FACTORS INFLUENCING BRAIN AND TISSUE LEVELS OF TRYPTAMINE: SPECIES, DRUGS AND LESIONS

With a modification of the spectrophotofluorometric (SPF) method of HESS & UDENFRIEND (1959) (J. Pharmac. exp. Ther. 127 , 175-177), brain tryptamine levels in the rat (20.9 ng/g) and guinea-pig (20.7 ng/g) were found to be less than those in the dog (32.1 ng/g) and cat (52.2 ng/g). Regional distribution studies in the dog and cat showed that tryptamine was present in all major brain regions with highest concentrations in the spinal cord. Blood levels of tryptamine in the guinea-pig, dog and cat (6-7 ng/ml) were lower than brain levels. Pargyline significantly increased brain tryptamine in both the dog and cat; whereas, isocarboxazid (after 4 h) increased brain tryptamine levels in the dog but decreased brain levels in the cat. Reserpine (0.5-1.0 mg/kg per day for 1-4 days) did not significantly decrease brain, spinal cord or blood tryptamine levels in the dog. Spinal cord transection did not decrease tryptamine levels below the lesion in the chronic spinal dog.     

Activation of hypoxanthine/guanine phosphoribosyltransferase from yeast by divalent zinc and nickel ions

We have observed previously that the reactions catalyzed by hypoxanthine/guanine phosphoribosyltransferase (HGPRTase) are activated by Mg(II), Mn(II), and Co(II), and we have defined the mechanism by which these activations proceed [Biochemistry 22, 3419-3424 (1983)]. A more extensive survey of the kinds of metal ions that will activate the HGPRTase catalysis now has been completed through the use of an HPLC assay procedure. Although Fe(II) and Ca(II) are unable to activate this reaction, a significant activation was achieved with the addition of spectroscopically pure Zn(II) to the assay solution. In addition some IMP synthesis resulted from the addition of Ni(II) to the assay mixture. Both the Zn(II) and Ni(II) kinetic effects on HGPRTase over a limited metal ion concentration range have been analyzed through the use of curve-fitting exercises. These results, in addition to the similar pH profiles for the activations by Mg(II), Mn(II), Co(II), and Zn(II), suggest that all of these metal ions activate the HGPRTase-catalyzed synthesis of IMP by way of the same mechanism [model II as defined by London and Steck, Biochemistry 8, 1767-1779 (1969)], during which two divalent ions bind to the HGPRTase active site per molecule of PRibPP.     

Plasmid-encoded lysostaphin endopeptidase gene of Staphylococcus simulans biovar staphylolyticus

A 12.2-kilobase (kb) BclI fragment containing the lysostaphin endopeptidase gene was cloned from Staphylococcus simulans biovar staphylolyticus into Escherichia coli. The gene was expressed in E. coli and the gene product apparently was secreted into the periplasmic space. The gene was localized to a 3.3-kb region of the cloned fragment and this region was shown to contain a staphylococcal promoter for the endopeptidase gene. By hybridization analysis, the endopeptidase gene was shown to reside on the largest of five plasmids in S. simulans biovar staphylolyticus. No additional copies of this gene were detected in the genome.     

Pancreatic polypeptide immunoreactivity in medullary carcinoma of the thyroid: identification and characterisation by radioimmunoassay, immunocytochemistry and high performance liquid chromatography

Pancreatic polypeptide immunoreactivity has been identified in primary medullary carcinoma of thyroid using radioimmunoassay and immunocytochemistry and subsequently characterised by HPLC. Two region-specific PP antisera were used in the study; one C-terminal and one non-C-terminal. These antisera demonstrate variable cross-reactivity with the molecular species of PP identified in the tumours. The immunoreactive material in the tumours corresponded to human PP and not PYY or NPY on the basis of immunoreactivity and HPLC behaviour. It was identified in all patients with familial-type disease but not in the two sporadic cases examined. We propose that estimation of the PP content of medullary carcinoma of thyroid may be a useful means of differentiating familial and sporadic types.     

Human METTL16 is a N6‐methyladenosine (m6A) methyltransferase that targets pre‐mRNAs and various non‐coding RNAs

N⁶‐methyladenosine (m⁶A) is a highly dynamic RNA modification that has recently emerged as a key regulator of gene expression. While many m⁶A modifications are installed by the METTL3–METTL14 complex, others appear to be introduced independently, implying that additional human m⁶A methyltransferases remain to be identified. Using crosslinking and analysis of cDNA (CRAC), we reveal that the putative human m⁶A “writer” protein METTL16 binds to the U6 snRNA and other ncRNAs as well as numerous lncRNAs and pre‐mRNAs. We demonstrate that METTL16 is responsible for N⁶‐methylation of A43 of the U6 snRNA and identify the early U6 biogenesis factors La, LARP7 and the methylphosphate capping enzyme MEPCE as METTL16 interaction partners. Interestingly, A43 lies within an essential ACAGAGA box of U6 that base pairs with 5′ splice sites of pre‐mRNAs during splicing, suggesting that METTL16‐mediated modification of this site plays an important role in splicing regulation. The identification of METTL16 as an active m⁶A methyltransferase in human cells expands our understanding of the mechanisms by which the m⁶A landscape is installed on cellular RNAs.     

