Preoperative breast MRI and mortality in older women with breast cancer.

PURPOSE: The survival benefit from detecting additional breast cancers by preoperative magnetic resonance imaging (MRI) continues to be controversial. METHODS: We followed a cohort of 4454 women diagnosed with non-metastatic breast cancer (stage I-III) from 2/2005-6/2010 in five registries of the breast cancer surveillance consortium (BCSC). BCSC clinical and registry data were linked to Medicare claims and enrollment data. We estimated the cumulative probability of breast cancer-specific and all-cause mortality. We tested the association of preoperative MRI with all-cause mortality using a Cox proportional hazards model. RESULTS: 917 (20.6%) women underwent preoperative MRI. No significant difference in the cumulative probability of breast cancer-specific mortality was found. We observed no significant difference in the hazard of all-cause mortality during the follow-up period after adjusting for sociodemographic and clinical factors among women with MRI (HR 0.90; 95% CI 0.72-1.12) compared to those without MRI. CONCLUSION: Our findings of no breast cancer-specific or all-cause mortality benefit supplement prior results that indicate a lack of improvement in surgical outcomes associated with use of preoperative MRI. In combination with other reports, the results of this analysis highlight the importance of exploring the benefit of preoperative MRI in patient-reported outcomes such as women's decision quality and confidence levels with decisions involving treatment choices.

Proton conduction through full-length gp91phox requires histidine 115.

The NADPH oxidase of neutrophils is a transmembrane electron transfer complex, containing a flavin adenine dinucleotide and two hemes, all of which are suggested to be contained within gp91 (phox), one of four subunits of the enzyme. The transfer of electrons through the NADPH oxidase is associated with an efflux of protons. gp91 (phox) has previously been demonstrated to function as the proton conduction pathway. The mutation of histidines 111, 115, and 119 to leucines and of histidine 115 to leucine within the N-terminal 230-amino-acid fragment of gp91 (phox) has previously been demonstrated to result in the loss of proton conduction through this N-terminal fragment. In this paper we have investigated the role of these histidines in proton conduction by the full-length gp91 (phox). Stable CHO cell lines were established which expressed full-length gp91 (phox) in which histidines 111, 115, and 119 had been mutated to leucines (CHO91H111/115/119) and in which histidine 115 had been mutated to leucine (CHO91H115L). The expression of gp91 (phox) and its cellular localisation in these cell lines were comparable between wild-type and the mutant gp91 (phox). The mutation of histidines 111, 115, and 119 to leucines or just histidine 115 to leucine resulted in an almost total loss of both the arachidonate-activated influx and efflux of protons, in comparison with that observed for wild-type gp91 (phox). Therefore, histidine 115 is required for proton conduction by both full-length gp91 (phox) and the N-terminal 230-amino-acid fragment of gp91 (phox). Histidine 115 has recently been proposed to act as a coordinating ligand for the outer heme iron of the NADPH oxidase. On the basis of observations for cytochrome c oxidase, we propose a model for this dual role of histidine 115.

NADPH oxidase subunit gp91phox: a proton pathway.

The generation of superoxide by the NADPH oxidase is an electrogenic process resulting in a rapid depolarisation of the membrane potential of the cell. The efflux of H+ ions through an arachidonate-activatable, Zn(2+)-inhibitable H+ pathway accompanies the efflux of electrons and provides the necessary charge compensation. Inhibition of H+ flux leads to inhibition of superoxide generation. The protein gp91phox, a transmembrane component of the NADPH oxidase, was demonstrated to be capable of acting as the NADPH oxidase-associated H+ channel in a stable CHO cell line, CHO91. The N-terminal 230 amino acids contain all that is required for the protein to form an H+ channel and specifically histidine 115 is important to the ability of gp91phox to conduct H+ ions. The recording of outward currents from CHO91 cells, in the whole-cell configuration, demonstrated that gp91phox is also capable of functioning as a voltage-gated H+ conductance pathway. The similarity in properties between voltage-elicited outward currents, from both wild type and the mutations, and the arachidonate-activated H+ flux strongly suggests that these H+ pathways are one in the same. Among the recently identified homologues of gp91phox only NOH-1S has so far been demonstrated to also act as an H+ conductance pathway.

