Assessment of fetal growth trajectory identifies infants at high risk of perinatal mortality.

OBJECTIVE: To analyze perinatal risks associated with three distinct scenarios of fetal growth trajectory in the latter half of pregnancy compared with a reference group. METHODS: This cohort study included women with a singleton pregnancy that delivered between 32 + 0 and 41 + 6 weeks' gestation and had two or more ultrasound scans, at least 4 weeks apart, from 18 + 0 weeks. We evaluated three different scenarios of fetal growth against a reference group, which comprised appropriate-for-gestational-age fetuses with appropriate forward-growth trajectory. The comparator growth trajectories were categorized as: Group 1, small-for-gestational-age (SGA) fetuses (estimated fetal weight (EFW) or abdominal circumference (AC) persistently < 10th centile) with appropriate forward growth; Group 2, fetuses with decreased growth trajectory (decrease of ≥ 50 centiles) and EFW or AC ≥ 10th centile (i.e. non-SGA) at their final ultrasound scan; and Group 3, fetuses with decreased growth trajectory and EFW or AC < 10th centile (i.e. SGA) at their final scan. The primary outcome was overall perinatal mortality (stillbirth or neonatal death). Secondary outcomes included stillbirth, delivery of a SGA infant, preterm birth, emergency Cesarean section for non-reassuring fetal status and composite severe neonatal morbidity. Associations were analyzed using logistic regression. RESULTS: The final study cohort comprised 5319 pregnancies. Compared to the reference group, the adjusted odds of perinatal mortality were increased significantly in Group 2 (adjusted odds ratio (aOR), 4.00 (95% CI, 1.36-11.22)) and Group 3 (aOR, 7.71 (95% CI, 2.39-24.91)). Only Group 3 had increased odds of stillbirth (aOR, 5.69 (95% CI, 1.55-20.93)). In contrast, infants in Group 1 did not have significantly increased odds of demise. The odds of a SGA infant at birth were increased in all three groups compared with the reference group, but was highest in Group 1 (aOR, 111.86 (95% CI, 62.58-199.95)) and Group 3 (aOR, 40.63 (95% CI, 29.01-56.92)). In both groups, more than 80% of infants were born SGA and nearly half had a birth weight < 3rd centile. Likewise, the odds of preterm birth were increased in all three groups compared with the reference group, being highest in Group 3, with an aOR of 4.27 (95% CI, 3.23-5.64). Lastly, the odds of composite severe neonatal morbidity were increased in Groups 1 and 3, whereas the odds of emergency Cesarean section for non-reassuring fetal status were increased only in Group 3. CONCLUSION: Assessing the fetal growth trajectory in the latter half of pregnancy can help identify infants at increased risk of perinatal mortality and birth weight < 3rd centile for gestation. © 2024 International Society of Ultrasound in Obstetrics and Gynecology.

Effect of reverse osmosis and ultra-high-pressure homogenization on the composition and microstructure of sweet buttermilk.

