Pulmonary arterial hypertension - progress in understanding the disease and prioritizing strategies for drug development.

Pulmonary arterial hypertension (PAH), at one time a largely overlooked disease, is now the subject of intense study in many academic and biotech groups. The availability of new treatments has increased awareness of the condition. This in turn has driven a change in the demographics of PAH, with an increase in the mean age at diagnosis. The diagnosis of PAH in more elderly patients has highlighted the need for careful phenotyping of patients and for further studies to understand how best to manage pulmonary hypertension associated with, for example, left heart disease. The breadth and depth of expertise focused on unravelling the molecular pathology of PAH has yielded novel insights, including the role of growth factors, inflammation and metabolic remodelling. The description of the genetic architecture of PAH is accelerating in parallel, with novel variants, such as those reported in potassium two-pore domain channel subfamily K member 3 (KCNK3), adding to the list of more established mutations in genes associated with bone morphogenetic protein receptor type 2 (BMPR2) signalling. These insights have supported a paradigm shift in treatment strategies away from simply addressing the imbalance of vasoactive mediators observed in PAH towards tackling more directly the structural remodelling of the pulmonary vasculature. Here, we summarize the changing clinical and molecular landscape of PAH. We highlight novel drug therapies that are in various stages of clinical development, targeting for example cell proliferation, metabolic, inflammatory/immune and BMPR2 dysfunction, and the challenges around developing these treatments. We argue that advances in the treatment of PAH will come through deep molecular phenotyping with the integration of clinical, genomic, transcriptomic, proteomic and metabolomic information in large populations of patients through international collaboration. This approach provides the best opportunity for identifying key signalling pathways, both as potential drug targets and as biomarkers for patient selection. The expectation is that together these will enable the prioritization of potential therapies in development and the evolution of personalized medicine for PAH.

Single TRAM domain RNA-binding proteins in Archaea: functional insight from Ctr3 from the Antarctic methanogen Methanococcoides burtonii.

TRAM domain proteins present in Archaea and Bacteria have a ß-barrel shape with anti-parallel ß-sheets that form a nucleic acid binding surface; a structure also present in cold shock proteins (Csps). Aside from protein structures, experimental data defining the function of TRAM domains is lacking. Here, we explore the possible functional properties of a single TRAM domain protein, Ctr3 (cold-responsive TRAM domain protein 3) from the Antarctic archaeon Methanococcoides burtonii that has increased abundance during low temperature growth. Ribonucleic acid (RNA) bound by Ctr3 in vitro was determined using RNA-seq. Ctr3-bound M. burtonii RNA with a preference for transfer (t)RNA and 5S ribosomal RNA, and a potential binding motif was identified. In tRNA, the motif represented the C loop; a region that is conserved in tRNA from all domains of life and appears to be solvent exposed, potentially providing access for Ctr3 to bind. Ctr3 and Csps are structurally similar and are both inferred to function in low temperature translation. The broad representation of single TRAM domain proteins within Archaea compared with their apparent absence in Bacteria, and scarcity of Csps in Archaea but prevalence in Bacteria, suggests they represent distinct evolutionary lineages of functionally equivalent RNA-binding proteins.

Genome-wide differentiation in closely related populations: the roles of selection and geographic isolation.

Population divergence in geographic isolation is due to a combination of factors. Natural and sexual selection may be important in shaping patterns of population differentiation, a pattern referred to as 'isolation by adaptation' (IBA). IBA can be complementary to the well-known pattern of 'isolation by distance' (IBD), in which the divergence of closely related populations (via any evolutionary process) is associated with geographic isolation. The barn swallow Hirundo rustica complex comprises six closely related subspecies, where divergent sexual selection is associated with phenotypic differentiation among allopatric populations. To investigate the relative contributions of selection and geographic distance to genome-wide differentiation, we compared genotypic and phenotypic variation from 350 barn swallows sampled across eight populations (28 pairwise comparisons) from four different subspecies. We report a draft whole-genome sequence for H. rustica, to which we aligned a set of 9493 single nucleotide polymorphisms (SNPs). Using statistical approaches to control for spatial autocorrelation of phenotypic variables and geographic distance, we find that divergence in traits related to migratory behaviour and sexual signalling, as well as geographic distance, together explain over 70% of genome-wide divergence among populations. Controlling for IBD, we find 42% of genomewide divergence is attributable to IBA through pairwise differences in traits related to migratory behaviour and sexual signalling alone. By (i) combining these results with prior studies of how selection shapes morphological differentiation and (ii) accounting for spatial autocorrelation, we infer that morphological adaptation plays a large role in shaping population-level differentiation in this group of closely related populations.