Identifying and characterizing one-time and never users of mammography screening services among medicare eligible women.

BACKGROUND: Despite eligibility for a screening mammogram once every 2 years from 1991 to 1997, only a small percentage of Medicare women utilized this benefit. We examined mammography use among 388,707 North Carolina Medicare women from 1994 to 1997 to identify characteristics of one-time and never users of mammography. METHODS: Data were obtained from North Carolina Medicare mammography claims and enrollment files from 1994 to 1997. Women ages 65+ as of 01/01/1994, continuously enrolled in Medicare from 1994 to 1997, and alive as of 12/31/1997 were included in the sample (n = 388,707). Mammogram use was categorized as never, once, or at least twice during 1994/1995 and 1996/1997. Women with at least one mammography claim during 1994/1995 and at least one mammography claim during 1996/1997 were called repeat users, women with one mammography claim during the 4 years were labeled one-time users, and women with zero mammography claims during the 4 years were termed never users. Multivariate logistic regression analyses were conducted to determine associations between characteristics and mammography frequency. RESULTS: Biennial mammography claims data rates were 35.3% in 1994/1995 and 41.8% in 1996/1997. Compared with all other users, one-time users (n = 108,899) were more likely to be ages 65-74 (vs 75-84 and 85+), live in an urban versus rural county, and be eligible for Medicare only versus Medicare and Medicaid. Never users (n = 184,545) were more likely to be ages 85+, be non-Caucasian, live in a rural county, and be eligible for both Medicare and Medicaid versus Medicare. CONCLUSIONS: These results demonstrate different demographic characteristics for one-time and never mammography users. This approach of using multiple years of claims data to segment the targeted population provides the opportunity to tailor interventions to subgroups.

Surface electrode somatosensory-evoked potentials in spinal surgery: implications for indications and practice.

STUDY DESIGN: A retrospective study of 442 major spinal operations with spinal cord monitoring performed in a University Hospital between 1982 and 1992 was performed. OBJECTIVES: To assess the reliability of the authors' method for monitoring by somatosensory-evoked potential recording, to determine criteria for intraoperative corrective action, and to redefine the need for the wake-up test. SUMMARY OF BACKGROUND DATA: The routine use of somatosensory-evoked potential monitoring in spinal surgery remains controversial. In Nottingham, the authors have used a method of recording from either scalp or high cervical electrodes. METHODS: The recordings and outcomes of all monitored spinal operations between the years 1982 and 1992 were reviewed. RESULTS: In 442 procedures, 23 technical failures (no reliable monitoring) occurred. Most technical failures were in patients with severe preoperative neurology, identifiable by somatosensory-evoked potential recording before operation. In the remaining 419 procedures, a significant intraoperative change in response occurred in 70 procedures (16.7%). Using the definitions of the American EEG Society, the authors identified 10 true-positives and 60 false-positives. There were no false-negatives. A wake-up test was performed if an amplitude drop greater than 50% from baseline value persisted after attempts to correct any possible identifiable intraoperative cause. This occurred in only 21 patients (5%). In the true-positive group, somatosensory-evoked potential recordings remained persistently abnormal despite an apparently normal subsequent wake-up test. The sensitivity of the method according to current definitions was 100% and the specificity 85.33%. CONCLUSIONS: Modified guidelines are needed for routine intraoperative use of somatosensory-evoked potential monitoring in spinal surgery. Such guidelines should avoid the term "false-positive" as currently defined and concentrate on the causative analysis of abnormal responses that warn of critical spinal cord dysfunction before that becomes irreversible and allow for appropriate action. Information from this monitoring method alerted the surgeon to the possible need for corrective action in an additional 9.78% of the reported patients, who traditionally would have been regarded as false-positives. A wake-up test still is indicated in patients with persistent suppression of their somatosensory-evoked potential despite correction of any identifiable cause and in cases of technical failure. The current method proved flexible, versatile, and reliable for future use.

