Ovarian stimulation for fertility treatments and risk of breast cancer: a matched cohort study.

STUDY QUESTION: Is there a difference in the breast cancer risk among women who underwent ART treatments compared to those who underwent medically assisted reproduction (MAR) infertility treatments or women of reproductive age in the general population? SUMMARY ANSWER: The risk of breast cancer among women treated by ART was similar to the risk among women treated by MAR and women who did not undergo fertility treatments. WHAT IS KNOWN ALREADY: Studies investigating breast cancer risk in women who have undergone fertility treatments have provided conflicting results. STUDY DESIGN, SIZE, DURATION: A retrospective, population-based cohort study included women who underwent ART or MAR treatments and women who did not undergo fertility treatments from 1994 to 2019. PARTICIPANTS/MATERIALS, SETTING, METHODS: Women who underwent ART were matched one to one with women who underwent MAR treatments and one to one with woman from the general population of reproductive age, by year of birth and year of first delivery or nulliparity status. MAR women were also matched to ART women by treatment initiation calendar year. All included women were members of Maccabi Healthcare Services. Data regarding demographics, fertility treatments, BRCA mutation and possible confounders were obtained from the computerized database of electronic health records. The incidence of breast cancer after fertility treatments was compared to the matched controls. MAIN RESULTS AND THE ROLE OF CHANCE: Of 8 25 721 women of reproductive age, 32 366 women who underwent ART were matched with patients treated by MAR (n = 32 366) and 32 366 women of reproductive age. A total of 984 women (1.0%) were diagnosed with breast cancer (mean follow-up period, 9.1 ± 6.3 years; interquartile range [IQR], 3.8-13.7 years). The incidence rates of breast cancer per 10 000 person-years were 11.9 (95% CI, 10.7-13.3), 10.7 (95% CI, 9.6-11.9) and 10.7 (95% CI, 9.6-12.0) in the ART group, MAR group and general population, respectively. The crude risk for breast cancer was similar in the ART group compared with the general population (hazard ratio (HR) = 1.10, 95% CI, 0.94-1.28) and in the ART group compared with the MAR group (HR = 1.00, 95% CI, 0.86-1.16). Further adjustment for age, BMI, smoking, socioeconomic status and parity did not substantially impact the hazard rates for breast cancer (ART vs general population: HR = 1.10, 95% CI, 0.94-1.28; ART vs MAR: HR = 0.99, 95% CI, 0.85-1.16). Among women diagnosed with breast cancer, the prevalence of BRCA1/2 mutations and tumour staging did not differ between the ART, MAR and general population groups. Among women who underwent ART, no correlation was found between breast cancer and the number of ART cycles or the use of recombinant medications or urine-derived medications. LIMITATIONS, REASONS FOR CAUTION: The mean age of women at the end of follow-up was only 42 years thus the study was not powered to detect potential differences in the risk of postmenopausal breast cancer. In addition, we did not sub-classify the exposed patients by the reason for infertility. WIDER IMPLICATIONS OF THE FINDINGS: Breast cancer incidence following ART was comparable to that in the general population or following MAR. Women undergoing fertility treatments and their clinicians may be reassured about the safety of assisted reproduction technologies in terms of premenopausal breast cancer risk. STUDY FUNDING/COMPETING INTEREST(S): No specific funding was used and there are no competing interests. TRIAL REGISTRATION NUMBER: N/A.

An open-label, long-term evaluation of the safety, efficacy and tolerability of avanafil in male patients with mild to severe erectile dysfunction.

AIM: Determine the long-term efficacy, safety and tolerability of avanafil, a highly specific, rapidly absorbed phosphodiesterase type 5 inhibitor in male patients with mild to severe erectile dysfunction (ED), with or without diabetes. METHODS: This was a 52-week, open-label extension of two 12-week, randomised, placebo-controlled, phase 3 trials. Patients were assigned to avanafil 100 mg, but could request 200 mg (for increased efficacy; '100/200-mg' group) or 50 mg (for improved tolerability). Primary end points included percentage of sexual attempts ending in successful vaginal penetration [Sexual Encounter Profile 2 (SEP2)] and intercourse (SEP3) and erectile function domain score per the International Index of Erectile Function (IIEF-EF). RESULTS: Some 712 patients enrolled; 686 were included in the intent to treat population and contributed to the data. All primary end points showed sustained improvement. SEP2 and SEP3 success rates improved from 44% to 83% and from 13% to 68% (100-mg group) and from 43% to 79% and from 11% to 66% (100/200-mg group), respectively. Mean IIEF-EF domain scores improved from 13.6 to 22.2 (100-mg group) and from 11.9 to 22.7 (100/200-mg group). Avanafil was effective in some patients ≤ 15 min and > 6 h postdose. Sixty-five per cent (112/172) of 'nonresponders' to avanafil 100 mg responded to the 200-mg dose. The most common (≥ 2%) treatment-emergent adverse events were headache, flushing, nasopharyngitis and nasal congestion; < 3% of patients discontinued therapy because of adverse events. CONCLUSIONS: The long-term tolerability and improvement in sexual function, coupled with rapid onset, suggest that avanafil is well suited for the on-demand treatment of ED.