Phenotypic differentiation is associated with divergent sexual selection among closely related barn swallow populations.

Sexual selection plays a key role in the diversification of numerous animal clades and may accelerate trait divergence during speciation. However, much of our understanding of this process comes from phylogenetic comparative studies, which rely on surrogate measures such as dimorphism that may not represent selection in wild populations. In this study, we assess sexual selection pressures for multiple male visual signals across four barn swallow (Hirundo rustica) populations. Our sample encompassed 2400 linear km and two described subspecies: European H. r. rustica (in the Czech Republic and Romania) and eastern Mediterranean H. r. transitiva (in Israel), as well as a potential area of contact (in Turkey). We demonstrate significant phenotypic differentiation in four sexual signalling axes, despite very low-level genomic divergence and no comparable divergence in an ecological trait. Moreover, the direction of phenotypic divergence is consistent with differences in sexual selection pressures among subspecies. Thus, H. r. transitiva, which have the darkest ventral plumage of any population, experience directional selection for darker plumage. Similarly, H. r. rustica, which have the longest tail feathers of any population, experience directional selection for elongated tail feathers and disruptive selection for ventral plumage saturation. These results suggest that sexual selection is the primary driver of phenotypic differentiation in this species. Our findings add to growing evidence of phenotypic divergence with gene flow. However, to our knowledge, this is the first study to relate direct measures of the strength and targets of sexual selection to phenotypic divergence among closely related wild populations.

Survival of HIV-positive individuals with hepatitis B and C infection in Michigan.

We sought to estimate mortality and associated factors in HIV-hepatitis co-infected individuals in Michigan using a retrospective cohort study. For the study period of 1 January 2006 to 31 December 2009, all HIV-infected individuals were matched to hepatitis B and C cases. In the final Cox proportional hazards regression model, individuals of other [hazard ratio (HR) 2·2, 95% confidence interval (CI) 1·4-3·2] and black (HR 1·3, 95% CI 1·1-1·6) race had decreased survival compared to white race. Similarly, injecting drug users (IDUs) (HR 2·1, 95% CI 1·6-2·6), men who have sex with men (MSM)/IDUs (HR 1·5, 95% CI 1·1-2·2), individuals with undetermined risk (HR 1·5, 95% CI 1·2-1·9) and heterosexual practices (HR 1·4, 95% CI 1·1-1·8) had decreased survival compared to MSM. Additionally, an interaction was found between current HIV status and co-infection. Mortality in HIV-hepatitis co-infected individuals remains a continuing problem. Our study can help in planning interventions to reduce mortality in HIV-infected individuals.

Hepatitis B and C co-infection in HIV/AIDS population in the state of Michigan.

A retrospective cohort study was conducted from 1 January 2006 to 31 December 2009 in Michigan to estimate the prevalence of HIV and hepatitis co-infection and identify associated factors. The prevalence of co-infection was 4.1% [95% confidence interval (CI) 3.8-4.5]. Multivariable logistic regression analysis revealed a significant association between co-infection and being male and: of Black race [odds ratio (OR) 2.0, 95% CI 1.2-3.6] and of Other race (OR 3.5, 95% CI 1.7-7.0) compared to Hispanic race. A significant association was found between co-infection and risk categories of blood products (OR 11.1, 95% CI 6.2-20.2), injecting drug user (IDU) (OR 3.6, 95% CI 2.7-4.8) and men who have sex with men/IDU (OR 3.4, 95% CI 2.4-4.9) in addition to two interactions; one between sex and current HIV status and the other between current HIV status and age at HIV diagnosis. Our results document the changing epidemiology of HIV-hepatitis co-infection which can guide preventive measures and interventions to reduce the prevalence of hepatitis co-infection.