Evidence that the product of the human X-linked CGD gene, gp91-phox, is a voltage-gated H(+) pathway.

Expression of gp91-phox in Chinese hamster ovary (CHO91) cells is correlated with the presence of a voltage-gated H(+) conductance. As one component of NADPH oxidase in neutrophils, gp91-phox is responsible for catalyzing the production of superoxide (O(2).(2)). Suspensions of CHO91 cells exhibit arachidonate-activatable H(+) fluxes (Henderson, L.M., G. Banting, and J.B. Chappell. 1995. J. Biol. Chem. 270:5909-5916) and we now characterize the electrical properties of the pathway. Voltage-gated currents were recorded from CHO91 cells using the whole-cell configuration of the patch-clamp technique under conditions designed to exclude a contribution from ions other than H(+). As in other voltage-gated proton currents (Byerly, L., R. Meech, and W. Moody. 1984. J. Physiol. 351:199-216; DeCoursey, T.E., and V.V. Cherny. 1993. Biophys. J. 65:1590-1598), a lowered external pH (pH(o)) shifted activation to more positive voltages and caused the tail current reversal potential to shift in the manner predicted by the Nernst equation. The outward currents were also reversibly inhibited by 200 microM zinc. Voltage-gated currents were not present immediately upon perforating the cell membrane, but showed a progressive increase over the first 10-20 min of the recording period. This time course was consistent with a gradual shift in activation to more negative potentials as the pipette solution, pH 6.5, equilibrated with the cell contents (reported by Lucifer yellow included in the patch pipette). Use of the pH-sensitive dye 2'7' bis-(2-carboxyethyl)-5(and 6) carboxyfluorescein (BCECF) suggested that the final intracellular pH (pH(i)) was approximately 6.9, as though pH(i) was largely determined by endogenous cellular regulation. Arachidonate (20 microM) increased the amplitude of the currents by shifting activation to more negative voltages and by increasing the maximally available conductance. Changes in external Cl(-) concentration had no effect on either the time scale or the appearance of the currents. Examination of whole cell currents from cells expressing mutated versions of gp91-phox suggest that: (a) voltage as well as arachidonate sensitivity was retained by cells with only the NH(2)-terminal 230 amino acids, (b) histidine residues at positions 111, 115, and 119 on a putative membrane-spanning helical region of the protein contribute to H(+) permeation, (c) histidine residues at positions 111 and 119 may contribute to voltage gating, (d) the histidine residue at position 115 is functionally important for H(+) selectivity. Mechanisms of H(+) permeation through gp91-phox include the possible protonation/deprotonation of His-115 as it is exposed alternatively to the interior and exterior faces of the cell membrane (see Starace, D.M., E. Stefani, and F. Bezanilla. 1997. Neuron. 19:1319-1327) and the transfer of protons across an "H-X-X-X-H-X-X-X-H" motif lining a conducting pore.

Role of histidines identified by mutagenesis in the NADPH oxidase-associated H+ channel.

