A contrast set mining based approach for cancer subtype analysis.

The task of detecting common and unique characteristics among different cancer subtypes is an important focus of research that aims to improve personalized therapies. Unlike current approaches mainly based on predictive techniques, our study aims to improve the knowledge about the molecular mechanisms that descriptively led to cancer, thus not requiring previous knowledge to be validated. Here, we propose an approach based on contrast set mining to capture high-order relationships in cancer transcriptomic data. In this way, we were able to extract valuable insights from several cancer subtypes in the form of highly specific genetic relationships related to functional pathways affected by the disease. To this end, we have divided several cancer gene expression databases by the subtype associated with each sample to detect which gene groups are related to each cancer subtype. To demonstrate the potential and usefulness of the proposed approach we have extensively analysed RNA-Seq gene expression data from breast, kidney, and colon cancer subtypes. The possible role of the obtained genetic relationships was further evaluated through extensive literature research, while its prognosis was assessed via survival analysis, finding gene expression patterns related to survival in various cancer subtypes. Some gene associations were described in the literature as potential cancer biomarkers while other results have been not described yet and could be a starting point for future research.

Immune response in breast cancer surgery. A comparative and prospective study of different anesthetic techniques.

INTRODUCTION: The main reason for high mortality in breast cancer is local recurrence and metastasis, despite surgery as the first therapeutic option. The anesthesia used in the operation room can determine the immune response. METHODS: A prospective, comparative and non- randomised study in patients undergoing breast cancer surgery was conducted in our hospital after obtaining approval from the Hospital's Institutional Review Board. Patients were divided in two groups: Group A received general anesthesia with propofol and opioids. Group B, in addition to general anesthesia, three interfascial blocks (Pec I, Pec II and BRILMA) were performed in all patients. Three blood samples were taken 1) previous anesthetic induction; 2) two hours after the end of the surgery and 3) 24-48 h after surgery. Leukocytes, CD3, CD4, CD8 and Natural Killer cells were determined at each time. RESULTS: 103 patients were included. 59 (group A) received general anesthesia and 54 (group B) general anesthesia and interfascial blocks. Regarding baseline characteristics, age was significantly higher in the group that received general anesthesia and mastectomy was more frequent in the group that received interfascial blocks. We observed after surgery an increase in leukocytes level that returns close to baseline levels. On the other hand, a reduction in the immune response was observed that also returns to the previous level 48 h after surgery. Group A and B get similar results and also subgroups of hormonal receptors (HER+, PR and/or ER+). CONCLUSIONS: Interfascial blocks in chest wall added to general anesthesia in breast cancer surgery has not shown a significant difference in the inflammatory response or immunological depression compared to general anesthesia as the only anesthetic technique. It seems to trend less immunological depression in the interfascial block group.

Cytotoxicity profiling of deep eutectic solvents to human skin cells.

The tailor-made character of deep eutectic solvents (DES) turns them very attractive to be used in several applications, including in health-related areas such as pharmaceutical, nutraceutical, and cosmetic industries. However, although DES has been touted as "green" solvents, several works proved that their potential toxicity should not be neglected. Using the premise of DES applicability in the cosmetic and pharmaceutical sectors, we chose two cell lines to work as a skin model (keratinocytes HaCaT and tumor melanocytes MNT-1), to assess DES cytotoxicity. The effect of three different hydrogen bond acceptors (HBA) ([Chol]Cl, [N1111]Cl and [N4444]Cl) and three different hydrogen bond donors (HBD) (hexanoic and butanoic acid, ethylene glycol, 1-propanol and urea) were evaluated through a common viability assay (MTT assay). Results were promising since [Chol]Cl and [N1111]Cl- based DES showed good biocompatibility for the tested cells. [N4444]Cl-based DES, however, showed cytotoxicity for both cell lines, with the HBA being the driver of the toxicity. Interestingly, some compounds increased cell viability in the HaCaT cell line, namely [Chol]Cl, ethylene glycol, hexanoic acid, urea, and all [Chol]Cl and [N1111]Cl-based DES and should be considered as targets for future studies. These results highlight their possible use in cosmetic or pharmaceutical formulations.