The assessment of vascular risk in men with erectile dysfunction: the role of the cardiologist and general physician.

Erectile dysfunction (ED) and cardiovascular disease (CVD) share risk factors and frequently coexist, with endothelial dysfunction believed to be the pathophysiologic link. ED is common, affecting more than 70% of men with known CVD. In addition, clinical studies have demonstrated that ED in men with no known CVD often precedes a CVD event by 2-5 years. ED severity has been correlated with increasing plaque burden in patients with coronary artery disease. ED is an independent marker of increased CVD risk including all-cause and especially CVD mortality, particularly in men aged 30-60 years. Thus, ED identifies a window of opportunity for CVD risk mitigation. We recommend that a thorough history, physical exam (including visceral adiposity), assessment of ED severity and duration and evaluation including fasting plasma glucose, lipids, resting electrocardiogram, family history, lifestyle factors, serum creatinine (estimated glomerular filtration rate) and albumin:creatinine ratio, and determination of the presence or absence of the metabolic syndrome be performed to characterise cardiovascular risk in all men with ED. Assessment of testosterone levels should also be considered and biomarkers may help to further quantify risk, even though their roles in development of CVD have not been firmly established. Finally, we recommend that a question about ED be included in assessment of CVD risk in all men and be added to CVD risk assessment guidelines.

Mechanisms of lysosomal enzyme release from leukocytes exposed to immune complexes and other particles.

Human PMN release lysosomal enzymes (beta-glucuronidase, acid phosphatase) when exposed to immune complexes, but do not release cytoplasmic LDH. The cells remain viable, and failure of LDH to appear in supernatants is not due to selective absorption or inactivation. Release of enzymes is not due to platelet contamination and is only partially enhanced by fresh serum. The selective release of lysosomal enzymes after uptake of complexes resembles that induced by inert particles of zymosan, and can be distinguished from the concurrent release of all enzymes after cell death induced by membrane-lytic crystals of MSU. Uptake of complexes, zymosan, or MSU particles is accompanied by concomitant increases in C-1 oxidation of glucose. Although MSU-induced damage can be retarded by the presence of Tris buffer, immune complexes and zymosan selectively release lysosomal hydrolases in the presence or absence of Tris buffer. Agents which elevate the level, within cells, of cAMP (PGE(1), theophylline, 2-CA) and cAMP itself inhibit the selective extrusion of acid hydrolases from leukocytes without affecting the viability of cells. Leukocytes may respond to immune particles by regurgitating a portion of their lysosomal hydrolases during phagocytosis.

Influence of local anesthetics upon human polymorphonuclear leukocyte function in vitro. Reduction of lysosomal enzyme release and superoxide anion production.

Cationic local anesthetics have been reported to influence cellular responses to surface stimuli by interfering with the function of microtubules and microfilaments. Since unimpaired microtubule and microfilament functions are required by human polymorphonuclear leukocytes in order to respond normally to surface stimulation, we have studied effects of the local anesthetic, tetracaine on the function and morphology of these cells in vitro. Tetracaine (0.25--1.0 mM) significantly reduced extracellular release of the lysosomal enzymes, beta-glucuronidase and lysozyme from polymorphonuclear leukocytes exposed to serum-treated zymosan (a particulate stimulus), zymosan-treated serum (a soluble stimulus), and to the surface-active lectin, concanavalin A. Tetracaine also significantly reduced superoixde anion production (superoxide dismutase-inhibitable cytochrome c reduction) by these cells. Tetrancaine was not cytotoxic and its effects could be reversed completely by washing cells once with buffer. Electron microscope examination of tetracaine-treated cells revealed marked alterations of surface membranes. Microtubules and microfilaments appeared normal in "resting" polymorphonuclear leukocytes, but the increase in microtubules normally observed in stimulated cells was not seen after tetracaine treatment. These results suggest that tetracaine interferes with those interactions between immune reactants and the polymorphonuclear leukocyte cell surface which provoke exocytosis and increased oxidative metabolism.