Milk Contamination by Mycobacterium tuberculosis Complex, Implications for Public Health in Amazonas, Brazil.

ABSTRACT: In Brazil, contamination of raw milk with Mycobacterium tuberculosis complex (MTC) has been reported in several states. The highest rate of consumption of raw milk and its derivatives in Brazil occurs in Amazonas. This state also has the highest prevalence of tuberculosis in both humans and livestock. We assessed the contamination of cow's milk and buffalo's milk with MTC in Amazonas, focusing on Mycobacterium bovis, the species most commonly found in cattle and buffalo. In 2019, 250 samples of raw milk (91 from cattle, 159 from buffalo) were collected before processing from three milk plants in the state of Amazonas. The samples were placed into 21 pools and analyzed using shotgun metagenomic sequencing and taxonomic classification with Kraken 2 and MegaBLAST. To confirm the identity of mycobacterial species found, BLASTN was used to identify specific genomic positions in the TbD1 and RD1 regions and flanking RD4 region. MTC genetic material was identified in all pools of raw milk. Genetic material consistent with M. bovis was identified in seven pools of raw milk (1 from cattle, 6 from buffalo). Buffalo's milk had significantly higher MTC reads than did cow's milk. The common practice of consumption of raw milk and its derivatives in Amazonas presents a risk to public health. Urgent measures to prevent transmission of foodborne tuberculosis are needed in the Amazon region. Greater efforts and resources also should be directed toward elimination of bovine tuberculosis in cattle and buffalo herds in Amazonas and the rest of Brazil.

Iron deficiency in pulmonary arterial hypertension: a potential therapeutic target.

Iron deficiency is known to be common and detrimental in chronic left heart failure, where parenteral iron treatment has been shown to improve exercise capacity, New York Heart Association functional class and patient wellbeing. There is now increasing interest in the role of iron in the natural history of pulmonary arterial hypertension (PAH). Iron availability influences the pulmonary vasoconstrictor response to hypoxia and accumulating evidence indicates that iron deficiency is prevalent in idiopathic and heritable forms of PAH, iron status being related to exercise capacity, symptoms and poorer survival in patients with idiopathic PAH (IPAH). Potential mechanisms behind iron deficiency in IPAH include inhibition of dietary iron uptake by the master iron regulator hepcidin. High hepcidin levels underlie the anaemia of chronic disease. Possible stimuli of the observed high levels of hepcidin in IPAH include dysfunctional bone morphogenetic protein receptor type II signalling and inflammation. Iron status may influence outcomes through modulation of the pulmonary circulation as well as myocardial and skeletal muscle function. Two parallel studies, from our centre (Hammersmith Hospital, London, UK) and others in the UK and Amsterdam (the Netherlands), investigating the safety and potential benefit of iron supplementation in patients with PAH are currently under way.

Tetrahydrobiopterin and the regulation of hypoxic pulmonary vasoconstriction.

Tetrahydrobiopterin (BH(4)) is an essential cofactor for nitric oxide synthases (NOS). This study investigated the effect of increasing BH(4) levels on hypoxia-induced pulmonary vasoconstriction (HPV). Sprague Dawley rats and hph-1 (BH(4) deficient) mice were given BH(4) before and during HPV in an isolated perfused lung preparation. BH(4) inhibited HPV in a concentration-dependent manner and increased NO metabolites in the perfusate. Bradykinin-induced reductions in HPV were blunted in hph-1 mice and pre-administration of BH(4) restored the response. The effect of BH(4) was attenuated by l-NAME (NOS inhibitor), PTIO (NO scavenger), and catalase (H(2)O(2) catalyser) administered prior to HPV but enhanced by MnTMPyP (superoxide dismutase mimetic). The effect of BH(4) on HPV was partially recapitulated by NH(4), a stereoisomer that shares antioxidant properties with BH(4) but is not a NOS cofactor. The bioavailability of BH(4) is an important determinant of the pulmonary vascular response to hypoxia. Its effects are mediated via nitric oxide, hydrogen peroxide and its antioxidant properties, and are attenuated by oxidant stress. Pharmacological administration of BH(4) may have therapeutic potential in pulmonary hypertension.