The efficacy of HRAS and CDK4/6 inhibitors in anaplastic thyroid cancer cell lines.

PURPOSE: Anaplastic thyroid carcinomas (ATCs) are non-responsive to multimodal therapy, representing one of the major challenges in thyroid cancer. Previously, our group has shown that genes involved in cell cycle are deregulated in ATCs, and the most common mutations in these tumours occurred in cell proliferation and cell cycle related genes, namely TP53, RAS, CDKN2A and CDKN2B, making these genes potential targets for ATCs treatment. Here, we investigated the inhibition of HRAS by tipifarnib (TIP) and cyclin D-cyclin-dependent kinase 4/6 (CDK4/6) by palbociclib (PD), in ATC cells. METHODS: ATC cell lines, mutated or wild type for HRAS, CDKN2A and CDKN2B genes, were used and the cytotoxic effects of PD and TIP in each cell line were evaluated. Half maximal inhibitory concentration (IC50) values were determined for these drugs and its effects on cell cycle, cell death and cell proliferation were subsequently analysed. RESULTS: Cell culture studies demonstrated that 0.1 µM TIP induced cell cycle arrest in the G2/M phase (50%, p < 0.01), cell death, and inhibition of cell viability (p < 0.001), only in the HRAS mutated cell line. PD lowest concentration (0.1 µM) increased significantly cell cycle arrest in the G0/G1 phase (80%, p < 0.05), but only in ATC cell lines with alterations in CDKN2A/CDKN2B genes; additionally, 0.5 µM PD induced cell death. The inhibition of cell viability by PD was more pronounced in cells with alterations in CDKN2A/CDKN2B genes (p < 0.05) and/or cyclin D1 overexpression. CONCLUSIONS: This study suggests that TIP and PD, which are currently in clinical trials for other types of cancer, may play a relevant role in ATC treatment, depending on the specific tumour molecular profile.

Leishmania infantum exerts immunomodulation in canine Kupffer cells reverted by meglumine antimoniate.

Kupffer cells (KC) are the liver macrophage population that resides in the hepatic sinusoids and efficiently phagocyte pathogens by establishing an intimate contact with circulating blood. KC constitute the liver host cells in Leishmania infection, nevertheless little is described about their role, apart from their notable contribution in granulomatous inflammation. The present study aims to investigate how canine KC sense and react to the presence of Leishmania infantum promastigotes and amastigotes by evaluating the gene expression of specific innate immune cell receptors and cytokines, as well as the induction of nitric oxide and urea production. Complementarily, the impact of a leishmanicidal drug - meglumine antimoniate (MgA) - in infected KC was also explored. KC revealed to be susceptible to both parasite forms and no major differences were found in the immune response generated. L. infantum parasites seem to interact with KC innate immune receptors and induce an anergic state, promoting immune tolerance and parasite survival. The addition of MgA to infected KC breaks the parasite imposed silence and increased gene expression of Toll-like receptors (TLR) 2 and TLR4, possibly activating downstream pathways. Understanding how KC sense and react to parasite presence could bring new insights into the control or even elimination of canine leishmaniasis.

Cytotoxic and genotoxic potential of geraniol in peripheral blood mononuclear cells and human hepatoma cell line (HepG2).

Geraniol is an acyclic monoterpene alcohol present in the essential oil of many aromatic plants and is one of the most frequently used molecules by the flavor and fragrance industries. The literature also reports its therapeutic potential, highlighting itself especially as a likely molecule for the development of drugs against cancer. In view of these considerations, this study was designed to evaluate the cytotoxic and genotoxic potential of geraniol, in an in vitro protocol, using two types of human cells: one without the ability to metabolize (peripheral blood mononuclear cells - PBMC), and the other with this capability (human hepatoma cell line - HepG2) through the comet assay and the micronucleus test. Four concentrations (10, 25, 50, and 100 µg/mL) were selected for the genotoxic assessment for PBMC and three (1.25, 2.5, and 5 µg/mL) for HepG2 cells based on cytotoxicity tests (MTT assay). Results showed that geraniol did not present genotoxic or clastogenic/aneugenic effects on both cell types under the conditions studied. However, caution is advised in the use of this substance by humans, since a significant reduction in viability of HepG2 and a marked decrease in cell viability on normal PBMC were verified.