Thromboxane generation by human peripheral blood polymorphonuclear leukocytes.

Human peripheral blood polymorphonuclear leukocytes were stimulated to generate thromboxane B2 in a time- and concentration-dependent fashion upon exposure to serum-treated zymosan particles. Conversion by stimulated PMN of [14C] arachidonic acid to [14C]thromboxane B2 was confirmed by thin-layer radiochromatography, radio-gas chromatography, and mass spectrometry. Generation of thromboxane B2 was independent of platelet contamination and could be inhibited by the cyclooxygenase inhibitor, indomethacin. Cells rendered incapable of ingesting particles by treatment with cytochalasin B generated comparable amounts of thromboxane B2. These results suggest that human peripheral blood polymorphonuclear leukocytes synthesize thromboxanes in response to surface stimulation independently of phagocytosis.

Dimensions of the fetal facial profile in normal pregnancy.

OBJECTIVE: To determine sonographic dimensions of the fetal facial profile in normal pregnancy. METHODS: This was a prospective, cross-sectional study of 397 normal healthy fetuses at 14-33 weeks of gestation. After exclusion of the small numbers of patients at the upper GAs, 379 patients between 14.0 and 26.9 weeks of gestation were included in the analyses. The sagittal plane of the fetal facial profile was evaluated using transvaginal and transabdominal ultrasound. Distances from the tip of the nose to the mouth (the line between the lips), from the mouth to the gnathion (lower chin), from the upper philtrum to the mouth, and from the mouth to the upper concavity of the chin were measured and are presented according to gestational age (GA). RESULTS: There was a significant linear correlation between GA and the distance from the tip of the nose to the mouth (r = 0.943; P < 0.00001; y = -37.98 + 7.54 x GA), from the mouth to the gnathion (r = 0.946; P < 0.00001; y = -46.34 + 7.95 x GA), from the upper level of the philtrum to the mouth (r = 0.71; P < 0.00001; y = 0.22 + 3.33 x GA) and from the mouth to the upper concavity of the chin (r = 0.665; P < 0.00001; y = 1.65 + 2.95 x GA). The ratio between the distance from the tip of the nose to the mouth and that from the mouth to the gnathion was also almost constant throughout gestation, as was the ratio between the distance from the upper philtrum to the mouth and that from the mouth to the upper concavity of the chin. CONCLUSIONS: We provide normative data of the fetal facial profile across GA. Our data offer a potential tool for the prenatal diagnosis of abnormal fetal facial profile.

Early preservation of effector functions followed by eventual T cell memory depletion: a model for the delayed onset of the effect of thiopurines.

OBJECTIVE: The onset of the effect of thiopurines is delayed for several months. The aim of this study was to investigate immune mechanisms for this delay. METHODS: The effects of thiopurines on human peripheral blood T cells and on lamina propria lymphocytes were investigated for apoptosis induction by Annexin V/propidium iodide (PI) and for cytokine secretion by intracellular staining and ELISA assays. To investigate the mechanism of the effect of thiopurines in vivo, Balb/C mice were co-immunised with HEL/OVA (hen egg lysozyme/ovalbumin) antigens, and then repeatedly challenged by HEL only, while being treated by mercaptopurine or vehicle alone for either 4 or 20 weeks. The memory response of CD4+ splenocytes towards HEL/OVA was then determined by CFSE (carboxyfluorescein succinimidyl ester) dilution. RESULTS: Thiopurines arrested the proliferation of stimulated T cells but did not enhance the apoptosis of either resting T cells or activated T cells until day 5 poststimulation. Despite the proliferation arrest, stimulated T cells successfully differentiated into effector cells, as evidenced by their capacity for proinflammatory cytokine secretion, potent adhesion and cytotoxicity. Prolonged mercaptopurine treatment of mice for 20 weeks selectively reduced the CD4+ memory response to a repeatedly encountered HEL antigen, but did not affect the T cell memory pool to the previously presented OVA antigen. A shorter, 4 weeks, treatment with mercaptopurine did not inhibit the memory response to either antigen. CONCLUSIONS: T cells arrested from cycling by thiopurines can still differentiate into potent effector cells capable of propagating the inflammatory process. Thiopurine treatment results in depletion of antigen-specific memory T cells, but this effect is dependent upon repeated encounters with the antigen over a prolonged time course.

