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PubMed1988

Changes in activities of some enzymes of glycerolipid synthesis in brown adipose tissue of cold-acclimated rats.

1. Measurements were made of the activities of the following enzymes of glycerolipid synthesis in homogenates of interscapsular brown adipose tissue obtained from rats subjected to a 4 degrees C environment for time periods of 6 h up to 12 days: fatty acyl-CoA synthetase (FAS), mitochondrial and microsomal forms of glycerolphosphate acyltransferase (GPAT), monoacylglycerolphosphate acyltransferase (MGPAT) and Mg2+-dependent phosphatidate phosphohydrolase (PPH). 2. Relative to tissue DNA content, the activities of mitochondrial GPAT, MGPAT and Mg2+-dependent PPH were significantly increased after 1 day of exposure to cold, and continued to increase thereafter. By contrast, FAS and microsomal GPAT activities were unchanged relative to tissue DNA. 3. The time profile of the increase in MGPAT activity correlated well with a concomitant increase in the microsomal marker NADP+-cytochrome c reductase. Changes in mitochondrial GPAT and in Mg2+-dependent PPH activities were larger in amplitude than that of MGPAT. 4. It is proposed that these selective changes in enzyme activity may be associated with the onset of brown-adipose-tissue hyperplasia or possibly with an increase in triacylglycerol synthesis during cold-acclimation.

1-Acylglycerol-3-Phosphate O-Acyltransferase↗
PubMed2000

Rabies virus-induced membrane fusion pathway.

Fusion of rabies virus with membranes is triggered at low pH and is mediated by the viral glycoprotein (G). The rabies virus-induced fusion pathway was studied by investigating the effects of exogenous lipids having various dynamic molecular shapes on the fusion process. Inverted cone-shaped lysophosphatidylcholines (LPCs) blocked fusion at a stage subsequent to fusion peptide insertion into the target membrane. Consistent with the stalk-hypothesis, LPC with shorter alkyl chains inhibited fusion at lower membrane concentrations and this inhibition was compensated by the presence of oleic acid. However, under suboptimal fusion conditions, short chain LPCs, which were translocated in the inner leaflet of the membranes, considerably reduced the lag time preceding membrane merging, resulting in faster kinetics of fusion. This indicated that the rate limiting step for fusion is the formation of a fusion pore in a diaphragm of restricted hemifusion. The previously described cold-stabilized prefusion complex was also characterized. This intermediate is at a well-advanced stage of the fusion process when the hemifusion diaphragm is destabilized, but lipid mixing is still restricted, probably by a ring-like complex of glycoproteins. I provide evidence that this state has a dynamic character and that its lipid organization can reverse back to two lipid bilayers.

Liposomes↗
PubMed1990

Natural killer lines and clones with apparent antigen specificity.

Fresh CD3-, CD16+ lymphocytes that adhered to selected allogeneic lymphoblastoid cell lines (LCL) were cultured with LCL in the presence of IL-2-containing medium. The resulting lines as well as clones derived from these lines expressed CD16 and/or CD56, but lacked detectable CD3 or TCR-alpha/beta or TCR-gamma/delta complexes on the cell surface. Northern blot analysis failed to detect CD3 epsilon or TCR-beta transcripts, but revealed the presence of a TCR-gamma chain transcript in one of these lines. In addition to displaying potent cytolytic activity against K562 erythroleukemia cells (a classical NK target), the vast majority of these lines and clones lysed their specific stimulator LCL to a significantly greater extent than irrelevant LCL. This selective killing was inhibited by the addition of cold stimulator LCL or K562 cells, or anti-LFA 1 mAbs, but not by irrelevant LCL or mAbs to CD3, class I or class II MHC antigens. These results indicate that some CD3- lymphocytes, phenotypically indistinguishable from NK cells, can recognize and lyse allogeneic targets in a specific manner.

Antibodies, Monoclonal↗
PubMed2006

Long-chain polyunsaturated fatty acids and the pathophysiology of myalgic encephalomyelitis (chronic fatigue syndrome).

Evidence is put forward to suggest that myalgic encephalomyelitis, also known as chronic fatigue syndrome, may be associated with persistent viral infection. In turn, such infections are likely to impair the ability of the body to biosynthesise n-3 and n-6 long-chain polyunsaturated fatty acids by inhibiting the delta-6 desaturation of the precursor essential fatty acids--namely, alpha-linolenic acid and linoleic acid. This would, in turn, impair the proper functioning of cell membranes, including cell signalling, and have an adverse effect on the biosynthesis of eicosanoids from the long-chain polyunsaturated fatty acids dihomo-gamma-linolenic acid, arachidonic acid and eicosapentaenoic acid. These actions might offer an explanation for some of the symptoms and signs of myalgic encephalomyelitis. A potential therapeutic avenue could be offered by bypassing the inhibition of the enzyme delta-6-desaturase by treatment with virgin cold-pressed non-raffinated evening primrose oil, which would supply gamma-linolenic acid and lipophilic pentacyclic triterpenes, and with eicosapentaenoic acid. The gamma-linolenic acid can readily be converted into dihomo-gamma-linolenic acid and thence arachidonic acid, while triterpenes have important free radical scavenging, cyclo-oxygenase and neutrophil elastase inhibitory activities. Furthermore, both arachidonic acid and eicosapentaenoic acid are, at relatively low concentrations, directly virucidal.

