Biomedical subjects
Publications and source records attributed to L Yu.
Four kinds of Hawthorn Fruit, namely Large Chinese Hawthorn, Chinese Hawthorn, Nippon Hawthorn and Yunnan Hawthorn, were compared in animal experiments. Their effects on small intestine motion, myocardial ischemia and hypercholesterolemia have been found different from each other.
We studied the usefulness of serum ferritin for the assessment of iron deficiency (ID) or ID anaemia (IDA) of 186 lactating and 27 non-lactating Zairean women (15-45 years old). Haemoglobin (Hb), serum iron (SI), total iron binding capacity (TIBC), and transferrin saturation (TS) were also measured. Participants were recruited in rural Bas-Zaire State in the summers of 1986 and 1989. Serum ferritin ranged from 10 to 360 micrograms l-1 (median 62 micrograms l-1) in lactating women and from 14.2 to 120 micrograms l-1 (median 40 micrograms l-1) in non-lactating women. While mean levels of serum ferritin and TS were within the normal range in both groups of women, those of Hb were below normal (< 12 g dl-1), partly due to inflammation. The prevalence of anaemia was 66% in lactating women and 70% in non-lactating women, and did not change with time. It was higher in women with inflammation than in those without inflammation. Although ID (ferritin < 12 micrograms l-1) was almost absent, after raising the cut-off point of ferritin to 50 micrograms l-1 in women with inflammation, it was present in 28.8% of lactating women and 52% of non-lactating women. While the prevalence of ID assessed by serum ferritin was significantly higher in lactating women studied in 1989 (40.5%) than in those studied in 1986 (13.5%), it was similar in both groups of non-lactating women. ID defined by TS < 16% was present in 41% of lactating women and 21% of non-lactating women. In the presence as well as absence of inflammation, the use of TS identified a higher percentage of lactating women with either ID or IDA than did the use of serum ferritin. We conclude that, in the studied population, unless inflammation is taken into consideration, serum ferritin has a limited value in the diagnosis of ID.
Acute phase proteins (APP) are plasma proteins secreted by the liver following inflammation. APP reference levels are unknown for healthy female Zaireans. The purpose of this study was to determine: (a) the reference levels of alpha 1-acid glycoprotein (AGP), C-reactive protein (CRP), ceruloplasmin (Cp), and haptoglobin (Hp) in healthy Zairean adult women, and (b) whether the levels are different between lactating (Group I), pregnant (Group II) and non-pregnant/non-lactating women (Group III). APP were measured by radial immunodiffusion in 180, 212 and 27 women of Groups I, II, and III respectively (age range 15-45 years). The ranges for different APP were wider than for Western adults or those obtained in plasma from normal laboratory personnel. The means and medians of AGP were lower in pregnant that non-pregnant women, and those of CRP and Cp were higher. There were no differences in mean Hp levels between the three groups. In non-pregnant women, the mean levels and 95% confidence limits of CRP, Cp and Hp were within published ranges for healthy Western adults, while those of AGP were higher. In pregnant women only AGP and Hp were within published ranges. We conclude that, at least in non-pregnant women, the same cut-off points of CRP, Cp and Hp used for Western adults may also be used in the assessment of inflammation in Zairean women.
Tumours of mice are induced by administration of Inj. Hydroxyprogesteroni Caproatis Co. (EP) in a practical subthreshold dose of carcinogenesis or 2.5-5 times the human contraceptive dose (simply referred to as 2.5- to 5-fold dose) combined with whole-body 0.5 Gy gamma-ray irradiation. Malignant transformation of Syrian golden hamster embryo (SHE) cells is also induced by 5-fold dose of EP combined with 0.3 Gy gamma-ray irradiation in vitro, thereby indicating that synergistic carcinogenesis can be obtained by combined use of physical and chemical carcinogens. The mechanisms of synergistic carcinogenesis have been further explained by cytogenetics, damage extent of the target cell DNA and production of free radicals. The Chinese traditional medicine with antioxidating effect (Sulekang Capsule, SC), food additive--butylated hydroxyanisole (BHA) and green tea can effectively inhibit the carcinogenic effect of EP or EP combined with gamma rays in mice. They all have marked ability to scavenge or remove the free radicals and thereby reduce the DNA damage.