Mechanisms of lysosomal enzyme release from human polymorphonuclear leukocytes. Effects of phorbol myristate acetate.

PMA enhanced release of the azurophil granule enzyme, beta-glucuronidase, as well as lysozyme, from cytochalasin B-treated PMN's exposed to either zymosan particles or C5a. PMA was active at nanomolar concentrations, was not toxic to the cells, and was most effective when present for brief durations (0-1 min) before exposure of the cells to the stimuli. Beta-glucuronidase was not released in significant amounts from PMN's exposed to PMA alone, in the absence of stimuli such as zymosan or C5a. In contrast, only the specific granule enzyme, lysozyme, was released from unstimulated cells. Electron micrographs of cells exposed to PMA revealed an increase in the number of visible cytoplasmic microtubules as compared to control cells. Enhancement of lysosomal enzyme (beta-glucuronidase) release by PMA appears to be independent of effects on release of specific granule enzymes (lysozyme), but rather is likely due to PMA-induced elevations of cellular cGMP.

Role of microtubule assembly in lysosomal enzyme secretion from human polymorphonuclear leukocytes. A reevaluation.

The dose-related inhibition by colchicine of both lysosomal enzyme release and microtubule assembly was studied in human polymorphonuclear leukocytes (PMN) exposed to the nonphagocytic stimulus, zymosan-treated serum (ZTS). Cells were pretreated with colchicine (60 min, 37 degrees C) with or without cytochalasin B (5 microng/ml, 10 min) and then stimulated with ZTS (10%). Microtubule numbers in both cytochalasin B-treated and untreated PMN were increased by stimulation and depressed below resting levels in a dose-response fashion by colchicine concentrations above 10(-7) M. These concentrations also inhibited enzyme release in a dose-response fashion although the inhibition of microtubule assembly was proportionately greater than the inhibition of enzyme release. Other aspects of PMN morphology were affected by colchicine. Cytochalasin B-treated PMN were rounded, and in thin sections the retracted plasma membrane appeared as invaginations oriented toward centrally located centrioles. Membrane invaginations were restricted to the cell periphery in cells treated with inhibitory concentrations of colchicine, and the centrioles and Golgi apparatus were displaced from their usual position. After stimulation and subsequent degranulation, the size and number of membrane invaginations greatly increased. They remained peripheral in cells pretreated with greater than 10(-7) M colchicine but were numerous in the pericentriolar region in cells treated with less than 10(-7) M. Similarly, untreated PMN that were permitted to phagocytose immune precipitates had many phagosomes adjacent to the centriole. After colchicine treatment, phagosomes were distributed randomly, without any preferential association with the centrioles. These data suggest that microtubules are involved in maintaining the internal organization of cells and the topologic relationships between organelles and the plasma membrane.

Concanavalin A induces microtubule assembly and specific granule discharge in human polymorphonuclear leukocytes.

Human neutrophils stimulated by concanavalin A (Con A, 100 microng/ml) contained markedly enhanced numbers of microtubules and discharged peroxidase-negative (specific) but not peroxidase-position (azurophile) granules. Release of lysozyme from specific granules was dose and time dependent, could be inhibitied by alpha-methyl-D-mannoside, and enhanced by cytochalasin B. Many microtubules were associated with internalized plasma membrane bearing Con A binding sites.

Potassium and amino acid transport in human leukocytes exposed to phagocytic stimuli.