Buttermilk (BM), the by-product of butter making, is similar to skim milk (SM) composition. However, it is currently undervalued in dairy processing because it is responsible for texture defects (e.g., crumbliness, decreased firmness) in cheese and yogurt. One possible way of improving the incorporation of BM into dairy products is by the use of technological pretreatments such as membrane filtration and homogenization. The study aimed at characterizing the effect of preconcentration by reverse osmosis (RO) and single-pass ultra-high-pressure homogenization (UHPH) on the composition and microstructure of sweet BM to modify its techno-functional properties (e.g., protein gel formation, syneresis, firmness). The BM and RO BM were treated at 0, 15, 150, and 300 MPa. Pressure-treated and control BM and RO BM were ultracentrifuged to fractionate them into the following 3 fractions: a supernatant soluble fraction (top layer), a colloidal fraction consisting of a cloudy layer (middle layer), and a high-density pellet (bottom layer). Compositional changes in the soluble fraction [lipid, phospholipid (PL), protein, and salt], as well as its protein profile by PAGE analysis, were determined. Modifications in particle size distribution upon UHPH were monitored by laser diffraction in the presence and absence of sodium citrate to dissociate the casein (CN) micelles. Microstructural changes in pressure-treated and non-pressure-treated BM and RO BM particles were monitored by confocal laser scanning microscopy. Particle size analysis showed that UHPH treatment significantly decreased the size of the milk fat globule membrane fragments in BM and RO BM. Also, pressure treatment at 300 MPa led to a significant increase in the recovery of total lipids, CN, calcium, and phosphate in the BM soluble fraction (top layer) following ultracentrifugation. However, PL were primarily concentrated in the pellet cloud (middle layer), located above the pellet in BM concentrated by RO. In contrast, PL were evenly distributed between soluble and colloidal phases of BM. This study provides insight into the modifications of sweet BM constituents induced by RO and UHPH from a compositional and structural perspective.

Endoglin is required in Pax3-derived cells for embryonic blood vessel formation.

Mutations in endoglin, a TGFß/BMP coreceptor, are causal for hereditary hemorrhagic telangiectasia (HHT). Endoglin-null (Eng-/-) mouse embryos die at embryonic day (E)10.5-11.5 due to defects in angiogenesis. In part, this is due to an absence of vascular smooth muscle cell differentiation and vessel investment. Prior studies from our lab and others have shown the importance of endoglin expression in embryonic development in both endothelial cells and neural crest stem cells. These studies support the hypothesis that endoglin may play cell-autonomous roles in endothelial and vascular smooth muscle cell precursors. However, the requirement for endoglin in vascular cell precursors remains poorly defined. Our objective was to specifically delete endoglin in neural crest- and somite-derived Pax3-positive vascular precursors to understand the impact on somite progenitor cell contribution to embryonic vascular development. Pax3Cre mice were crossed with Eng+/- mice to obtain compound mutant Pax3(Cre/+);Eng+/- mice. These mice were then crossed with homozygous endoglin LoxP-mutated (Eng(LoxP/LoxP)) mice to conditionally delete the endoglin gene in specific lineages that contribute to endothelial and smooth muscle constituents of developing embryonic vessels. Pax3(Cre/+);Eng(LoxP/)(-) mice showed a variety of vascular defects at E10.5, and none of these mice survived past E12.5. Embryos analyzed at E10.5 showed malformations suggestive of misdirection of the intersomitic vessels. The dorsal aorta showed significant dilation with associated vascular smooth muscle cells exhibiting disorganization and enhanced expression of smooth muscle differentiation proteins, including smooth muscle actin. These results demonstrate a requirement for endoglin in descendants of Pax3-expressing vascular cell precursors, and thus provides new insight into the cellular basis underlying adult vascular diseases such as HHT.

Abnormally invasive placenta-prevalence, risk factors and antenatal suspicion: results from a large population-based pregnancy cohort study in the Nordic countries.

OBJECTIVE: The objective was to investigate prevalence, estimate risk factors, and antenatal suspicion of abnormally invasive placenta (AIP) associated with laparotomy in women in the Nordic countries. DESIGN: Population-based cohort study. SETTING AND POPULATION: A 3-year Nordic collaboration among obstetricians to identify and report on uterine rupture, peripartum hysterectomy, excessive blood loss, and AIP from 2009 to 2012 The Nordic Obstetric Surveillance Study (NOSS). METHODS: In the NOSS study, clinicians reported AIP cases from maternity wards and the data were validated against National health registries. MAIN OUTCOME MEASURES: Prevalence, risk factors, antenatal suspicion, birth complications, and risk estimations using aggregated national data. RESULTS: A total of 205 cases of AIP in association with laparotomy were identified, representing 3.4 per 10 000 deliveries. The single most important risk factor, which was reported in 49% of all cases of AIP, was placenta praevia. The risk of AIP increased seven-fold after one prior caesarean section (CS) to 56-fold after three or more CS. Prior postpartum haemorrhage was associated with six-fold increased risk of AIP (95% confidence interval 3.7-10.9). Approximately 70% of all cases were not diagnosed antepartum. Of these, 39% had prior CS and 33% had placenta praevia. CONCLUSION: Our findings indicate that a lower CS rate in the population may be the most effective way to lower the incidence of AIP. Focused ultrasound assessment of women at high risk will likely strengthen antenatal suspicion. Prior PPH is a novel risk factor associated with an increased prevalence of AIP. TWEETABLE ABSTRACT: An ultrasound assessment in women with placenta praevia or prior CS may double the awareness for AIP.