Use of multiple-locus variable number tandem repeat analysis and phage typing for subtyping of Salmonella Enteritidis from sporadic human cases in the United States.

AIMS: To investigate the genetic diversity among S. Enteritidis isolates from different geographic regions to evaluate the relationship between phage types (PTs) and variable number tandem repeat analysis (VNTR) loci. METHODS AND RESULTS: We performed multiple-locus variable number tandem repeat analysis (MLVA) and phage typing on 245 S. Enteritidis isolates collected from sporadic human clinical cases in Michigan, Minnesota, New York, and Washington states between 2000 and 2007. Ninety-four MLVA types and 22 different PTs were identified. Specific PTs were associated with a predominant allele for certain VNTR loci. Cluster analysis using a minimum-spanning tree demonstrated two major clusters (I, II) and one minor cluster of isolates. PTs 8, 13a, 13 and 34 were significantly associated with MLVA cluster I. Phage types 1, 4, 6a, and 18 were significantly associated with MLVA cluster II. CONCLUSIONS: We found significant association between MLVA-based clusters and PTs. Certain VNTR loci were associated with specific PTs and could serve as useful molecular markers for S. Enteritidis in epidemiological investigations. SIGNIFICANCE AND IMPACT OF THE STUDY: MLVA genotyping in combination with phage typing can be used for effective characterization of S. Enteritidis isolates. It can also be useful for tracing possible sources during investigations of sporadic and outbreak cases of S. Enteritidis.

Ecological Assembly Processes Are Coordinated between Bacterial and Viral Communities in Fractured Shale Ecosystems.

The ecological drivers that concurrently act upon both a virus and its host and that drive community assembly are poorly understood despite known interactions between viral populations and their microbial hosts. Hydraulically fractured shale environments provide access to a closed ecosystem in the deep subsurface where constrained microbial and viral community assembly processes can be examined. Here, we used metagenomic analyses of time-resolved-produced fluid samples from two wells in the Appalachian Basin to track viral and host dynamics and to investigate community assembly processes. Hypersaline conditions within these ecosystems should drive microbial community structure to a similar configuration through time in response to common osmotic stress. However, viral predation appears to counterbalance this potentially strong homogeneous selection and pushes the microbial community toward undominated assembly. In comparison, while the viral community was also influenced by substantial undominated processes, it assembled, in part, due to homogeneous selection. When the overall assembly processes acting upon both these communities were directly compared with each other, a significant relationship was revealed, suggesting an association between microbial and viral community development despite differing selective pressures. These results reveal a potentially important balance of ecological dynamics that must be in maintained within this deep subsurface ecosystem in order for the microbial community to persist over extended time periods. More broadly, this relationship begins to provide knowledge underlying metacommunity development across trophic levels.IMPORTANCE Interactions between viral communities and their microbial hosts have been the subject of many recent studies in a wide range of ecosystems. The degree of coordination between ecological assembly processes influencing viral and microbial communities, however, has been explored to a much lesser degree. By using a combined null modeling approach, this study investigated the ecological assembly processes influencing both viral and microbial community structure within hydraulically fractured shale environments. Among other results, significant relationships between the structuring processes affecting both the viral and microbial community were observed, indicating that ecological assembly might be coordinated between these communities despite differing selective pressures. Within this deep subsurface ecosystem, these results reveal a potentially important balance of ecological dynamics that must be maintained to enable long-term microbial community persistence. More broadly, this relationship begins to provide insight into the development of communities across trophic levels.

Simvastatin and sildenafil combine to attenuate pulmonary hypertension.