The effect of nasal mask CPAP administered during night in four central hypoventilation patients with various causes were investigated. On the basis of their histories, specific clinical manifestations, cranial CT and MRI, cerebral angiography and polysomnography. The patients (three males and one female) are diagnosed as primary alveolar hypoventilation and central hypoventilation, the later mainly caused by the compression of bulbar respiratory centre due to congenital deformity of right vertebral artery and deformity of foramen magnum. In order to assess the severity of nocturnal hypoventilation and blood oxygen level of the patients, polysomnography was taken before treatment, during 2L/min of oxygen via nasal tube and CPAP ventilation. Only oxygen administration did not ameliorate hypoventilation episodes and hypoxemia, lowest SaO2 is less than 50% on four patients, hypoventilation index is greater than 20 times/hour in three patients and 6.4 in a patient. Applying CPAP with the pressure fixed to 6-12 cm H2O can significantly improve hypoventilation index and hypoxemia with lowest SaO2 more than 70% (P < 0.001) and hypoventilation index less than 7 (P < 0.01). We conclude that nocturnal nasal mask CPAP is an effective treatment for central hypoventilation.
Opioid drugs, such as morphine, and the endogenous opioid peptides, namely the enkephalins, endorphins, and dynorphins, exert a wide spectrum of physiological and behavioral effects, including effects on pain perception, mood, motor control, and autonomic functions. These effects are mediated via membrane-bound receptors, of which the best characterized are the kappa, delta, and mu receptors. The existence of these distinct types of opioid receptors has recently been confirmed by molecular cloning. In the present study, we have examined the pharmacological profiles of the cloned kappa, delta, and mu receptors using a battery of widely employed opioid agents. Our results suggest that the cloned kappa and mu receptors have pharmacological characteristics similar to those of the endogenously expressed kappa 1 and mu receptors, respectively. The cloned delta receptor displays a pharmacological profile consistent with that of a delta 2 receptor. Opioid agents with abuse potential possess high affinities for the mu receptor. The availability of the cloned receptors will facilitate the identification and development of more specific and selective compounds with greater therapeutic potential and fewer undesirable side effects.
In Xenopus oocytes, Ca2+ influx through an endogenous voltage-gated Ca2+ channel activates a transient outward Cl- current (ICl(Ca)), which is potentiated by cAMP increase. The site of cAMP effect appears to be the Ca2+ channel instead of the Ca(2+)-activated Cl- channel, because cAMP potentiates the Ba2+ current through the Ca2+ channel in a similar way to the ICl(Ca), and cAMP does not potentiate the Ca(2+)-dependent Cl- current in cells treated with Ca2+ ionophore. Using the catalytic subunit of protein kinase A (PKA) and PKA inhibitors, it was shown that PKA is both necessary and sufficient for the cAMP effect on ICl(Ca). Furthermore, the cAMP/PKA-mediated potentiation of ICl(Ca) was inhibited by both type 1 and type 2A protein phosphatases.