CD99 regulates neural differentiation of Ewing sarcoma cells through miR-34a-Notch-mediated control of NF-κB signaling.

Sarcomas are mesenchymal tumors characterized by blocked differentiation process. In Ewing sarcoma (EWS) both CD99 and EWS-FLI1 concur to oncogenesis and inhibition of differentiation. Here, we demonstrate that uncoupling CD99 from EWS-FLI1 by silencing the former, nuclear factor-κB (NF-κB) signaling is inhibited and the neural differentiation program is re-established. NF-κB inhibition passes through miR-34a-mediated repression of Notch pathway. CD99 counteracts EWS-FLI1 in controlling NF-κB signaling through the miR-34a, which is increased and secreted into exosomes released by CD99-silenced EWS cells. Delivery of exosomes from CD99-silenced cells was sufficient to induce neural differentiation in recipient EWS cells through miR-34a inhibition of Notch-NF-κB signaling. Notably, even the partial delivery of CD99 small interfering RNA may have a broad effect on the entire tumor cell population owing to the spread operated by their miR-34a-enriched exosomes, a feature opening to a new therapeutic option.

Influence of Cytoplasmatic Folding on Mitochondrial Import.

The eukaryotic cell, with its organelle organization, represents a challenge for protein traffic. Contrary to what occurs in the endoplasmic reticulum, mitochondrial protein import is proposed to occur postranslationaly, as proteins are synthesized in cytoplasmic ribosomes and only then imported to the organelle. Because the diameter of the Tom and Tim pores is too narrow for the passage of a folded protein, it is assumed that polypeptides must be already in an unfolded, import competent, state for organelle entry. However, it has been suggested that mitochondria might be able to actively unfold proteins itself at the outer membrane. Here we discuss the influence of cytoplasmatic protein folding on mitochondrial import. Despite the contribution of active mitochondrial unfolding to protein import is not excluded, this mechanism is inconsistent with a number of experimental evidences. Accordingly, other alternative models for mitochondrial import are here discussed. Understanding the molecular constraints regulating this process is of crucial importance, since its failure can lead to a number of pathological situations.

Prognostic significance of miR-34a in Ewing sarcoma is associated with cyclin D1 and ki-67 expression.

BACKGROUND: At diagnosis, identification of reliable biological indicators of prognosis to allow stratification of patients according to different risks is an important but still unresolved aspect in the treatment of Ewing sarcoma (EWS) patients. This study aimed to explore the role of miR-34A expression on prognosis of EWS patients. PATIENTS AND METHODS: Specimens from 109 patients with non-metastatic EWS treated at the Rizzoli Institute with neoadjuvant chemotherapy (protocols ISG/SSGIII, EW-1, EW-2, EW-REN2, EW-REN3, EW-PILOT) and 17 metastases were studied. Sixty-eight patients (62%) remained disease-free and 41 (38%) relapsed (median follow-up: 67 months, range 9-241 months). Expression of miR-34a and of some of its targets (cyclin D1, bcl-2, SIRT1 and YY1) was evaluated by qRT-PCR using TaqMan MicroRNA Assays and/or by immunohistochemistry on tissue microarrays from the same patients. RESULTS: High expression of miR-34a in localized tumors was significantly related to better event-free and overall survival (P = 0.004). Relevance of miR-34a was confirmed by using different calibrators (normal mesenchymal stem cells and different normal tissues). By multivariate Cox regression analysis, low miR-34a expression as well as nontotal necrosis and high levels of lactate dehydrogenase were all confirmed as independent risk factors associated with poor outcome. Expression of miR-34a was lower in metastases than in primary tumors. It inversely correlated with expression of cyclin D1 and Ki-67. CONCLUSIONS: By demonstrating its relationship with clinical outcome, we propose evaluation of miR-34a at diagnosis of EWS patients to allow early risk stratification. Validation of these results would nonetheless ultimately need a prospective assessment.