The efflux of protons through a H+ channel acts as the charge compensation pathway for the electrogenic generation of superoxide (O-2) by human neutrophil NADPH oxidase. It has previously been shown that the N-terminal 230 amino acids of the product of the X-linked chronic granulomatous gene gp91(phox) contain all that is required for it to function as the arachidonate-activable, NADPH oxidase-associated H+ channel (Henderson, L. M., Thomas, S., Banting, G., and Chappell, J. B. (1997) Biochem. J. 325, 701-705). To identify functionally important amino acids, Chinese hamster ovary (CHO) cell lines were constructed that expressed point mutations in the N terminus of gp91(phox). No H+ flux was observed in CHO cell lines expressing the N-terminal gp91(phox) mutants H111L, H115L, and H119L, or H115L, or H115K. Partial retention of H+ channel function was, however, observed in the H115D CHO cell line. The addition of arachidonic acid to R91E,R92E CHO cells elicited a full H+ channel response. The buffering capacity and response of 2', 7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein to H+ were the same in all cell lines. Therefore, it can be concluded that His-115 is important to the ability of gp91(phox) to function as the NADPH oxidase-associated H+ channel and that the mechanism of H+ conduction involves protonation and deprotonation of an amino acid with an appropriate pK value.

Aneuploidy: a report of an ECETOC task force.

Aneuploidy plays a significant role in adverse human health conditions including birth defects, pregnancy wastage and cancer. Although there is clear evidence of chemically induced aneuploidy in experimental systems, to date there are insufficient data to determine with certainty if chemically induced aneuploidy contributes to human disease. However, since there is no reason to assume that chemically induced aneuploidy will not occur in human beings, it is prudent to address the aneugenic potential of chemicals in the safety assessment process. A wide range of methods has been described for the detection of chemically induced aneuploidy including subcellular systems, tests with fungi, plants and Drosophila as well as in vitro mammalian systems and in vivo mammalian somatic and germ cell assays. However, none of these methods is sufficiently validated or widely used in routine screening. Underlying the efforts to develop aneuploidy-specific assays is the presumption that current genetic toxicology tests do not detected chemicals that have aneuploidy-inducing potential. To address this, we have critically evaluated data from standard genetic toxicology assays for 16 known or suspected aneugens. The conclusions from the review are listed below. 1. At present there are only nine chemicals that can be classified as definitive aneugens, as determined by positive results in in vivo rodent assays. 2. As expected, the majority of definitive and suspected aneugens are negative in the bacterial mutation assay. 3. The majority of definitive aneugens evaluated induce polyploidy in vitro. With few exception, they also induced structural chromosome aberrations in vitro. 4. All of the definitive aneugens that have been sufficiently tested induce micronuclei in rodent bone marrow cells in vivo. A number of these chemicals also induced structural chromosome aberrations in vivo. 5. There is no evidence for a unique germ cell aneugen, that is a chemical that induces aneuploidy in germ cells and not in somatic cells. Furthermore, an analysis of several databases indicates the proportion of chemicals which induce polyploidy and not chromosome aberrations in vitro is low. Based on these conclusions, the following recommendations are made: for screening purposes, a standard genotoxicity test battery (including an in vitro cytogenetic assay with an assessment of polyploidy and clastogenicity at the same harvest time) should be performed; in the absence of polyploidy induction in vitro no further evaluation of aneuploidy-inducing potential is needed; if polyploidy is observed, in vitro follow-up testing to investigate further the aneuploidy-inducing potential should be conducted; such follow-up testing will generally start with the conduct of a standard in vivo somatic cell micronucleus assay; if the in vivo somatic cell micronucleus assay is negative, with adequate evidence of exposure of the bone marrow to the test compound, no further testing of aneuploidy-inducing potential is needed; if the in vivo somatic cell micronucleus assay is positive, further information on mechanisms of micronucleus induction can be obtained by using kinetochore/centromeric staining in vitro and/or in vivo; an assessment of potential germ cell aneuploidy activity may then be considered; aneuploidy induction which does not involve the direct interaction of a chemical or its metabolite(s) with DNA is expected to have a threshold. This must be considered in the risk assessment of such chemicals; this is not addressed by current risk assessment guidelines.

The arachidonate-activatable, NADPH oxidase-associated H+ channel is contained within the multi-membrane-spanning N-terminal region of gp91-phox.