Long-term effects of prenatal progesterone exposure: neurophysiological development and hospital admissions in twins up to 8 years of age.

OBJECTIVES: To perform a neurophysiological follow-up at 48 or 60 months of age in children exposed prenatally to progesterone compared with a placebo and evaluate their medical histories up to 8 years of age. METHODS: In this study, Danish participants of the PREDICT study, including 989 surviving children from 498 twin pregnancies, were followed-up. PREDICT was a placebo-controlled randomized clinical trial examining the effect of progesterone for prevention of preterm delivery in unselected twin pregnancies. Medical histories of the children were reviewed and neurophysiological development was evaluated by the parent-completed Ages and Stages Questionnaire (ASQ) at either 48 or 60 months after the estimated date of delivery. We used the method of generalized estimating equation to account for the correlation within twins. RESULTS: A total of 492 children had been exposed prenatally to progesterone and 497 to placebo. There was no difference in the number of admissions to or length of stay in hospital between the treatment groups, and we found no overall difference in the rates of diagnoses made. However, the odds ratios (ORs) for a diagnosis concerning the heart was 1.66 (95% CI, 0.81-3.37), favoring placebo, among all children, 2.38 (95% CI, 1.07-5.30) in dichorionic twins and 8.19 (95% CI, 1.02-65.6) in all children when excluding diagnoses made at outpatient clinic visits. ASQ scores were available for 437 children (progesterone, n = 225; placebo, n = 212). Mean ASQ score was slightly higher in the progesterone group compared with the placebo group (P = 0.03). In dichorionic twins, the risk of having a low ASQ score (< 10(th) centile) was decreased in the progesterone group (OR, 0.34 (95% CI, 0.14-0.86)). CONCLUSION: Second- and third-trimester exposure of the fetus to progesterone does not seem to have long-term harmful effects during childhood, but future studies should focus on cardiac disease in the child. Copyright © 2016 ISUOG. Published by John Wiley & Sons Ltd.

The personality trait self-directedness predicts the amygdala's reaction to appetizing cues in fMRI.

Personality and neural response to food cues in various mesolimbic brain structures have been linked to eating disorders. We investigated the question of whether personality traits in healthy individuals correlate with the brain activation induced on confrontation with appetizing visual stimuli. Personality was assessed in 27 normal-weight participants (14 women, mean age=26.0, SD=3.3 years) with the Temperament and Character Inventory (TCI). After an overnight fast, participants viewed blocks of pictures, half containing appetizing food and the other half showing scrambled pictures as control. After each block, participants rated their appetite. Brain activation was measured using a 3T MR scanner. Food compared to control stimuli elicited a significantly higher appetite rating, as well as strong activation in the ventral and dorsal visual stream, the fusiform gyrus and consecutive limbic centres such as the parahippocampal gyrus, the amygdala, the thalamus, the insula, the ventral striatum and the orbitofrontal cortex. In a region-of-interest analysis, the TCI trait self-directedness was negatively correlated with mean blood oxygenation level dependent (BOLD) signal change in the right amygdala (r=-.43, p=.025). Ultimately, amygdala reactivity might provide a risk factor for the development of eating disorders.