A novel androgen signalling pathway uses dihydrotestosterone, but not testosterone, to activate the EGF receptor signalling cascade in prostate stromal cells.

BACKGROUND AND PURPOSE: Human prostate growth and function are tightly controlled by androgens that are generally thought to exert their effects by regulating gene transcription. However, a rapid, non-genomic steroid action, often involving an elevation of intracellular calcium ([Ca(2+) ]i ), has also been described in a number of cell types. In this study we investigate whether androgens acutely regulate [Ca(2+) ]i in stromal cells derived from the human prostate. EXPERIMENTAL APPROACH: Human-cultured prostatic stromal cells (HCPSCs) were loaded with the calcium-sensitive fluorophore, fura-2-acetoxymethyl ester (FURA-2AM) (10 µM). Changes in [Ca(2+) ]i in response to the androgens, dihydrotestosterone (DHT) and testosterone, as well as EGF were measured by fluorescence microscopy. KEY RESULTS: DHT, but not testosterone (0.03-300 nM), elicited concentration-dependent elevations of [Ca(2+) ]i within 1 min of addition. These responses were blocked by the androgen receptor antagonist, flutamide (10 µM); the sarcoplasmic reticulum ATPase pump inhibitor, thapsigargin (1 µM); the inositol trisphosphate receptor inhibitor, 2-aminoethyldiphenyl borate (50 µM) and the PLC inhibitor, U-73122 (1 µM). Responses were also blocked by the L-type calcium channel blocker, nifedipine (1 µM), and by removal of extracellular calcium. A similar transient elevation of [Ca(2+) ]i was elicited by EGF (100 ng·mL(-1) ). The EGF receptor inhibitor, AG 1478 (30 nM), and the MMP inhibitor, marimastat (100 nM), blocked the DHT-induced elevation of [Ca(2+) ]i . CONCLUSIONS AND IMPLICATIONS: These studies show that DHT elicits an androgen receptor-dependent acute elevation of [Ca(2+) ]i in HCPSC, most likely by activating EGF receptor signalling.

Novel drug targets for the pharmacotherapy of benign prostatic hyperplasia (BPH).

Benign prostatic hyperplasia (BPH) is the major cause of lower urinary tract symptoms in men aged 50 or older. Symptoms are not normally life threatening, but often drastically affect the quality of life. The number of men seeking treatment for BPH is expected to grow in the next few years as a result of the ageing male population. Estimates of annual pharmaceutical sales of BPH therapies range from $US 3 to 10 billion, yet this market is dominated by two drug classes. Current drugs are only effective in treating mild to moderate symptoms, yet despite this, no emerging contenders appear to be on the horizon. This is remarkable given the increasing number of patients with severe symptoms who are required to undergo invasive and unpleasant surgery. This review provides a brief background on prostate function and the pathophysiology of BPH, followed by a brief description of BPH epidemiology, the burden it places on society, and the current surgical and pharmaceutical therapies. The recent literature on emerging contenders to current therapies and novel drug targets is then reviewed, focusing on drug targets which are able to relax prostatic smooth muscle in a similar way to the α(1) -adrenoceptor antagonists, as this appears to be the most effective mechanism of action. Other mechanisms which may be of benefit are also discussed. It is concluded that recent basic research has revealed a number of novel drug targets such as muscarinic receptor or P2X-purinoceptor antagonists, which have the potential to produce more effective and safer drug treatments.

Isoflavones isolated from red clover (Trifolium pratense) inhibit smooth muscle contraction of the isolated rat prostate gland.