The reactions of a beef heart cytochrome c1 preparation containing the hinge protein with horse cytochrome c derivatives labeled at specific lysine amino groups with (dicarboxybipyridine)(bisbipyridine)ruthenium(II) (Ru(II)) were studied by flash photolysis. All of the ruthenium-cytochrome c derivatives formed complexes with cytochrome c1 in low ionic strength buffer (5 mM sodium phosphate, pH 7). Excitation of Ru(II) to Ru(II*) with a 0.4-microseconds laser flash resulted in rapid electron transfer to the ferric heme group in cytochrome c, followed by electron transfer from the ferrous heme group of cytochrome c to the ferric heme group of cytochrome c1. The kinetic difference spectra displayed maxima at 546 nm and minima at 554 nm characteristic of electron transfer between the two cytochromes. The rate constants were independent of concentration at low ionic strength, indicating intracomplex electron transfer. The rate constants were 4,800, 6,800, 22,000, and 22,000 s-1 for cytochrome c derivatives modified at lysines 13, 27, 25, and 72, respectively. The observed rate constants were independent of ionic strength up to about 50 nM and then decreased progressively with further increases in ionic strength indicating dissociation of the complex. Second-order kinetics were observed at 310 mM ionic strength, with rate constants of 1.0 x 10(6), 1.6 x 10(7), 1.2 x 10(8), and 3.0 x 10(7) M-1 s-1 for the derivatives modified at lysines 13, 27, 25, and 72, respectively. The ionic strength dependence of the second-order rate constants is comparable to that involving native horse cytochrome c and is consistent with electron transfer reactions between oppositely charged proteins.
By screening a rat brain cDNA library using a cloned mu opioid receptor cDNA as probe, a clone was identified that is very similar to both the mu and delta opioid receptor sequences. Transient expression of this clone in COS-7 cells showed that it encodes a kappa opioid receptor, designated KOR-1, which is capable of high-affinity binding to kappa-selective ligands. Treatment of transfected cell membranes with bremazocine, a kappa-selective agonist, resulted in a 53% decrease in adenylate cyclase activity, indicating that this kappa opioid receptor displays inhibitory coupling to adenylate cyclase. Thus, one member from each of the three opioid receptor types, mu, kappa and delta, has been molecularly cloned. Analysis of sequence similarities among these opioid receptors, as well as between opioid receptors and other G-protein-coupled receptors, revealed regions of sequence conservation that may underlie the ligand-binding and functional specificities of opioid receptors.
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Resonance Raman spectroscopy (RRS) has been employed to characterize cytochromes c1 isolated from bc1 complexes of beef heart mitochondria and Rhodopseudomonas sphaeroides. The data obtained in this study extend the physical characterization of cytochromes c1 and focus on the effects of the local protein environment on the heme active site. While the general characteristics of the cytochromes c1 are similar to those of smaller soluble cytochromes c, the behavior of several core-size and ligation-sensitive heme modes reveal that significant systematic differences exist between those species. These, most likely, result from changes in the heme axial-ligand interactions.
Galactosyl beta-1,3-N-acetyl galactosamine (Gal beta-1,3-GalNAc) (Thomsen Friedenreich antigen), the Class I core sequence in O-linked oligosaccharide chains, behaves as an oncofetal antigen showing increased expression in many epithelial malignancies. Previous work has shown that peanut agglutinin (PNA), a lectin that binds Gal beta-1,3-GalNAc, stimulates proliferation in HT-29 (human colon cancer) cells and normal human colonic epithelium and this implies that cell surface glycoproteins which express Gal beta-1,3-GalNAc may play an important role in the regulation of epithelial cell proliferation. We have now studied the effect on epithelial cells of another dietary Gal beta-1,3-GalNAc-binding lectin, the edible mushroom Agaricus bisporus lectin (ABL). This differs from PNA in its ability to bind also to sialylated Gal beta-1,3-GalNAc. In contrast to PNA, ABL (25 micrograms/ml) inhibited incorporation of [3H]-thymidine into DNA of HT29 colon cancer cells by 87% (95% confidence limit, 85-89%), Caco-2 colon cancer cells by 16% (95% confidence limit, 12-20%), MCF-7 breast cancer cells by 50% (95% confidence limit, 47-52%), and Rama-27 rat mammary fibroblasts by 55% (95% confidence limit, 51-60%) when these cells were grown for 24 h in serum-free medium. When assessed by cell count, similar inhibition of proliferation of HT29 cells by ABL was found. In the presence of 2% fetal calf serum (which contains the ABL-binding glycoprotein fetuin), the inhibitory effect of ABL on cell proliferation was still demonstrable but at increased ABL concentration (60 micrograms/ml for 49% inhibition). Ten micrograms/ml ABL completely abolished the stimulatory effect on [3H]thymidine incorporation of epidermal growth factor (100 pg/ml) and PNA (25 micrograms/ml) and markedly inhibited the stimulatory effect of insulin (50 ng/ml). ABL (0.2 mg/ml) caused no cytotoxicity to HT29, MCF-7, and Rama-27 cells as measured by trypan blue exclusion, and inhibition of proliferation in HT29 cells caused by 50 micrograms/ml ABL was reversible after removal of the lectin. Binding studies with 125I-labeled ABL suggested a single class of binding site with an apparent Kd value of (4.12 +/- 0.29) x 10(-7) M with (3.6 +/- 0.3) x 10(7) binding sites/cell. A. bisporus lectin is a reversible noncytotoxic inhibitor of epithelial cell proliferation which deserves study as a potential agent for cancer therapy.