Interobserver agreement in analysis of cardiotocograms recorded during trial of labor after cesarean.

Introduction: To examine interobserver agreement in intrapartum cardiotocography (CTG) classification in women undergoing trial of labor after a cesarean section (TOLAC) at term with or without complete uterine rupture. Materials and methods: Nineteen blinded and independent Danish obstetricians assessed CTG tracings from 47 women (174 individual pages) with a complete uterine rupture during TOLAC and 37 women (133 individual pages) with no uterine rupture during TOLAC. Individual pages with CTG tracings lasting at least 20 min were evaluated by three different assessors and counted as an individual case. The tracings were analyzed according to the modified version of the Federation of Gynaecology and Obstetrics (FIGO) guidelines elaborated for the use of STAN (ST-analysis). Occurrence of defined abnormalities was recorded and the tracings were classified as normal, suspicious, pathological, or preterminal. The interobserver agreement was evaluated using Fleiss' kappa. Results: Agreement on classification of a preterminal CTG was almost perfect. The interobserver agreement on normal, suspicious or pathological CTG was moderate to substantial. Regarding the presence of severe variable decelerations, the agreement was moderate. No statistical difference was found in the interobserver agreement between classification of tracings from women undergoing TOLAC with and without complete uterine rupture. Conclusions: The interobserver agreement on classification of CTG tracings from high-risk deliveries during TOLAC is best for assessment of a preterminal CTG and the poorest for the identification of severe variable decelerations.

[Reirradiation of head and neck cancers]. / Réirradiation des cancers des voies aérodigestives supérieures.

Locoregional relapse in previously irradiated region for head and neck tumours is associated with a bad locoregional and distant prognosis. Reirradiation might be exclusive, or feasible in addition with surgery and/or chemotherapy, according to histopronostic factors. Data show that reirradiation is feasible with some severe toxicity due to the bad prognosis of this situation. Hyperfractionnated regimen with split course or normofractionnated regimen without split course are possible with similar efficacy. If tumour size is small, stereotactic ablative radiotherapy may be considered, and if the treatment centre has proton therapy, it could be proposed because of better organs at risk sparing. There is no standard regarding reirradiation schedules and several trials have to be done in order to determine the best technique. Nevertheless, it is agreed that a total dose of 60Gy (2Gy per fraction) is needed. Other trials testing the association with new systemic agents have to be performed, among them agents targeting the PD1/PD-L1 axis.

Regulation of targeted chemotherapy with cytotoxic lutenizing hormone-releasing hormone analogue by epidermal growth factor.

Targeting chemotherapy selectively to cancers can reduce the toxic side effects. AN-152, a conjugate of doxorubicin and [D-Lys6]-luteinizing hormone-releasing hormone (LH-RH), is more potent against LH-RH receptor-bearing cancers and produces less peripheral toxicity than doxorubicin. Many cancers, e.g., 50% of breast cancers, but few normal tissues express these receptors, providing a selective target for this cytotoxic conjugate. In this study, the effectiveness of AN-152 was heightened by receptor up-regulation. The cytotoxic effect of AN-152 can be regulated by the number of active LH-RH receptors on cancer cells. LH-RH receptor-positive (MCF-7) and -negative (UCI-107) cancer cells were treated with epidermal growth factor (EGF) or the somatostatin analogue, RC-160. EGF and RC-160 have been shown previously to regulate LH-RH receptors through phosphorylation. The effect of receptor regulation, by hormone exposure, on the cytotoxicity of AN-152 and doxorubicin and on the cellular uptake of AN-152, [D-Lys6]LH-RH, or doxorubicin was assessed by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and by two-photon laser scanning microscopy. The results demonstrated that the cellular entry of the conjugate was: (a) specific for cancers with LH-RH receptors; (b) up-regulated by EGF; (c) down-regulated by RC-160; and (d) the cytotoxicity of the AN-152 paralleled the efficiency of entry. This study illustrates the potential use of receptor regulation for increasing the efficacy of chemotherapeutic approaches that are directed to cell surface receptors.