This study investigated whether red clover contains any bioactive constituents which may affect contractility of rat prostatic smooth muscle in an attempt to determine whether its medicinal use in the treatment of benign prostatic hyperplasia is supported by pharmacological effects. A commercially available red clover extract was chemically fractionated and various isoflavones (genistein, formononetin and biochanin A) were isolated from these fractions and their effects on contractility were examined on preparations of the isolated rat prostate gland. Contractile effects of the isolated fractions were compared with commercially available isoflavones (genistein, formononetin and biochanin A). Pharmacological tools were used to investigate the mechanism of action modifying smooth muscle contraction. Crude red clover extract (Trinovin) inhibited electrical field stimulation induced contractions of the rat prostate across a range of frequencies with an IC(50) of approximately 68 microg/ml. Contractions of the rat prostate elicited by exogenous administration of acetylcholine, noradrenaline or adenosine 5'-triphosphate (ATP) were also inhibited. Chromatographic separation, and final purification by high performance liquid chromatography (HPLC) permitted the isolation of the isoflavones: daidzein, calycosin, formononetin, prunetin, pratensin, biochanin A and genistein. Genistein, formononetin and biochanin A (100 microM) from either commercial sources or isolated from red clover extract inhibited electrical field stimulation induced contractions of the isolated rat prostate. It is concluded that isoflavones contained in red clover are able to inhibit prostatic smooth muscle contractions in addition to their antiproliferative effects. However, the high concentrations required to observe these smooth muscle relaxant effects mean that a therapeutic benefit from this mechanism is unlikely at doses used clinically.

Regional vascular responses to ATP and ATP analogues in the rabbit kidney in vivo: roles for adenosine receptors and prostanoids.

BACKGROUND AND PURPOSE: Our knowledge of the effects of P2-receptor activation on renal vascular tone comes mostly from in vitro models. We aimed to characterise the pharmacology of ATP in the renal circulation in vivo. EXPERIMENTAL APPROACH: In pentobarbitone anaesthetized rabbits, we examined total renal and medullary vascular responses to ATP (0.2 and 0.8 mg kg(-1)), beta, gamma-methylene ATP (beta, gamma-mATP, 7 and 170 microg kg(-1)), alpha, beta-mATP (0.2 and 2 microg kg(-1)) and adenosine (2 and 6 microg kg(-1)) using transit-time ultrasound and laser Doppler flowmetry, respectively. We also determined whether adenosine receptors, NO or prostanoids contribute to the actions of the purinoceptor agonists. KEY RESULTS: Renal arterial boluses of ATP, beta,gamma-mATP, and adenosine produced biphasic changes; ischaemia followed by hyperaemia, in total renal and medullary blood flow. alpha,beta-mATP induced only ischaemia. The adenosine receptor antagonist 8-(p-sulphophenyl)theophylline reduced the responses to adenosine and the hyperaemic responses to ATP and beta,gamma-mATP only. NO synthase inhibition (Nomega-nitro-L-arginine) did not significantly alter responses to the P2 receptor agonists. Subsequent cyclooxygenase inhibition (ibuprofen) reduced the ATP- and beta, gamma-mATP-induced increases in renal blood flow. All other responses remained unchanged. CONCLUSIONS AND IMPLICATIONS: In the rabbit kidney in vivo, alpha, beta-mATP sensitive receptors mediate vasoconstriction. beta,gamma-mATP and ATP induce vasodilation at least partly through adenosine receptors. ATP induced renal vasodilatation is independent of NO and partly dependent on prostanoids in the bulk of the kidney, but not in the vasculature controlling medullary blood flow.

Lack of contribution of P2X receptors to neurally mediated vasoconstriction in the rabbit kidney in vivo.