Characterization of the mouse liver cell line FL83B

The previously reported mouse liver cell line FL83B has been characterized more completely with respect to its light and electron microscopic appearance, chromosomal composition, and ability to secrete various serum proteins. These cells bear many striking morphological similarities to parenchymal liver cells. Chromosomal analysis showed that the cells were transformed. The ability of these cells to grow in a completely chemically defined medium permitted the unequivocal demonstration of the synthesis of at least 12 mouse serum proteins including albumin and high density lipoprotein.     

Stage of the Estrous Cycle Does Not Influence Myocardial Ischemia–Reperfusion Injury in Rats (Rattus norvegicus)

Even though cardiovascular disease is the leading cause of death for men and women, the vast majority of animal studies use male animals. Because female reproductive hormones have been associated with cardioprotective states, many investigators avoid using female animals because these hormones are cyclical and may introduce experimental variability. In addition, no studies have investigated the specific effects of the estrous cycle on cardiac ischemic injury. This study was conducted to determine whether the estrous cycle stage influences the susceptibility to ischemic injury in rat hearts. Estrous cycle stage was determined by using vaginal smear cytology, after which hearts underwent either in vivo (surgical) or ex vivo (isolated) ischemia–reperfusion injury. For in vivo studies, the left anterior coronary artery was ligated for 25 min of ischemia and subsequently released for 120 min of reperfusion. Infarct sizes were 42% ± 6%; 49% ± 4%; 40% ± 9%; 47% ± 9% of the zone-at-risk for rats in proestrus, estrus, metestrus, and diestrus, respectively. For ex vivo studies, isolated, perfused hearts underwent global ischemia and reperfusion for 25 and 120 min, respectively. Similar to our in vivo studies, the ex vivo rat model showed no significant differences in susceptibility to infarction or extent of cardiac arrhythmia according to estrous stage. To our knowledge, these studies provide the first direct evidence that the stage of estrous cycle does not significantly alter cardiac ischemia–reperfusion injury in rats.Abbreviations: VF, ventricular fibrillation; VT, ventricular tachycardiaCardiovascular disease remains the leading cause of morbidity and mortality throughout the industrialized world, with ischemic heart disease being a major manifestation of cardiovascular disease. Many investigators use animal models to advance our understanding of the etiology and mechanisms involved. Although ischemic heart disease is the leading cause of death for both men and women, the overwhelming majority of studies use male animals. Perhaps the most common reason for this practice is that physiologic fluctuations in female reproductive hormones such as estrogen may be a confounding variable, given the influence of female reproductive hormones on various organ systems.²⁵ Despite the assertion that cyclical variations in female reproductive hormones may confound experimental studies, few data are available that support estrous-cycle–dependent variations in susceptibility to ischemic heart injury.Epidemiologic studies suggest that, compared with men, women have lower cardiac mortality prior to undergoing menopause.⁴⁰ Consistent with human studies, experimental models in several species commonly show that the degree of cardiac injury in young female animals is lower than that in male counterparts.^7,9,21,22,42 Exogenous administration of estrogen has a clear effect in reducing injury,^14,15 but whether endogenous cyclical variations in female reproductive hormones affect cardiac injury is not known.Rats and mice are commonly used species to examine cardiac ischemia–reperfusion injury. Unlike humans, rodents do not undergo menstruation, during which the uterine endometrium sloughs off and is expelled through the vagina, but rather the uterine lining of rodents is reabsorbed during an estrous cycle.²⁴ The rat estrous cycle is typically 4 to 5 d in length and is defined by 4 separate stages: proestrus, estrus, metestrus, and diestrus. Proestrus is characterized by increasing levels of estrogen. At the end of proestrus, ovulation (signaled by luteinizing hormone) occurs and marks the beginning of the estrus cycle. During metestrus and diestrus, the uterine lining regenerates, and the cycle starts again.^24,33 These stages induce changes in the composition of the epithelium of the vagina and the presence of inflammatory cells, which can easily be detected by using vaginal cytology.^18,35We conducted the current study to determine whether estrous cycle stage influences the susceptibility to ischemia–reperfusion injury in the rat heart. Because the stage of the estrous cycle may influence cardiac injury either directly (via a direct effect of circulating hormones), or indirectly (by inducing changes that are intrinsic to the heart), we used both in vivo and ex vivo models of injury.     

Immunomodulatory Agents as Adjunctive Therapy for the Treatment of Resistant Candida Species

Invasive Candida infections have increased fivefold over the past 20 years. During this time, the incidence of antifungal resistance and infection due to non-albicans species has risen with the increasing use of broad spectrum antifungals. As few new antifungal agents are in development, strategies to improve outcomes in the treatment of Candida infections are sorely needed. The use of immunotherapy to augment the host immune response as an adjunctive treatment for Candida infections is a potentially robust and promising approach. The purpose of this review is to focus on new developments in the use of adjunctive immunotherapy for the treatment of Candida infections, and discuss the potential impact of antifungal resistance on the host immune response.