Placebo-blinded study of morphine sulfate sustained-release tablets and immediate-release morphine sulfate solution in outpatients with chronic pain due to advanced cancer.

PURPOSE: This study was conducted to compare the relative analgesic efficacy and safety of an every-4-hour immediate-release oral morphine (IRM) solution with that of an every-12-hour sustained-release oral morphine (SRM) formulation. PATIENTS AND METHODS: This was a double-blind, placebo-blinded, crossover study in 34 adult male and female outpatients with pain due to advanced cancer. Baseline data were collected on day 1. On days 2 and 3, randomly assigned patients received either placebo plus IRM (Roxanol; Roxane Laboratories, Inc, Columbus, OH; 20 mg/mL) at 2, 6, and 10 am, and 2, 6, and 10 pm, or alternatively SRM (Oramorph SR; Roxane Laboratories, Inc; 30 mg) at 10 AM and 10 PM. Patients were then crossed over to the alternate treatment for days 4 through 6. Pain relief was measured using a conventional 100-mm visual analog scale (VAS) and by recording the incidence of breakthrough pain. Information on side effects was obtained from VAS scores for sedation, nausea, anxiety, and depression; by directly questioning the patient as to mental confusion, bowel movements, and laxative use; and from Karnofsky performance status scores. VAS data were analyzed using a linear statistical model. Breakthrough pain data were analyzed using analysis of variance (ANOVA). RESULTS: There were no statistically significant differences between IRM and SRM treatments with respect to VAS pain scores, side effect scores, or incidence of breakthrough pain data. Karnofsky performance scores remained stable for all patients throughout the study. CONCLUSION: It was concluded that every-12-hour administration of SRM and every-4-hour administration of IRM provide similar analgesic effectiveness and side effect profiles in the treatment of chronic pain in cancer patients.

[Management of locally advanced anal canal carcinoma with modulated arctherapy and concurrent chemotherapy]. / Place de l'arcthérapie modulée et de la chimiothérapie concomitante dans la prise en charge des cancers du canal anal localement évolués.

The standard treatment of locally advanced (stage II and III) squamous cell carcinoma of the anal canal consists of concurrent chemoradiotherapy (two cycles of 5-fluoro-uracil, mitomycin C, on a 28-day cycle), with a dose of 45 Gy in 1.8 Gy per fraction in the prophylactic planning target volume and additional 14 to 20 Gy in the boost planning target volume (5 days per week) with a possibility of 15 days gap period between the two sequences. While conformal irradiation may only yield suboptimal tumor coverage using complex photon/electron field junctions (especially on nodal areas), intensity modulated radiation therapy techniques (segmented static, dynamic, volumetric modulated arc therapy and helical tomotherapy) allow better tumour coverage while sparing organs at risk from intermediate/high doses (small intestine, perineum/genitalia, bladder, pelvic bone, etc.). Such dosimetric advantages result in fewer severe acute toxicities and better potential to avoid a prolonged treatment break that increases risk of local failure. These techniques also allow a reduction in late gastrointestinal and skin toxicities of grade 3 or above, as well as better functional conservation of anorectal sphincter. The technical achievements (simulation, contouring, prescription dose, treatment planning, control quality) of volumetric modulated arctherapy are discussed.

Attitudes of Colorado health professionals toward breast and cervical cancer screening in Hispanic women.