AIM: The contribution of adenosine triphosphate (ATP) to the neural control of regional renal perfusion in vivo remains unknown. We therefore examined whether P2X receptors mediate renal vascular responses to electrical stimulation of the renal nerves (RNS) in pentobarbitone anaesthetized rabbits. METHODS: Responses to RNS were tested before and during renal arterial infusion of alpha,beta-methylene ATP (alpha,beta-mATP, 7-56 microg kg(-1) min(-1)) to desensitize P2X1 receptors. RNS consisted of 3 min trains at graded frequencies and short trains of RNS (4-32 pulses). RESULTS: Three-minute trains of RNS reduced renal blood flow (RBF), cortical laser Doppler flux (CLDF), and medullary LDF (MLDF) by -90 +/- 3%, -89 +/- 3% and -31 +/- 11%, respectively, at 4 Hz. MLDF was reduced less than CLDF or RBF. During short train RNS, RBF, CLDF and MLDF were reduced by -22 +/- 2%, -15 +/- 2% and -12 +/- 2%, respectively, for 32 s at 1 Hz. CLDF and MLDF were reduced to a similar extent. Infusion of alpha,beta-mATP induced transient reductions in RBF, CLDF and MLDF, but within 5 min these variables had recovered to control levels. Vascular responses to RNS were not significantly altered by alpha,beta-mATP treatment. CONCLUSIONS: In the rabbit kidney in vivo, alpha,beta-mATP-sensitive receptors mediate vasoconstriction and reduce perfusion in both cortical and medullary vascular beds. However, these receptors do not mediate neurally induced reductions in renal perfusion.

Detailed molecular comparison between the inhibition mode of A/B-type carboxypeptidases in the zymogen state and by the endogenous inhibitor latexin.

Treatment of advanced stages of prostate carcinoma with histone-deacetylase inhibitors entails expression of human procarboxypeptidase-A4 (hPCPA4). The three-dimensional structure of hPCPA4 has been solved and shows the features of related metallocarboxypeptidase zymogens, with a preformed alpha/beta/-hydrolase active-enzyme moiety (hCPA4) and an inhibiting pro-domain (PD). The protease moiety recalls a sphere, out of which a spherical cone has been cut. This results in a funnel-like structure, at the bottom of which the active-site cleft resides. The border of this funnel is shaped by loops, which are responsible for the interaction with the PD, characterised by a large interface area and relatively few contacts. Such an inhibitory mode is evocative of the recently reported structure of the human inhibitor latexin in its complex with hCPA4. The main contacting structure of latexin is similar to the one employed for PD inhibition. In both cases, active-site blocking relies mainly on a loop provided by the central part of a beta sheet.

The in vitro and in vivo pharmacological activity of Boiga dendrophila (mangrove catsnake) venom.

The great taxonomic and prey base diversity of colubrids (non-front-fanged snakes) suggests that their venoms may represent a 'literal gold mine' for scientists eager to find novel pharmacological probes. While pharmacological characterization is lacking for most of these venoms, this is even more so with regard to activity of colubrid venoms on the mammalian autonomic nervous system. This study characterizes the activity of venom from the colubrid, Boiga dendrophila using in vitro smooth muscle preparations and the anaesthetized rat. In the prostatic segment of the rat vas deferens, cumulative additions of venom (1-150 microg ml(-1)) induced concentration-dependent inhibition of electrically evoked (0.2 Hz, 0.3 ms, 70-100 V) twitches. The inhibitory effect of venom (100 microg ml(-1)) was attenuated by 8-phenyltheophylline (8-PT) (20 microM) and 8-cyclopentyl-1, 3-dipropylxanthine (20 microM) but not idazoxan (1 microM), or a combination of ranitidine (0.2 microM) and thioperamide (10 microM). The inhibitory effect of venom (100 microg ml(-1)) was augmented by dipyridamole (10 microM) but abolished by pretreatment with adenosine deaminase (7.5 units/100 microl) suggesting that it contains components with adenosine A(1) receptor activity, most likely adenosine. In isolated segments of guinea-pig ileum, venom (10-100 microg ml(-1)) caused concentration-dependent contractions which were inhibited by the muscarinic receptor antagonist atropine (0.1 microM) but not by the histamine receptor antagonist mepyramine (0.5 microM). In the anaesthetized rat, venom (5-7.5 mg kg(-1), i.v.) caused a hypotensive effect. Our data suggest that the venom contains components with purinergic and muscarinic receptor activity.