Influxes of potassium and amino acids were measured in suspensions of human polymorphonuclear leukocytes (PMNs) under resting conditions and after various phagocytic stimuli. Both ouabain-sensitive (or pump) and ouabain-insensitive (or leak) influxes of K were determined. In 5 mM external K, mean total K influx was 0.69 nmol/10(6) cells x min, of which 52% was ouabain-sensitive. Ouabain binding was irreversible, and, as in erythrocytes, was inhibited by K. At external concentrations of 0.1 mM, influxes of lysine and leucine were entirely carrier-mediated, with means of 0.021 nmol/10(6) cells x min, and 0.019 nmol/10(6) cells x min, respectively. After incubation of PMNs with zymosan or latex particles, the K pump was reduced more than 60%, whereas amino acid influxes were inhibited only by 30%. PMNs were also exposed to cytochalasin B before challenge by particles: the drug prevented phagocytosis but not surface binding of zymosan, nor did it influence transport of K or amino acids. After pretreatment of PMNs with cytochalasin B, interaction of zymosan with their surface resulted in the same degree of inhibition of influxes of K and amino acids as when the cells were permitted to phagocytose the particles. In contrast, exposure of PMN to latex particles, which do not bind to cytochalasin B-treated cells, after pretreatment of cells with cytochalasin B did not result in inhibition of influxes. Treatment of cells with colchicine had no effect on either membrane transport or its inhibition after exposure to various phagocytic stimuli. These results indicate that the surface membranes of PMNs are functionally heterogeneous with respect to the association of transport sites for the different solutes. Moreover, loss of specific membrane functions from phagocytosing cells may result from the surface-at-tachment phase of particle-cell interactions, since the interactions of zymosan particles with PMNs in the absence of phagocytosis also inhibited transport of solutes.

Potential for Converting Wood into Plastics: Chemicals from wood may regain importance as the cost of petroleum continues to rise.

The conversion of wood into chemicals for the production of most of our synthetic plastics, fibers, and rubbers is technically feasible. With refinements in technology a large integrated plant utilizing all components of the wood for production of ethanol (to be further processed to ethylene and butadiene), phenols and furfural would be approaching economic feasibility as well at current petrochemical prices. If crude oil prices continue to climb at a faster rate than wood costs, the economic feasibility of chemicals for polymers from wood would become certain. Although technical feasibility has not been established, synthetic oils from liquefaction of wood might serve as feedstocks for cracking to chemicals in the same way that crude oil is presently used. The fulfillment of all our polymer needs from wood as a raw material should not place an impossible burden on our wood supply, but might actually improve the availability of wood for lumber, plywood and pulp by providing a use for less valuable wood which would allow reforestation and improved forest management.

P53 in blind subterranean mole rats--loss-of-function versus gain-of-function activities on newly cloned Spalax target genes.

A tumor suppressor gene, p53, controls cellular responses to a variety of stress conditions, including DNA damage and hypoxia, leading to growth arrest and/or apoptosis. Recently, we demonstrated that in blind subterranean mole rats, Spalax, a model organism for hypoxia tolerance, the p53 DNA-binding domain contains a specific Arg174Lys amino acid substitution. This substitution reduces the p53 effect on the transcription of apoptosis genes (apaf1, puma, pten and noxa) and enhances it on human cell cycle arrest and p53 stabilization/homeostasis genes (mdm2, pten, p21 and cycG). In the current study, we cloned Spalax apaf1 promoter and mdm2 intronic regions containing consensus p53-responsive elements. We compared the Spalax-responsive elements to those of human, mouse and rat and investigated the transcriptional activity of Spalax and human Arg174Lys-mutated p53 on target genes of both species. Spalax and human-mutated p53 lost induction of apaf1 transcription, and increased induction of mdm2 transcription. We conclude that Spalax evolved hypoxia-adaptive mechanisms, analogous to the alterations acquired by cancer cells during tumor development, with a bias against apoptosis while favoring cell arrest and DNA repair.

Birth order, atopy, and symptoms of allergy and asthma among inner-city children attending Head Start in New York City.

BACKGROUND: In past research, children with older siblings were more likely than others to wheeze at age 2 years, but less likely by age 6 years. Higher infection transmission and a down-regulated allergic immune response as a result of these infections, respectively, were suggested as the causes. However, in a study of children aged 0-3 years in a low-income urban community in New York City, USA, with high asthma prevalence, we observed no birth-order effect. OBJECTIVE: To evaluate the association between birth order and atopy and respiratory symptoms in 4-year-old children attending Head Start programs in NYC. METHODS: Respiratory symptoms were assessed by questionnaire for 1005 children (mean age 4.0 years) living in high asthma prevalence neighbourhoods. Serum was collected from a subgroup of the children (n=494) and specific IgE responses to dust mite, cockroach, mouse, and cat allergens were measured. RESULTS: Prevalence of specific IgE (> or =0.35 IU/mL) did not differ significantly among first (35%), second (35%), and later-born children (28%) (P=0.23). Increasing birth order was associated with increasing prevalence of respiratory symptoms in the prior year, including wheeze (first 20%, second 27%, third or later 35%; P<0.001), being awakened at night by cough (28%, 33%, 38%; P=0.005), emergency department visits (14%, 17%, 21%; P=0.02) and hospitalizations for difficulty breathing (6.1%, 6.6%, 10%; P=0.04). The associations of birth order with respiratory symptoms were statistically significant only for the non-seroatopic children and those without an asthmatic parent. CONCLUSIONS: Non-seroatopic children with older siblings were more likely than those without older siblings to have respiratory symptoms at age 4 years. Although the stability of these associations over time remains to be determined, the differences in findings between this study and our previous NYC birth cohort study suggest that patterns of asthma development may vary even among low-income populations within the same city.