Statins have been proposed to be a potential treatment for pulmonary arterial hypertension. If introduced into clinical practice, the statin would have to be used in conjunction with established therapy. We investigated the effects of combining simvastatin with a phosphodiesterase type-5 inhibitor, sildenafil, in the rat model of hypoxia-induced pulmonary hypertension. Rats were allocated to either: 1) a prevention protocol, to receive simvastatin 20 mg x kg(-1) x day(-1) by intraperitoneal injection or sildenafil 75 mg x kg(-1) x day(-1) orally or the combination (or vehicle) for 2 weeks beginning at the start of exposure to hypoxia (10% inspired oxygen); or 2) a treatment protocol, where the same agents were administered in the last 2 weeks of a 4-week period of hypoxia. In both protocols, the combination of sildenafil and simvastatin lowered pulmonary artery pressure and produced a significantly greater reduction in right ventricular hypertrophy and pulmonary vascular muscularisation than either drug alone. Moreover, the combination augmented significantly endothelial nitric oxide synthase expression and cGMP levels in the lung and right ventricle above that produced by either drug independently and resulted in greater inhibition of RhoA activity. These data suggest that simvastatin can be usefully combined with sildenafil in the treatment of pulmonary arterial hypertension to achieve greater therapeutic benefit.

Guidelines for the diagnosis and management of osteoporosis in postmenopausal women and men from the age of 50 years in the UK.

In 1999 and 2000 the Royal College of Physicians published guidelines for the prevention and treatment of osteoporosis [Royal College of Physicians. Osteoporosis: clinical guidelines for the prevention and treatment. London: Royal College of Physicians; 1999; Royal College of Physicians and Bone and Tooth Society of Great Britain. Update on pharmacological interventions and an algorithm for management. London, UK: Royal College of Physicians; 2000.; Royal College of Physicians. Glucocorticoid-induced osteoporosis. Guidelines on prevention and treatment; Bone and Tooth Society of Great Britain, National Osteoporosis Society and Royal College of Physicians. London, UK: Royal College of Physicians; 2002]. Since then, there have been significant advances in the field of osteoporosis including the development of new techniques for measuring bone mineral density, improved methods of assessing fracture risk and new treatments that have been shown to significantly reduce the risk of fractures. Against this background, the National Osteoporosis Guideline Group (NOGG), in collaboration with many Societies in the UK, have updated the original guidelines [Royal College of Physicians, National Osteoporosis Guideline Group on behalf of the Bone Research Society, British Geriatrics Society, British Orthopaedic Association, British Society of Rheumatology, National Osteoporosis Society, Osteoporosis 2000, Osteoporosis Dorset, Primary Care Rheumatology Society, Society for Endocrinology. Osteoporosis. Clinical guideline for prevention and treatment, Executive Summary. University of Sheffield Press; 2008], a practical summary of which is detailed below. The management algorithms are underpinned by a health economic analysis applied to the epidemiology of fracture in the UK.

Language issues in the community pharmacy: a perspective from Wales.

OBJECTIVES: This study aimed to obtain the views of Welsh speakers to explore the role of the Welsh language in community pharmacies in bilingual communities in Wales. METHODS: Two communities with a high proportion of Welsh speakers were purposively identified for the research: one in North Wales and one in West Wales. Stage 1: semi-structured interviews with a purposive sample of Welsh speakers to identify key themes. Data collection continued until no new themes emerged. Interviews were tape-recorded, transcribed verbatim, coded manually and analysed thematically. Stage 2: self-complete questionnaire developed based on the interview results. The anonymous, bilingual questionnaire and covering letter, with a postage-paid envelope, were delivered to 500 homes (250 in each community) for completion by the person in the household who visited a pharmacy most often. There was no follow-up mailing due to anonymity. Data were analysed using SPSS version 12. KEY FINDINGS: Results from both interviews (n = 36) and questionnaires (response rate was 52%, 82% of whom were Welsh speakers) found that the majority of Welsh speakers in the study were able to understand English but preferred to use Welsh in the pharmacy. They would find it easier to explain symptoms and would ask more about their medication if they could speak Welsh with the pharmacist. In addition, the study participants would generally feel more at ease with a Welsh-speaking pharmacist and would feel they were getting a better service if they could use their first language. CONCLUSIONS: This study of Welsh speakers indicates that language choice is important for bilingual people who may prefer to use their native, minority language for consultation with health professionals. Further, it is clear that a concordant partnership between patient and pharmacist is less likely where one party is using a language with which they are not confident or comfortable. Pharmacists need to be aware of the linguistic needs and preferences of bilingual clients.