The generation of superoxide by the NADPH oxidase of neutrophils is accompanied by the efflux of H+ ions through a H+ channel. gp91-phox, a protein component of the oxidase, has been shown previously to function as a H+ channel [Henderson, Banting and Chappell (1995) J. Biol. Chem. 270, 5909-5916]. We have constructed a CHO cell line (CHO-N) that expresses an N-terminal fragment of gp91-phox containing the predicted multiple transmembrane domains of the protein. These cells exhibit H+ fluxes in response to an imposed proton motive force and in the presence of arachidonate (to open the channel). The H+ fluxes were indistinguishable from those observed in cells expressing full-length gp91-phox. Therefore the N-terminal 230 amino acids of gp91-phox contain all that is required to function as the NADPH oxidase-associated H+ channel.

The arachidonate-activable, NADPH oxidase-associated H+ channel. Evidence that gp91-phox functions as an essential part of the channel.

The human neutrophil NADPH oxidase-associated H+ channel acts as a charge compensator for the electrogenic generation of superoxide (O2-.). The expression of the channel activity was found to increase in parallel with that of the stimulatable generation of O2-. in differentiated HL60 cells. HL60 cells induced to differentiate in the presence of succinyl acetone (a inhibitor of heme synthesis) were unable to generate O2-., failed to express p22-phox but retained H+ channel activity. EBV transformed B lymphocyte cell lines from normal and CGD patients lacking expression of either p47-phox or p67-phox all expressed unaltered channel activity; however, the activity was completely absent in the lymphocyte cell line lacking gp91-phox. CHO cells and undifferentiated HL60 cells transfected with gp91-phox cDNA expressed H+ channel activity correlating with the expression of gp91-phox. We therefore conclude that the large subunit of the NADPH oxidase cytochrome b (gp91-phox) is the arachidonate activable H+ channel of human neutrophils.

Assembly of contact-phase factors on the surface of the human neutrophil membrane.

H-kininogen (HK), a major factor involved in contact-phase activation, was recently immunolocalized on the external surface of human neutrophils. Experiments were, therefore, designed to consider the question of whether the complete assembly of contact factors occurs on the outer surface of the neutrophil membrane. By immunolocalization techniques, and using specific antibodies directed against the various contact factors, we now demonstrate that plasma prekallikrein (PK), factor XI (FXI), and factor XII (FXII) are present on the exterior face of the human neutrophil. Failure to localize HK, PK, or FXI by monoclonal antibodies directed to their reciprocal binding sites, and displacement of PK/FXI by peptide HK31, which mimics the relevant binding site(s) of HK, suggested that prekallikrein and FXI are anchored to the neutrophil membrane through attachment to the kininogen molecule. Probing of the kinin moiety by a specific antibody showed that kininogen molecules bound to the neutrophil cell membrane contain the kinin sequence, which can be released by plasma kallikrein or by tissue kallikrein. Our results led us to the novel conclusion that neutrophils provide a circulating platform for the components of the contact-phase system.

Recovery of Aujeszky's disease virus recombinants from experimentally co-infected swine.

Tissue homogenates were obtained from swine experimentally co-infected with two vaccine strains of Aujeszky's disease virus (ADV). Viral isolates were derived by serial plaque purification directly from tissue homogenates, without an intervening step of isolation and amplification on cell cultures. Use of limiting dilutions and recovery of virus isolates as individual plaques minimized the likelihood of in vitro recombination serving as a confounding source of recombinant ADV. The tyhmidine kinase and glycoprotein X gene sequences were classified as wildtype or deleted, using a battery of polymerase chain reaction assays. On the basis of pairwise combinations of the allelic forms of the thymidine kinase and glycoprotein X genes, the isolates were characterized as recombinant and parental genotypes. The results substantiate the observation that ADV vaccine strains can form genetic recombinants in vivo after experimentally induced co-infection. A full description of this study is available (Am. J. Vet. Res. 54 (4) 540-545, 1993).

Direct isolation and identification of recombinant pseudorabies virus strains from tissues of experimentally co-infected swine.