Cholinergic innervation and function in the prostate gland.

The mammalian prostate is densely innervated by hypogastric and pelvic nerves that play an important role in regulating the growth and function of the gland. While there has been much interest in the role of the noradrenergic innervation and adrenoceptors in prostate function, the role of cholinergic neurones in prostate physiology and pathophysiology is not well understood. This review focuses on the role of acetylcholine and cholinoceptors in prostate function. Nitric oxide, vasoactive intestinal polypeptide, and/or neuropeptide Y are co-localised with cholinesterase and/or acetylcholine transporter in some of the nerve fibres supplying the prostate. Their roles are also briefly discussed in this review. A dense network of cholinesterase-staining fibres supplies both prostate epithelium and stroma, suggesting a role of acetylcholine and/or co-localised neuropeptides in the modulation of prostatic secretions, as well as smooth muscle tone. A predominantly epithelial location for prostate muscarinic receptors indicated a major secretomotor role for acetylcholine. The muscarinic receptor subtype mediating muscarinic agonist-induced smooth muscle contraction or enhancement of contractions evoked by nerve stimulation differs in different species. In the human, there is evidence for M(1) receptors on the epithelium, M(2) receptors on the stroma, and both M(1) and M(3) receptors in some prostate cancer cell lines. Several recent investigations indicate that muscarinic receptors may also mediate or modulate normal, benign, and malignant prostate growth. The role of muscarinic agonists and their receptors and the influences of age, testicular, and other steroids in regulating the effects are reviewed.

Births: final data for 1999.

OBJECTIVES: This report presents 1999 data on U.S. births according to a wide variety of characteristics. Data are presented for maternal demographic characteristics including age, live-birth order, race, Hispanic origin, marital status, and educational attainment; maternal characteristics (medical risk factors, weight gain, tobacco and alcohol use); medical care utilization by pregnant women (prenatal care, obstetric procedures, complications of labor and/or delivery, attendant at birth, and method of delivery); and infant characteristics (period of gestation, birthweight, Apgar score, abnormal conditions, congenital anomalies, and multiple births). Also presented are birth and fertility rates by age, live-birth order, race, Hispanic origin, and marital status. Selected data by mother's State of residence are shown, as well as data on month and day of birth, sex ratio, and age of father. Trends in fertility patterns and maternal and infant characteristics are described and interpreted. METHODS: Descriptive tabulations of data reported on the birth certificates of the 3.96 million births that occurred in 1999 are presented. RESULTS: Overall birth and fertility rates changed less than 1 percent in 1999. Teenage birth rates fell 2 to 6 percent. The rate for women aged 20-24 years declined slightly, while rates for women in their late twenties and their thirties rose 2 to 3 percent each. The number of births to unmarried women, the birth rate, and the percent of births that were to unmarried women each rose 1 percent or less. Smoking by pregnant women overall dropped again, but rose among women aged 18-24 years. Improvements in prenatal care utilization continued. The cesarean delivery rate increased for the third year after declining for 7 consecutive years. The proportion of multiple births continued to rise; however, higher order multiple births (e.g., triplets, quadruplets) declined for the first time in over a decade, following increases of 13 percent per year during 1990-98. The percent low birthweight remained at 7.6 percent, while preterm births rose to 11.8 percent. These trends are in large part the result of increases in multiple births.

Effects of adenine nucleosides and nucleotides on neuromuscular transmission to the prostatic stroma of the rat.