Hypoglycaemic and amylase inhibitory activities of leaves of Spondias mombin Linn.

Suppressing the production of glucose by inhibiting a-amylase / a-glucosidase activity is one of the therapeutic approaches for decreasing postprandial hyperglycaemia and a strategy for evaluating antidiabetic activity. We investigated leaves of Spondias mombin because our previous ethnobotanical survey showed that it is used by traditional healers to manage diabetes in South West Nigeria. We report a bioactivity-guided study of S. mombin using glucose loading (1 g/kg) alloxan-induced diabetic rats and inhibition of a-amylase as basis for isolation of active constituents. Hyperglycaemia was induced in albino rats and blood glucose levels monitored for 180 mins using a glucometer. Powdered leaves were macerated with 80% Methanol. The active extract was fractionated on column chromatography packed with silica gel G6OA eluting with gradient mixtures of pet. ether and ethylacetate. The most active a-amylase inhibiting fraction was purified on thin layer chromatography (TLC) and pure compound identified by spectroscopy. Peak decrease in blood glucose of 41.4% (p < 0.05) was recorded after 60 mins. This activity-guided study produced an active TLC band (69.8% amylase inhibition, p < 0.05) from which a-sitosterol was characterized as the main inhibitor. This is first report of hypoglycaemic and amylase inhibitory activities of S. mombin. The role of phytosterols in control of diabetes mellitus is discussed. This study justifies the ethnopharmacological use of this species in recipes for management of diabetes mellitus.

Non-canonical cyclic AMP SMAD1/5/8 signalling in human granulosa cells.

Development of mammalian ovarian follicles is promoted by the combined action of endocrine cues and paracrine factors. Follicle stimulating hormone (FSH), through the action of cAMP drives follicular growth and development. The oocyte secretes powerful growth factors such as bone morphogenetic protein 15 (BMP15) to regulate granulosa cell proliferation, metabolism, steroidogenesis and differentiation through the activation of SMAD1/5/8. This study investigated the role of the cAMP signalling pathway on SMAD1/5/8 action in human granulosa cells. Cyclic AMP enhanced BMP15-induction of a SMAD1/5/8-specific BRE reporter. Moreover, in the absence of BMP ligand, cAMP also activated SMAD1/5/8-induced BRE activity. Cyclic AMP increased canonical downstream targets of BMP signalling such as inhibitor of differentiation (ID) mRNA expression. The observed effects were not mediated by secretion of BMPs as cAMP did not promote BMP ligand mRNA expression and a BMP extracellular antagonist, the BMP type II receptor ectodomain, did not affect cAMP-induced ID mRNA expression. Finally, the ERK1/2 pathway was shown to be required for the maintenance of cAMP-induced SMAD1/5/8 activity. Together our results suggest a novel and non-canonical pathway for cAMP signalling in human granulosa cells. Cyclic AMP appears to promote SMAD1/5/8 pathway activity intracellularly and has the ability to activate canonical SMAD1/5/8 downstream targets. Our results add another layer of complexity to the interactions between endocrine signalling and oocyte-secreted BMP ligands during folliculogenesis. Given the importance of both cAMP and SMAD1/5/8 pathways in follicular development, these interactions are likely required for the fine-tuning of oocyte paracrine signalling by endocrine stimuli.

Phosphodiesterase inhibitors for the treatment of pulmonary hypertension.