Tissue homogenates were obtained from swine co-infected with 2 vaccine strains of pseudorabies virus (PRV). Viral isolates derived by serial plaque purification directly from tissue homogenates, without an intervening step of isolation and amplification on cell cultures, were characterized as recombinant and parental PRV genotypes on the basis of thymidine kinase and glycoprotein X gene combinations. Use of limiting dilutions and recovery of virus isolates as individual plaques minimized the likelihood of in vitro recombination serving as a confounding source of recombinant PRV. The thymidine kinase and glycoprotein X gene sequences were classified as wild-type or deleted, using a battery of polymerase chain reaction assays. Results substantiate the observation that PRV vaccine strains can form genetic recombinants in vivo after experimentally induced co-infection.

Immunovisualization of high (HK) and low (LK) molecular weight kininogens on isolated human neutrophils.

An immunocytochemical study was performed to examine the cellular localization and the subcellular distribution of kininogens in human blood cells. Kininogens were visualised using the immunogold-silver staining method and confocal scanning laser microscopy. We confirmed the existence of high molecular weight kininogen in human neutrophils and describe for the first time the presence of low molecular weight kininogen on these cells. Both high and low molecular weight kininogens were restricted to the neutrophils where they localized as clusters of immunogold particles on the cell membrane. No labeling was observed intracellularly in organelles such as mitochondria, endoplasmic reticulum, and azurophilic or specific granules after permeabilization of the neutrophils with Triton X-100, a procedure that permitted the visualization of elastase in the azurophilic granules. Clusters of high molecular weight kininogen molecules attached to the neutrophil surface could serve as receptors for plasma kallikrein and/or be the source of substrate for a discrete and circumscribed formation of kinins that may in turn facilitate the local diapedesis of neutrophils and the transudation of plasma constituents during acute inflammation.

Immunovisualisation of plasma prekallikrein and H-kininogen on human neutrophils and in human hepatocytes.

Both plasma prekallikrein and H-kininogen were immunolocalised in human hepatocytes by the use of immunocytochemical techniques in conjunction with the confocal optical scanning microscopy. In contrast, both proteins were demonstrated on the external surface of human blood neutrophils. However, detection of H-kininogen on non-fixed but not on fixed neutrophils with the anti-domain 6 antibody (directed at the prekallikrein binding site on H-kininogen), suggested that access to the epitope was blocked by the presence of the bound plasma prekallikrein. Therefore, we prepose that H-kininogen provides the binding site for plasma prekallikrein on circulating neutrophils.

Superoxide generation by EBV-transformed B lymphocytes. Activation by IL-1 beta, TNF-alpha and receptor independent stimuli.

The generation of superoxide by Epstein-Barr virus (EBV)-transformed human B lymphocytes can be stimulated by a range of compounds; receptor-dependent stimuli include tumour necrosis factor-alpha (TNF-alpha), interleukin-1 beta (IL-1 beta) and lipopolysaccharides (LPS), and independent stimuli include AlF3, A21387 and ionomycin. The stimuli suggest that the activation pathway for the lymphocyte oxidase is similar to that proposed for the neutrophil oxidase. Although the rate of superoxide production was lower than that by neutrophils, the respiratory burst was much prolonged. It is possible that this superoxide generation by lymphocytes may have a biological function.

Uptake of L-carnitine, D-carnitine and acetyl-L-carnitine by isolated guinea-pig enterocytes.