Asthma, body mass, gender, and Hispanic national origin among 517 preschool children in New York City.

BACKGROUND: Striking differences in asthma prevalence have been reported among Hispanic adults and children living in different cities of the USA. Prevalence is highest among those of Puerto Rican and lowest among those of Mexican origin. We hypothesized that body size would mediate this association. METHODS: Parents of children in New York City Head Start programs completed a questionnaire including demographic factors, health history, a detailed history of respiratory conditions, lifestyle, and home environment. Children's height and weight were measured in home visits. Logistic regression was used to model the association of asthma with body mass index percentile (<85th percentile, gender/age specific vs>or=85th percentile, gender/age specific), national origin, and other factors. RESULTS: Of 517 children at mean age of 4.0 +/- 0.6 years, 34% met the study criteria for asthma, and 43% were above the 85th percentile. Asthma was strongly associated with non-Mexican national origin, male gender, allergy symptoms, and maternal asthma, and marginally with body size. The odds of asthma among boys of non-Mexican origin was 5.9 times that among boys of Mexican origin [95% confidence interval (CI): 2.9-12.2]; the comparable odds ratio (OR) among girls was 1.8 (95% CI: 0.9-3.6). Body mass was associated with asthma among girls [OR = 2.0 (95% CI: 1.1-3.7)], but not boys [OR = 1.4 (95% CI: 0.8-2.6)]. CONCLUSIONS: The association of asthma with both body mass and national origin was gender-specific among the children in our study. Ours is one of the first studies to report on pediatric asthma in different Hispanic populations in the same city, by gender.

Leukotriene B4 omega-hydroxylase in human polymorphonuclear leukocytes. Partial purification and identification as a cytochrome P-450.

Human polymorphonuclear leukocytes (PMN) not only synthesize and respond to leukotriene B4 (LTB4), but also catabolize this mediator of inflammation rapidly and specifically by omega-oxidation. To characterize the enzyme(s) responsible for omega-oxidation of LTB4, human PMN were disrupted by sonication and subjected to differential centrifugation to yield membrane, granule, and cytosol fractions (identified by biochemical markers). LTB4 omega-hydroxylase activity was concentrated (together with NADPH cytochrome c reductase activity) only in the membrane fraction (specific activity increased 10-fold as compared to whole sonicates, 41% recovery). Negligible activity was detected in granule or cytosol fractions. LTB4 omega-hydroxylase activity in isolated PMN membranes was linear with respect to duration of incubation and protein concentration, was maximal at pH 7.4, had a Km for LTB4 of 0.6 microM, and was dependent on oxygen and on reduced pyridine nucleotides (apparent Km for NADPH = 0.5 microM; apparent Km for NADH = 223 microM). The LTB4 omega-hydroxylase was inhibited significantly by carbon monoxide, ferricytochrome c, SKF-525A, and Triton X-100, but was not affected by alpha-naphthoflavone, azide, cyanide, catalase, and superoxide dismutase. Finally, isolated PMN membranes exhibited a carbon monoxide difference spectrum with a peak at 452 nm. Thus, we have partially purified the LTB4 omega-hydroxylase in human PMN and identified the enzyme as a membrane-associated, NADPH-dependent cytochrome P-450.

A specific inhibitor of complement (C5)-derived chemotactic activity in serum from patients with systemic lupus erythematosus.