BACKGROUND: A variety of economic, cultural, and communication barriers appear to be involved in breast and cervical cancer screening among Hispanic women. These barriers include culture-based embarrassment both for mammography and for Pap smears and fear and hopelessness concerning a diagnosis of cancer. Cost and access barriers are shared by low-income women from various ethnic and racial groups, as is a purported lack of physician referral. Hispanic women may have the latter problem enhanced by a language barrier between physicians and patients when the physicians do not speak or understand Spanish. PURPOSE: The goal of this project, conducted by the Cancer Education Division of the University of Colorado Cancer Center, has been to determine the attitudes and practices among health care providers in areas of Colorado with relatively large Hispanic populations (concerning screening mammography, clinical breast examination, breast self-examination, and Pap testing) and to design interventions to address any deficiencies or problems recognized. These studies were coordinated with telephone surveys and focus groups involving Hispanic women, directed by E. Flores in the Department of Sociology of the University of Colorado at Boulder and by C. Chrvala at the Colorado Department of Health. METHODS: Data were collected from 520 primary care physicians, nurses, and allied health personnel in 11 Colorado counties through focus groups and mailed questionnaires. Responses were analyzed by considering a variety of demographic characteristics of the respondents and by stratifying the associated practices by percent of Hispanic patients. RESULTS: The physicians involved in the focus groups and responding to the questionnaires, as well as their associated nurses and other health care personnel, are generally familiar with the breast and cervical cancer-screening guidelines as developed and disseminated by several organizations, including the National Cancer Institute and the American Cancer Society. Major barriers to screening Hispanic women, as perceived by these health care providers, appear to be cost; lack of transportation, child care, and release from work; fear of diagnosis of cancer; patients considering the test unnecessary; discomfort; and embarrassment. The prompt use of colposcopy to evaluate patients whose Pap smears indicated dysplasia appeared less than optimal, especially among internists. CONCLUSIONS: Familiarity with guidelines for breast and cervical cancer screening is widespread among Colorado physicians and associated health care personnel, including those with high percentages of Hispanic patients in their practices. Increased continuing education efforts may be indicated concerning the application of colposcopy to the evaluation of women with abnormal Pap smears and concerning the application of computer technology to cancer-screening reminder systems. IMPLICATIONS: Educational approaches to primary care professionals may improve the effectiveness of breast and cervical cancer screening, although a variety of other approaches will also be necessary to decrease barriers to screening of Hispanic women.

Measles immunity among community hospital employees.

OBJECTIVE: To define measles immunity rates among employees at 2 hospitals during a community outbreak in 1990. DESIGN: Cohort survey using enzyme-linked immunosorbent assay (ELISA) and questionnaire. SETTING: Two community hospitals. PARTICIPANTS: Seventy-six percent of 2,060 employees. RESULTS: Seven percent (115/1566) of participants lacked ELISA-defined measles immunity. Among employees whose ages were known, 14% (64/467) of those born after 1956 and 5% (50/1086) of those born before 1957 lacked serologic evidence of immunity. Fifty-eight percent of the susceptible persons had substantial patient contact. With ELISA results as the reference for immunity, the predictive value of an undocumented positive history of measles disease or vaccination was 95%; the predictive value of a negative history of both was 52%. Measles developed in 7 employees. CONCLUSIONS: A substantial number of hospital employees lacked ELISA-defined measles immunity, including many who had patient contact or who had been born before 1957. Undocumented disease and vaccination histories were not adequate predictors of serologic status. This study supports the recommendations and suggestions of the Immunization Practices Advisory Committee that hospitals should require documented evidence of measles immunity from employees who have patient contact.

Molecular basis of hyperparathyroidism and potential targets for drug development.