The pulmonary vascular bed is both a source of and target for a number of vasoactive factors. Among the most important for pulmonary vascular homeostasis are factors that utilise cyclic guanosine monophosphate (cGMP) as an intracellular second messenger. These include nitric oxide and the natriuretic peptide family (atrial, brain and C-type natriuretic peptides). In the search for therapeutic strategies that engage the cGMP signalling pathway for the treatment of pulmonary arterial hypertension (PAH), inhibition of cGMP metabolism by phosphodiesterase type 5 (PDE5)-targeted compounds has proven most successful to date. One PDE5 inhibitor, sildenafil, has been shown to improve pulmonary haemodynamics and exercise capacity in patients with PAH and is now an approved treatment. Others are under investigation. An interesting, although still tentative, observation is the potential of sildenafil to reduce pulmonary vascular resistance without adversely affecting ventilation-perfusion matching. Another is the expression of phosphodiesterase type 5 in the hypertrophied right ventricle. These data suggest that phosphodiesterase type 5 inhibitors may have effects that distinguish them from other treatments for pulmonary hypertension and merit further study.

A hand-held surface plasmon resonance biosensor for the detection of ricin and other biological agents.

There is an ongoing need for field-deployable biosensor devices. We have constructed a fully self-contained, hand-held biosensor, based on the surface plasmon resonance technique. The dimensions of the sensor unit are 15 x 8 cm, the weight is 600 g and it is powered by a 9 V battery. We have characterised the responsiveness of the sensor using calibrated sucrose solutions and were able to measure changes as small as 3.3 x 10(-6) refractive index units. To demonstrate functionality of the sensor, we have prepared surfaces with an antibody fragment specific for the biological toxin ricin. We were able to detect ricin at 200 ng/mL in 10 min, which is approximately 2500 times less than the minimum lethal dose. We were also able to verify positive binding within a second 10 min window. This sensor demonstrates important steps required for the development of fully integrated, hand-held sensor devices and will form the basis of a multi-analyte system, to be developed in the near future. It also represents the first completely hand-held SPR device, not requiring external power or a computer connection to operate.

Augmentation of the natriuretic activity of exogenous and endogenous atriopeptin in rats by inhibition of guanosine 3',5'-cyclic monophosphate degradation.

To investigate the relationship between AP, cyclic GMP, and sodium excretion, we studied the effect of a cyclic GMP phosphodiesterase inhibitor (M + B22948) on the natriuretic response to (a) an infusion of AP (103-126) and (b) acute volume expansion in rats. The phosphodiesterase inhibitor markedly potentiated the effect of low-dose AP infusions on urinary sodium and cyclic GMP excretion without potentiating the fall in blood pressure. Acute volume expansion (1% body wt) led to small but significant (P less than 0.01) rises in plasma AP and urinary cyclic GMP levels. Pretreatment with the phosphodiesterase inhibitor enhanced the natriuretic and cyclic GMP response to volume loading, an effect that was attenuated by administration of a monoclonal antibody directed against AP. These data indicate that cyclic GMP mediates the natriuretic activity of AP and AP and cyclic GMP play active roles in the natriuresis of acute volume expansion. Moreover, pharmacological manipulation of cyclic GMP levels may prove a useful therapeutic strategy for facilitating the natriuretic but not the hypotensive effects of AP.

Differential regulation of natriuretic peptide receptor messenger RNAs during the development of cardiac hypertrophy in the rat.

The heart expresses the three natriuretic peptide receptors (NPR), namely NPR-A, NPR-B, and NPR-C. We have examined the temporal relationship between the expression of mRNA transcripts for atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) and their receptors in the heart during the development of cardiac hypertrophy in the aortovenocaval fistula rat. Messenger RNAs were measured by cDNA amplification. Progressive cardiac hypertrophy was accompanied by increased ANP mRNA prevalence throughout the heart and increased BNP mRNA in the left atrium. The most striking observation was the gradual disappearance of NPR-C transcripts (the putative "clearance" receptor) in all chambers; this was in marked contrast to the increase in mRNA levels for NPR-A and NPR-B (the guanylyl cyclase-linked receptors). Our observations have important therapeutic implications if the transcript changes are mirrored at the receptor protein level because (a) the apparent down-regulation of NPR-C may enhance the local action of natriuretic peptides on the heart, and (b) the loss of NPR-C, particularly if it is widespread, may reduce the rate of elimination of the natriuretic peptides, restricting the therapeutic potential of specific NPR-C ligands designed to reduce peptide clearance.