitor, PD89059, but not by an inhibitor of PKC. Direct activation of PKC by a phorbol ester activated MAPK and stimulated peptide secretion without a concomitant increase in [Ca(2+)](i). Inhibition of MEK blocked both bombesin- and phorbol 12-myristate 13-acetate-induced secretion. GRP-R-regulated secretion is initiated by an increase in [Ca(2+)](i); however, elevated [Ca(2+)](i) is insufficient to stimulate secretion in the absence of activation of PKC and the downstream MEK/MAPK pathways. We demonstrated that the activity of MEK is important for maintaining elevated [Ca(2+)](i) levels induced by GRP-R activation, suggesting that MEK may affect receptor-regulated secretion by modulating the activity of Ca(2+)-sensitive PKC.

Demonstration of functional oxytocin receptors in human breast Hs578T cells and their up-regulation through a protein kinase C-dependent pathway.

Oxytocin (OT) receptors (OTRs) have been demonstrated in a number of human breast tumors and tumor cells, but it was not clear whether the receptors were functional. We examined the regulation and function of OTR in a tumor cell line, Hs578T, derived from human breast. These cells expressed moderate levels of OTR when cultured in 10% FBS, as demonstrated by RT-PCR and binding analyses. Serum deprivation resulted in the loss of OTRs, with no effect on cell viability. Restoration of serum and addition of 1 microM dexamethasone (DEX) increased OTR levels by about 9-fold. Up-regulation was blocked by the addition of phospholipase C and PKC inhibitors. Serum/DEX treatment also increased steady state OTR messenger RNA levels. OT increased intracellular Ca2+ in a time- and dose-responsive manner, and the effects of OT were lost when OTRs were down-regulated by serum starvation. Serum/DEX up-regulation of OTR restored the responsiveness to OT. OT also stimulated ERK-2 (extracellular signal-regulated protein kinase) phosphorylation and PGE2 synthesis in Hs578T cells. In addition to showing that OTRs in the breast tumor cells are functional, these studies show that Hs578T cells can be used to study molecular regulation of OTR gene expression and intracellular signaling pathways stimulated by OT.

The role of micturating cystourethrography in antenatally detected mild hydronephrosis.

BACKGROUND: The postnatal imaging of infants with antenatally detected mild hydronephrosis remains controversial. OBJECTIVE: Our aim was to establish the role and timing of micturating cystourethrography (MCUG) in mild hydronephrosis. MATERIALS AND METHODS: We performed a retrospective study of 61 infants (122 kidneys) referred with an antenatal diagnosis of hydronephrosis who showed persistent postnatal dilatation. All had follow-up postnatal ultrasound (US) and MCUG performed. The degree of dilatation at each follow-up scan was recorded. RESULTS: Of the 122 kidneys, 65 showed mild hydronephrosis. A substantial proportion of these (21.5%) demonstrated reflux. Serial US of these infants showed that the 6-week scan was the most informative and that any changes that warranted further investigation had occurred by this time. CONCLUSION: We recommend that all infants with mild hydronephrosis should undergo MCUG. MCUG need not be delayed until 3 months but could be performed following a 6-week US scan.

Emergence of resistant variants of HIV in vivo during monotherapy with the proteinase inhibitor saquinavir.

We examined the phenotypic and genotypic properties of virus from the peripheral blood mononuclear cells (PBMC) and plasma of eight HIV-1-infected asymptomatic patients before and during monotherapy with the proteinase inhibitor saquinavir. Susceptibility of primary isolates to drug was assessed in PBMC culture by deriving IC50 and IC90 values. The observed increases in IC50 and IC90 after approximately one year of therapy with a dosage of 600 mg tds suggests the presence of virus resistant to saquinavir in vivo. The magnitude of this altered susceptibility ranged from three-fold to in one case 100-fold. In two patients a greater than eight-fold decrease in susceptibility to saquinavir was observed. Sequencing of the proteinase genes in viral RNA obtained from patient plasma and/or PBMC was carried out by PCR in parallel with sensitivity testing. In each case between nine and 12 clones were analysed. In the two patients from whom virus had greater than eight-fold reduction in susceptibility, a point mutation was observed in the viral proteinase (Leu90--> Met/Ile). Further mutations were observed at residues 36, 71 and 84 in these subjects. In a third patient, in whom an eight-fold increase in HIV IC50 of saquinavir was observed, no mutations were detected in the proteinase; sequencing of proteinase cleavage sites in viral gag-pol revealed no significant mutations. In no patient was a Gly48-->Val mutation observed, although this has been associated with resistance in vitro. The Leu90-->Met mutation was observed in five subjects, but a greater than eight-fold phenotypic change in antiviral susceptibility was seen in only two of these. Hence, in vivo, the Leu90-->Met but not the Gly48-->Val mutation is necessary, but not sufficient, for phenotypic resistance to saquinavir in HIV.

Effects of diflubenzuron and clanfenur on mouse bone marrow cells.