In the course of examining polymorphonuclear leukocyte (PMN) chemotaxis in patients with systemic lupus erythematosus (SLE), we have found a previously undescribed serum inhibitor of complement (C5)-derived chemotactic activity. Serum from a 25-yr-old Black female with untreated SLE, when activated with zymosan, failed completely to attract either her own or normal PMN. Incubation of normal PMN with the patient's serum did not affect their subsequent random motility or chemotactic response toward normal zymosan-treated serum (ZTS). The patient's serum, however, did inhibit the chemotactic activity of normal ZTS and of column-purified C5-derived peptide(s), but had no effect on the chemotactic activity of either the synthetic peptide, N-formylmethionyl leucyl-phenylalanine or a filtrate prepared from a culture of Escherichia coli (bacterial chemotactic factor). The inhibitory activity in the patient's serum resisted heating at 56 degrees C for 30 min and could be separated from C5-derived chemotactic activity in the patient's ZTS (or normal ZTS that had been incubated with the patient's serum) by chromatography on Sephadex G-75. Despite its effect on C5-derived chemotactic activity, the patient's serum did not influence two other C5-derived biologic activities: PMN lysosomal enzyme-releasing activity and PMN-aggregating activity. Chromatography of the patient's serum (65% ammonium sulfate pellet) on Sephadex G-200 yielded three distinct peaks of inhibitory activity. Two were heat labile and exhibited other properties of the previously described chemotactic factor inactivators of normal human serum. The third and most active peak, however, resisted heating at 56 degrees C for 30 min, eluted with an apparent mol wt of 50,000-60,000, and acted specifically on C5-derived chemotactic activity. This uniquely specific, heat-stable inhibitor of C5-derived chemotactic activity has been found thus far in serum from 4 of 11 patients with active SLE and may account, in part, for altered host defenses against infections caused by pyogenic microorganisms.

Attachment of human C5a des Arg to its cochemotaxin is required for maximum expression of chemotactic activity.

The chemotactic activity of human C5a des Arg is enhanced significantly by an anionic polypeptide (cochemotaxin) in normal human serum and plasma. We have found that the cochemotaxin attaches to the oligosaccharide chain of native C5a des Arg to form a complex with potent chemotactic activity for human polymorphonuclear leukocytes. Although capable of enhancing the chemotactic activity of native C5a des Arg, the cochemotaxin had no effect on the chemotactic activity of either deglycosylated C5a des Arg, native C5a, or N-formyl-methionyl-leucyl-phenylalanine. Of the known components of the oligosaccharide chain, only sialic acid prevented enhancement by the cochemotaxin of the chemotactic activity exhibited by native C5a des Arg. Sialic acid also prevented the formation of C5a des Arg-cochemotaxin complexes, detected by acid polyacrylamide gel electrophoresis, molecular sieve chromatography on polyacrylamide gels, and sucrose density gradient ultracentrifugation.

A nitric oxide-like factor mediates nonadrenergic-noncholinergic neurogenic relaxation of penile corpus cavernosum smooth muscle.

This study was initiated to characterize nonadrenergic-noncholinergic (NANC) inhibitory neurotransmission in penile corpus cavernosum. Using organ baths, isometric tension measurements were made in strips of human and rabbit corpus cavernosum. In examining endothelium-mediated responses, cumulative additions of exogenous acetylcholine elicited dose-dependent relaxations which were significantly reduced or completely inhibited in tissues treated with NG-monomethyl L-arginine (L-NMMA; an inhibitor of nitric oxide synthesis), oxyhemoglobin (a nitric oxide scavenger), or methylene blue (a guanylate cyclase blocker). Tissues exposed to hypoxic conditions (PO2 = 5-10 mmHg) also did not respond to exogenous acetylcholine. Mechanical removal of the endothelium in human corporal strips or in situ treatment of rabbit corpora with detergent blocked the relaxation to acetylcholine. Transmural electrical stimulation of corporal tissue strips denuded of functional endothelium, in the presence of adrenergic blockade with bretylium and muscarinic receptor blockade with atropine, caused frequency-dependent relaxation. This neurogenic relaxation was reduced or prevented by L-NMMA, oxyhemoglobin, methylene blue, and hypoxia. The effects of L-NMMA were reversed by L-arginine and the effects of hypoxia were readily reversed by normoxic conditions. Authentic, exogenous nitric oxide relaxed corporal strips which were contracted with adrenergic agonists and this effect was significantly inhibited by oxyhemoglobin. It is concluded that (a) endothelium-mediated responses of corpus cavernosum smooth muscle are mediated by a diffusible nitric oxide-like substance; (b) NANC neurogenic inhibitory responses do not require functional endothelium, and (c) nitric oxide, or a closely related substance, may act as an inhibitory neurotransmitter in penile corpus cavernosum smooth muscle.