Several new phosphonate-containing phospholipid analogues were synthesized as inhibitors of cobra venom (Naja naja naja) phospholipase A2. These phospholipid analogues contained a novel thioether at the sn-1 position, a tetrahedral phosphonate moiety in place of the scissile ester bond at the sn-2 position, and several different polar head groups, including phosphocholine, phospho(N,N-dimethylethanolamine), phospho(N-methylethanolamine), and phosphoethanolamine. The affinities of these analogues for the enzyme were evaluated in the well-defined Triton X-100 mixed micelle system using thio-PC and thio-PE substrates. These phosphonates inhibited thio-PC hydrolysis with very similar potencies. Inhibition of phospholipase A2 by phosphonates is known to be pH-dependent [Yu, L., & Dennis, E. A. (1991) Proc. Natl. Acad. Sci. U.S.A. 88, 9325-9329]. At pH 5.5, all of the new analogues had IC50s of about 2 x 10(-5) mol fraction. At this pH, these inhibitors are the most potent reversible inhibitors of phospholipase A2 reported to date. In contrast, at pH 8.5, the PE analogue was a potent inhibitor of thio-PC hydrolysis (IC50 1.8 x 10(-3) mol fraction) but was a very poor inhibitor of thio-PE hydrolysis (IC50 is not detectable). However, the inhibition of thio-PE hydrolysis was dramatically enhanced when the enzyme was activated by sphingomyelin, suggesting that the phosphonate inhibitors bind much more tightly to the activated enzyme than to the nonactivated enzyme. The activation and inhibition of the enzyme have different pH dependencies; the enzyme activation is not pH-dependent, whereas the enzyme inhibition is pH-dependent. These results confirm the presence of a functionally distinct activator site on this enzyme.
A selected ion monitoring method for determination of the tobacco alkaloids anabasine, anatabine, nornicotine, metanicotine, dihydrometanicotine, and 2,3'-bipyridyl in urine of smokers and smokeless tobacco users is described. The method involves conversion of the secondary amine alkaloids to tertiary amine derivatives by reductive alkylation using an aldehyde and sodium borohydride, and chromatography on a 5% phenylmethylsilicone capillary column. These derivatives have good chromatographic properties, allowing determination of concentrations as low as 1 ng/ml. The alkaloid 2,3'-bipyridyl is unaffected by the derivatization procedure and may be determined simultaneously with the other alkaloids. The structural analogues 2-(3-pyridyl)hexahydroazepine, 5-methyldihydrometanicotine, and 6-methyl-2,3'-bipyridyl were synthesized for use as internal standards. Using the method, concentrations and 24 h excretion of anabasine, anatabine, and nornicotine in urine of twenty-two smokers, eight chewing tobacco users, and six oral snuff users were determined and compared with concentrations and excretion of nicotine and its metabolite cotinine. Excretion of nicotine and cotinine was similar in all tobacco users, but excretion of anabasine, anatabine and nornicotine was substantially greater in urine of smokeless tobacco users, presumably due to absence of pyrolysis of these alkaloids in smokeless tobacco products.