Diflubenzuron (DFB) and Clanfenur (CFN) belong to a group of compounds called Benzoylphenyl Ureas (BPUs). Several BPUs regulate cell growth in insects and/or inhibit growth of B-16 murine melanomas. In view of potential clinical use for these compounds, DFB and CFN were selected as examples of BPUs and tested for effects on hematopoiesis in C57Bl/6 mice housed in a conventional environment. DFB and CFN exhibit anti-tumor activity in mice, cause little or no morbidity and mortality and rather than causing bone marrow suppression, which is usual for anti-cancer drugs, these agents stimulate hematopoiesis in vivo and in vitro. Stimulation in vivo was evidenced by increased (up to 112%) peripheral blood granulocytes 6 days after a single injection and enhanced granulopoiesis (approximately 25%) in bone marrow up to 18 days after treatment. That effects of DFB and CFN were on hematopoietic stem cells were indicated by 47% and 48%, respectively, increases in numbers of CFUs and 97% and 95%, respectively, increases in CFUgm. Further, bone marrow cells treated in vitro contained about twice the number of CFUs and CFUgm as control bone marrow cells. Almost all of the increase in number of spleen colonies, whether derived from donors treated in vivo or bone marrow cells treated in vitro, was accounted for by a corresponding increase in number of undifferentiated colonies. These data indicate that DFB and CFN treatment enhance numbers of pluripotential stem cells both in vivo and in vitro. The mechanism of enhancement, direct or indirect, remains to be determined.

Parathyroid hormone-related peptide production and action in a myoepithelial cell line derived from normal human breast.

Physiological roles for PTH-related peptide (PTHrP) appear varied, but remain to be clarified. The peptide is present in large amounts in milk, and PTHrP mRNA has been shown to be present in high amounts in lactating mammary gland. Because PTHrP can cause smooth muscle relaxation, we hypothesized that the peptide might affect the contractility of the breast myoepithelial cell and thereby affect milk ejection. To test this idea, we asked whether PTHrP might affect second messenger responses in a human mammary gland myoepithelial-cell line (Hs578Bst) derived originally from normal breast tissue. To verify the presumed origin of these cells, we also examined the effects of oxytocin. Cells were grown in culture in multiwell plates and exposed to test peptides for 15 min in buffer containing 1 mM isobutylmethylxanthine, and intracellular cAMP was measured by RIA. Both PTHrP-(1-34) and PTH-(1-34) increased cAMP in a dose-related fashion (ED50, 5 nM), with a maximal effect (3-fold) occurring at 100 nM. The ability of PTHrP to stimulate cAMP was inhibited by a 10- to 100-fold molar excess of the specific inhibitors, PTH-(3-34) or PTHrP-(7-34). Inhibitors alone did not alter cAMP. Oxytocin also produced an increase in cAMP, but the effect was inconsistent and occurred only with high doses (0.1-1 microM). Using cells grown on coverslips and loaded with fura-2AM, intracellular Ca2+ was monitored in cells exposed to test peptides. Oxytocin (0.2-20 nM) produced rapid dose-related increases in intracellular Ca2+, with a peak and plateau characteristic of initial mobilization of intracellular Ca2+, followed by entry of extracellular Ca. The plateau was eliminated by the Ca channel antagonist La3+ or by perfusion of cells with Ca-free medium. PTHrP (10-100 nM) altered the intracellular Ca2+ response to oxytocin in 66% of 39 preparations tested. PTHrP inhibited the Ca2+ response when given before oxytocin or transiently decreased the plateau phase of the Ca2+ response when given after oxytocin. Analysis of cellular mRNA using reverse transcription polymerase chain reaction indicated that these cells express the gene for PTHrP, and immunohistochemistry using antiserum to PTHrP revealed positive staining of cells. Measurement of immunoreactive PTHrP in conditioned medium confirmed that these cells can synthesize and secrete the peptide. The finding of a response of this cell line to oxytocin provides functional evidence of their myoepithelial derivation.(ABSTRACT TRUNCATED AT 400 WORDS)

Reproducibility of cerebral artery Doppler measurements.

Intraobserver and interobserver variability were calculated for the measurement of cerebral artery pulsatility indices by five observers who made two recordings on each of 12 stable preterm infants. Variations in pulsatility indices of up to 0.21 were found; no measurement was below 0.55.

A novel presentation of Coxsackie B2 virus infection during pregnancy.

A 39-year-old woman presented at 34 weeks gestation with a febrile illness. This initially settled but recurred 6 days later, when uncontrolled vaginal bleeding necessitated caesarian section. At operation, vesicles were seen on the mother's hyperaemic uterus. The infant developed tachypnoea, bradycardia, cerebral irritation, an enlarged liver and spleen, intravascular coagulation and a transient rash. Mother and child made a full recovery. Coxsackie B2 virus was isolated from the infant and rises in the titre of Coxsackie B virus-specific IgM were detected in mother and child.