1. The aim of this study was to investigate the effects of adenine nucleosides and nucleotides on contractility of the smooth muscle of rat prostate gland. 2. Nerve terminals within rat isolated prostatic tissues were electrically field stimulated (60 V, 0.5 ms, 10 Hz, 20 pulses every 60 s). Adenosine 5'-triphosphate (ATP), adenosine 5'-diphosphate (ADP), adenosine 5'-monophosphate (AMP) and adenosine had no effect on baseline smooth muscle tone but concentration-dependently inhibited electrically-evoked contractile responses. The relative order of potency was ATP congruent with AMP congruent with adenosine>ADP. 3. The inhibition by ATP and adenosine of field stimulation-induced contractions in the rat prostate was antagonized by 8-phenyltheophylline (10 microM), but not by suramin (100 microM) and only slightly by reactive blue 2 (5 microM). 4. The adenosine metabolizing enzyme adenosine deaminase (0.1 unit ml(-1)) inhibited the inhibitory effects of ATP and adenosine. The P2 purinoceptor agonist 2-methylthio ATP (10 nM - 0.1 mM), had no effect on field stimulation-induced contractions of the rat prostate. 5. ATP and adenosine did not modify the contractile responses of the rat prostate to exogenously added noradrenaline (10 microM). 6. Inhibitory concentration-response curves to a number of adenosine analogues with differing stabilities and selectivities for the different adenosine receptors yielded a relative rank order of agonist potency of: N(6)-cyclopentyladenosine (CPA)>N(6)-cyclohexyladenosine (CHA) congruent with (-)-N(6)-(2-phenylisopropyl)-adenosine (R-PIA) congruent with 5'-(N-ethylcarboxamido)-adenosine (NECA)>(+)-N(6)-(2-phenylisopropyl)-adenosine (S-PIA)>2-p-[2-carboxyethyl]phenethyl-amino-5'-N-ethylcarboxamido-ade nosine (CGS 21680). 7. These results indicate that adenine nucleoside and nucleotide induced inhibition of electrically-evoked contractions in the rat prostate occurs through activation of adenosine but not ATP receptors. The relative order of potency of adenosine analogues is consistent with activation of receptors of the A(1)-adenosine receptor subtype. These receptors appear to be prejunctional.

Calcitonin gene-related peptide (CGRP) inhibits contractions of the prostatic stroma of the rat but not the guinea-pig.

This study investigated the presence and effects of calcitonin gene-related peptide (CGRP) within the rat and guinea-pig prostate glands. Immunohistochemical studies demonstrated that CGRP immunoreactive nerve fibres are sparsely distributed throughout the prostatic fibromuscular stroma in both species. These CGRP immunopositive nerve fibres shared a similar distribution profile but were not colocalized with tyrosine hydroxylase immunopositive nerve fibres which also innervate the prostatic stroma of these species. Nerve terminals within rat and guinea-pig prostatic tissues were electrically field stimulated (60 V, 0.5 ms, 10 Hz, 20 pulses every 60 s). In guinea-pig preparations, application of human alpha-CGRP, rat adrenomedullin or rat amylin (0.1 nM-1 microM) had no effect on responses to field stimulation. In contrast, both rat and human alpha-CGRP (10 pM-300 nM), rat adrenomedullin (0.3 nM-1 microM) and rat amylin (3 nM-1 microM) concentration-dependently inhibited electrically evoked contractile responses in the rat prostate. The relative order of potency was rat alpha-CGRP=human alpha-CGRP>rat adrenomedullin>rat amylin. The inhibition by rat alpha-CGRP of field stimulation-induced contractions in the rat prostate was competitively antagonized by human CGRP((8-37)) (1, 3 and 10 microM) with a pA(2) of 6.20+/-0.13. Rat alpha-CGRP (10 nM) attenuated contractile responses of the rat prostate to exogenously added noradrenaline (1-100 microM). Inhibitory concentration-response curves to rat alpha-CGRP in rat prostates were unaffected by preincubation in either glibenclamide (10-100 microM), N-nitro-L-arginine methyl ester (L-NAME) (10 microM), bestatin (10 microM), captopril (10 microM) or phosphoramidon (3 microM). Our results indicate that CGRP-induced inhibition of electrically evoked contractions in the rat prostate occurs through activation of postjunctional CGRP(2) receptors which act independently of a K(ATP) channel or nitrergic mechanisms. Degradation of rat alpha-CGRP via peptidases does not appear to occur in the rat prostate.