In a previous paper we showed that the C51D mutant of PsaC contains a [3Fe-4S] cluster in the FA site and a [4Fe-4S] cluster in the FB site and that the C14D mutant contains an uncharacterized cluster in the FB site and a [4Fe-4S] cluster in the FA site [Zhao, J. D., Li, N., Warren, P. V., Golbeck, J. H., & Bryant, D. A. (1992) Biochemistry 31, 5093-5099]. In this paper we describe the electrochemical and electron spin resonance properties of the recombinant C14D and C51D holoproteins after in vitro reinsertion of the iron-sulfur clusters. Unbound PsaC shows no significant resonances in the oxidized state, but the unbound C14D and C51D mutant proteins show an intense set of resonances at g approximately 2.02 and 1.99 characteristic of an oxidized [3Fe-4S]1+/0 cluster. The Em' values for the [3Fe-4S]1+/0 clusters in C14D (FB*) and C51D (FA*) are -98 mV, and both represent one-electron transfers. After reduction with dithionite at pH 10.0, wild-type PsaC shows a broad set of resonances resulting from the superposition of FA- and FB- characterized by a low-field peak at an apparent g value of 2.051 and a high-field trough at an apparent g value of 1.898. The FB resonances in C51D were slightly narrower, with a low-field peak at an apparent g value of 2.039 and high-field trough at an apparent g value of 1.908.(ABSTRACT TRUNCATED AT 250 WORDS)
The nicotinic acetylcholine receptor (AChR) was expressed in Xenopus oocytes from in vitro transcribed mRNA and was imaged by atomic force microscopy. A characteristic pentameric structure of AChR was readily observed on the extracellular face of the cell membrane, with a central pore surrounded by protruding AChR subunits. These structures were seen only in mRNA-injected oocytes that also gave acetylcholine-induced membrane currents. The size of individual AChR channels, the angles between subunits, and the interchannel spacing were all compatible with the current model of AChR. In addition, localized patches of microscopic AChR clustering were observed, with packing density approaching that at the neuromuscular junction. These findings show the potential of studying cloned membrane proteins in oocytes for both their surface topography and their structure-function relationship in native membrane without the need for crystallization.
We characterized the cell-free activating system of the superoxide (O2-)-producing NADPH oxidase of pig neutrophils. Activation of the oxidase required both the membrane and cytosolic fractions in the presence of sodium dodecyl sulfate. Chromatography on 2',5'-ADP-Sepharose resulted in separation of the cytosolic fraction into two fractions, the flow-through and bound fractions, which synergistically supported the O2- production with the membrane fraction in the absence of guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S), whereas only the bound fraction besides the membrane fraction was required for the activation in the presence of GTP gamma S. The effective factors in the bound fraction were further purified by gel filtration on Superdex G-200 and anion exchange chromatography on Mono Q and found to be p47-phox and p63-phox. The purified recombinant p47-phox and p65-phox replaced corresponding native factors for the activation. These results suggest that the membrane fraction from pig neutrophils contains the GTP-binding protein responsible for the activation. Furthermore, the presence of the GTP-binding protein for the activation in the flow-through fraction from 2',5'-ADP-Sepharose was also shown on the basis of the findings that extensive dialysis of the flow-through fraction resulted in complete loss of the ability to activate the oxidase with the recombinant factors and the washed membrane of human neutrophils which contained no GTP-binding protein for the activation and the lost ability was recovered by the addition of GTP gamma S. Thus, activation of the oxidase in the cell-free system of pig neutrophils absolutely requires the GTP-binding protein which localizes in the membrane fraction or in the cytosolic fraction.