Our appreciation of the molecular pathogenesis of primary hyperparathyroidism (HPT) has seen great advances over the past decade. This improved understanding may well lead to the development of new treatment options that are specifically targeted to defective pathways. This review summarizes recent advances in the molecular basis of HPT and associated endocrinopathies, and discusses the potential for these and future findings to provide targets for alternative approaches to therapy. The only proven contributors to common sporadic HPT, by virtue of clonal genetic abnormalities, are the cyclin D1 and MEN1 genes. Cyclin D1 is an oncogene that encodes a key regulator of the cell cycle, while MEN1 is a tumor suppressor gene that has also been implicated in familial multiple endocrine neoplasia type 1 (MEN1), in which primary HPT is common. In addition, other key parathyroid regulatory pathways may play a role in HPT pathogenesis. 1,25 (OH)2-vitamin D. Ca2+ and phosphate are regarded as principal regulators of parathyroid cell proliferation and PTH secretion. Therefore, prime candidate targets include the Ca2+ sensing receptor (CASR) gene, the vitamin D receptor (VDR) gene, a putative phosphate receptor gene, their cognate gene products, and other genes or proteins involved in their respective biochemical pathways. Attempts to identify new therapies based specifically on the defective pathways in HPT could complement or eventually supplant traditional approaches.

A controlled study of amniotic fluid immunoglobulin levels in intraamniotic infection.

The authors studied the levels of immunoglobulins (Ig) in the amniotic fluid of 46 patients with intraamniotic infection and of 46 matched controls. Amniotic fluid was collected through a transcervical intrauterine catheter. All infected patients had clinical signs of intraamniotic infection and at least 10(2) colony-forming units/milliliter of a high-virulence organism. None of the controls became infected. The matching criteria were gestational age, labor, interval from rupture of the membranes to delivery, and interval from rupture of the membranes to amniotic fluid collection. The authors tested each amniotic fluid sample for immunoglobulins G, M, and A by a single radial immunodiffusion technique. The mean IgG level in amniotic fluid for the intraamniotic infection group was 33.9 +/- 38.5 mg/dl, and the mean level for the control group was 17.9 +/- 11.1 mg/dl. The authors concluded that the mean IgG level in amniotic fluid from patients with intraamniotic infection is significantly higher than in controls (P less than .02).

AT4 receptor binding characteristics: D-amino acid- and glycine-substituted peptides.

The ability of angiotensin IV (AIV) analogs to compete for [125I]AIV binding in heat-treated bovine adrenal membranes was examined. Angiotensin IV displayed a Ki of 2.63 +/- 0.12 nM. Peptides containing mono-substitutions with glycine or the corresponding D-amino acid in positions one, two, or three possessed K(i)s greater than 100 nM. Conversely, substitutions at positions four, five, and six produced peptides with Kis less than 8 nM. These data suggest that the N-terminal domains of the AIV peptide are critical for receptor binding, while the C-terminal domains play a less decisive role in receptor specificity.

Autoradiographic identification of kidney angiotensin IV binding sites and angiotensin IV-induced renal cortical blood flow changes in rats.

The present investigation initially determined that specific binding sites for the hexapeptide angiotensin IV (AngIV) are present in the rat kidney cortex and outer medulla but not in the inner medulla, using in vitro autoradiographic techniques. This binding site has been termed AT4, is distinct from the previously characterized AT1 and AT2 sites, and does not bind the specific AT1 receptor antagonist DuP753 or the AT2 receptor antagonist PD123177. Renal artery infusions of AngIV produced a dose-dependent increase in cortical blood flow without altering systemic blood pressure. In contrast, the infusion of angiotensin II (AngII) induced a dramatic decrease in cortical blood flow, accompanied by a significant elevation in systemic blood pressure. The infusion of [D-Val(1)]AngIV, an analog that does not bind at the AT4 receptor site, and the C-terminal truncated analogs AngIV (1-4) and AngIV (1-5) that possess lower affinity for this site, produced no change in cortical blood flow. The infusion of [Nle1]AngIV and [Lys1]AngIV, analogs that bind with high affinity at the AT4 receptor site, produced increases in cortical blood flow with no influence on blood pressure. Pretreatment with a specific AT4 receptor antagonist, Divalinal-AngIV, completely blocked AngIV-induced elevations in blood flow, but failed to influence AngII-induced decreases in blood flow, suggesting that these ligands are acting at different receptor sites. Pretreatment with the nitric oxide synthase inhibitor, NG-Monomethyl-L-Arginine, also blocked subsequent AngIV-induced increases in cortical blood flow. These data support the notion that AngIV exerts a unique influence upon renal hemodynamics via the AT4 receptor subtype, and suggest that AngIV-induced elevations in blood flow may be mediated by nitric oxide.