Uptake and metabolism of L-carnitine, D-carnitine and acetyl-L-carnitine were studied utilizing isolated guinea-pig enterocytes. Uptake of the D- and L-isomers of carnitine was temperature dependent. Uptake of L-[14C]carnitine by jejunal cells was sodium dependent since replacement by lithium, potassium or choline greatly reduced uptake. L- and D-carnitine developed intracellular to extracellular concentration gradients for total carnitine (free plus acetylated) of 2.7 and 1.4, respectively. However, acetylation of L-carnitine accounted almost entirely for the difference between uptake of L- and D-carnitine. About 60% of the intracellular label was acetyl-L-carnitine after 30 min, and the remainder was free L-carnitine. No other products were observed. D-Carnitine was not metabolized. Acetyl-L-carnitine was deacetylated during or immediately after uptake into intestinal cells and a portion of this newly formed intracellular free carnitine was apparently reacetylated. L-Carnitine and D-carnitine transport (after adjustment for metabolism and diffusion) were evaluated over a concentration range of 2-1000 microM. Km values of 6-7 microM and 5 microM, were estimated for L- and D-carnitine, respectively. Ileal-cell uptake was about half that found for jejunal cells, but the labeled intracellular acetylcarnitine-to-carnitine ratios were similar for both cell populations. Carnitine transport by guinea-pig enterocytes demonstrate characteristics of a carrier-mediated process since it was inhibited by D-carnitine and trimethylaminobutyrate, as well as being temperature and concentration dependent. The process appears to be facilitated diffusion rather than active transport since L-carnitine did not develop a significant concentration gradient, and was unaffected by ouabain or actinomycin A.

Absorption and metabolism of adenine, adenosine-5'-monophosphate, adenosine and hypoxanthine by the isolated vascularly perfused rat small intestine.

Intestinal vascular perfusion and in vivo live animal experiments were conducted in order to evaluate the nature and extent of the intestinal metabolism of adenine, adenosine, adenosine-5'-monophosphate (AMP) and hypoxanthine in the rat. Radiolabeled purine substrates were administered intralumenally. Intestinal contents, tissue and/or portal flow were collected and evaluated for resultant purine metabolites by liquid and paper chromatography and paper electrophoresis. Adenosine, AMP and hypoxanthine were quantitatively metabolized to end products (primarily uric acid) within 15 minutes after administration. In contrast, the metabolism of adenine to uric acid was considerably slower. Up to 20% of the administered adenine was recovered unmetabolized in the portal vasculature. Nonetheless uric acid was the primary metabolite recovered from the portal circulation in the isolated intestine regardless of the purine substrate or concentration administered. Since lumenal inosine concentrations rose sharply with increasing doses of AMP, either transport or metabolism of inosine is a rate-limiting step in the intestinal metabolism of purines to uric acid in the rat. Finally, the large percentage of the radiolabel in uric acid recovered in the lumen is consistent with the hypothesis that the intestine is an extrarenal site for purine excretion.

Pyridine nucleotide synthesis in normal and neoplastic human pituitary cells in culture.

14C-Nicotinic acid (NA) incorporation into nicotinamide adenine dinucleotide (NAD) was studied in cultures from 7 normal human pituitaries and 13 chromophobe adenomas. 14C-NA (7.2 microM) was incubated with both normal and tumor cell cultures for periods up to 48 hours. Cells and culture media were examined separately for metabolites at 24 and 48 hours. NAD was the major labeled metabolite found in both normal and tumor cells, accounting for 65.4% for normal cells and 56.8% for tumor cells of the total cellular label at 48 hours. Nicotinamide (NAm), a product arising from NAD, was the only labeled metabolite found in the culture medium, aside from the 14C-NA added initially. Total incorporation of 14C-NA into NAD was estimated by adding the cellular 14C-NAD and labeled products of NAD (NMN, NAm and NADP) to the 14C-NAm found in the medium for each culture. Cultures derived from adenomas demonstrated greater than twice the rate of incorporation of 14C-NA into NAD as did normal cell cultures (P less than 0.01 at 24 hours and less than 0.001 at 48 hours). This difference did not appear to be related to the secretory status of the tumor, since both secreting and nonsecreting tumors demonstrated increased rates when compared to normal cells. This difference also persisted in two different culture media and with cells that had been maintained in culture for different lengths of time (6-day dispersed cell cultures and 6- to 16-week fragment cultures).