Characterization of the binding properties and physiological action of divalinal-angiotensin IV, a putative AT4 receptor antagonist.

Divalinal-Ang IV [V psi (CH2-NH2)YV psi (CH2-NH2)HPF] is being employed increasingly as a specific AT4 antagonist. This use, which necessitates a comprehensive physiological and pharmacological evaluation of Divalinal-Ang IV's functional and receptor binding characteristics in order to ensure its efficacy and specificity, was the stimulus for this study using bovine adrenal membranes. [125I]Ang IV and [125I]Divalinal-Ang IV were shown to bind with high affinity to a similar number of binding sites, suggesting that both bound the same receptor. This notion was verified by competition curves using [125I]Ang IV and [125I]Divalinal-Ang IV that indicated identical rank order affinities for several angiotensin-related peptides and 100% cross-displacement by Ang IV and Divalinal-Ang IV. Furthermore, an autoradiographic comparison of [125I]Ang IV and [125I]Divalinal-Ang IV in 20 microns sections of bovine adrenals revealed near identical binding distributions characterized by heavy binding in the glomerulosa layer and the medulla. Physiological studies in which test compounds were injected into the internal carotid of the rat and cerebral blood flor (CBF) was measured by laser Doppler flowmetry indicated that pretreatment with Divalinal-Ang IV, but not DuP 753 or PD123177, blocked the increased flow observed with Ang IV infusion. Conversely, DuP 753, but not Divalinal-Ang IV or PD123177, inhibited the decrease in flow witnessed with Ang II. Metabolic stability studies utilizing rat kidney homogenates as a peptidase source, demonstrated that the structural changes present in Divalinal-Ang IV greatly increased its resistance to metabolism as compared to Ang IV. Together, these studies show that Divalinal-Ang IV is a stable, efficacious and specific inhibitor of AT4 receptors.

Autoradiographic identification of brain angiotensin IV binding sites and differential c-Fos expression following intracerebroventricular injection of angiotensin II and IV in rats.

A unique angiotensin binding site specific for the hexapeptide, angiotensin II(3-8) (AngIV), has been previously reported by our laboratory in the guinea pig brain and is presently described in the rat brain. This angiotensin receptor subtype has been termed AT4 and is prominently distributed in cerebral cortex, piriform cortex, hippocampus, habenulae, colliculi, septum, periaqueductal gray, several thalamic nuclei, the arcuate nucleus of the hypothalamus and cerebellum. In the second part of the present investigation, separate groups of rats received i.c.v. injections of angiotensin II (AngII), AngIV or artificial cerebrospinal fluid (aCSF) and were euthanized 2 h later for the purpose of evaluating for brain c-Fos expression. After i.c.v.-injected AngIV, Fos-like immunoreactivity was present in the hippocampus and piriform cortex. This immunoreactivity was unaffected by i.c.v. pretreatment with the AT1 angiotensin receptor antagonist DuP 753 (losartan) or the AT2 receptor ligand PD123177 but was blocked by the AT4 angiotensin receptor antagonist, divalanal-AngIV. I.c.v. injection of AngII resulted in Fos-like immunoreactivity in the dorsal third and lateral ventricles, subfornical organ, lateral hypothalamus and amygdala. Pretreatment with losartan or PD123177 significantly interfered with this AngII-induced immunoreactivity while divalanal-AngIV did not. These results indicate that in both guinea pig and rat brains the AT4 receptor has a distribution different than that previously reported for AT1 and AT2 receptor subtypes. The c-Fos expression results suggest that different brain neuronal pathways are activated by i.c.v. injection of AngII and AngIV.