Eicosapentaenoic Acid↗
PubMed1995

A cold-passaged, attenuated strain of human respiratory syncytial virus contains mutations in the F and L genes.

In previous studies, a mutant (cp-RSV) of the RSV A2 strain derived from 52 serial cold passages in bovine embryonic tissue culture was highly attenuated in seropositive adults and children but caused upper respiratory tract disease in seronegative infants. We investigated the genetic basis for this attenuation phenotype by comparing the complete genomic RNA sequence of this virus with the published sequence of strain A2 as well as with that of its unattenuated wild-type parent (HEK-7) virus. RNA was extracted from virions grown in tissue culture, reverse transcribed, amplified by the polymerase chain reaction (PCR), cloned, and sequenced. Changes from the published A2 wild-type sequence were confirmed on independently derived cDNA clones and by direct sequencing of PCR fragments. The HEK-7 parent virus was then analyzed at these positions by direct sequencing of PCR fragments. Fifteen nucleotide differences between the published A2 wild-type virus and cp-RSV were found. None appeared to involve cis-acting RNA sequences. Of the 15 nucleotide differences, only 1 occurred outside a translational open reading frame (ORF), and 2 which did occur within ORFs were silent at the amino acid level. The remaining 12 nucleotide differences encoded amino acid changes. All 3 of the mutations which were silent at the amino acid level, and 8 of the 12 which resulted in amino acid differences, were also present in the HEK-7 parent virus and therefore were not changes acquired during the cold passages. Thus, the remaining 4 nucleotide differences and the attendant 4 amino acid changes are associated with the attenuation phenotype of the cp-RSV. Two of the changes occur in the F gene and two in the L gene. These results confirm the previously described RSV genomic sequence, provide the first sequence of a live attenuated RSV vaccine strain, provide the first sequence of an RSV strain which has been evaluated in chimpanzees and humans, and indicate that attenuation in humans of a pneumovirus can be associated with a relatively small number of nucleotide and amino acid changes.

Amino Acids↗
PubMed2003

Chronic intermittent cold stress activates ovarian sympathetic nerves and modifies ovarian follicular development in the rat.

We studied the effects of a chronic intermittent cold stress regime on sympathetic nerve activation and ovarian physiology. This paradigm (4 degrees C for 3 h/day, Monday-Friday, for 3 or 4 wk) does not affect basal plasma levels of corticosterone. After 3 wk of stress, we detected a decrease in noradrenaline (NA) in the ovary, but after 4 wk, this ovarian neurotransmitter concentration increased over that of unstressed control rats. To analyze whether this effect on NA is preceded by an activation of the neurotrophic factor system responsible for growth and survival of sympathetic neurons, we measured both nerve growth factor (NGF) (by enzyme immunoassay) and the intraovarian levels of its low affinity receptor mRNA (by reverse transcription-polymerase chain reaction). The activation of sympathetic nerves was followed by an increase in NGF concentration without affecting the ovarian levels of either NGF or the mRNA of its receptor. Interestingly, follicular development changed during the stress procedure; after 3 or 4 wk of stress, we found a decrease in preantral healthy follicles without a compensatory increase in atresia. Concomitantly with the increase in NA and NGF in the ovary, we observed that a new population of follicles with hypertrophied thecal cell layers appeared after 4 wk of stress. These results suggest that chronic stress, through an intraovarian neurotrophin-mediated sympathetic activation, produces changes in follicular development that could lead to an impairment of reproductive function.

Androstenedione↗
PubMed1986

A human monoclonal IgM kappa cold agglutinin recognizing oligosaccharides with immunodominant sialyl groups preferentially at the blood group M-specific peptide backbone of glycophorins: anti-PrM.

A kappa-monotypic IgM high titer cold agglutinin reacting like anti-Pr at low, like anti-M at higher temperatures, is described. It recognizes tetra- and/or trisaccharides with immunodominant sialyl groups on glycophorins A, B, C like anti-Pr. Its affinity to the oligosaccharides is, however, approximately 10-fold increased when they are attached to the M-specific peptide backbone of glycophorin A. The antibody, termed anti-PrM, occurred in a blood group MN patient with chronic cold agglutinin disease and caused autoimmune hemolytic anemia.

Agglutinins↗
PubMed1991

Characterization of autoantibodies in mixed-type autoimmune hemolytic anemia.

Three of 46 patients with autoimmune hemolytic anemia (AIHA) satisfied the diagnostic criteria for mixed-type AIHA in which both warm-type and cold-type autoantibodies against red blood cells (RBCs) are present. The specificities of these autoantibodies were analyzed. All of the warm-type autoantibodies were IgG-chi, and the specificities were not serologically classified. The autoantibody of patient 1 reacted to the 41- and 80-kD peptides on immunoblotting, and the epitope corresponding to it was papain sensitive. Two warm-type autoantibodies from patients 2 and 3 resembled each other in serological analyses and reacted with a protease- and neuraminidase-resistant antigen. However, the antigen corresponding to the autoantibody of patient 3 was located on the 37-kD peptide by immunoprecipitation. All of the cold-type autoantibodies were IgM-cha and showed high titer and high thermal amplitude. According to the reaction pattern with untreated and enzyme-treated RBCs, the cold-type autoantibodies of patients 2 and 3 were revealed to be anti-Om and anti-I, respectively. In patient 1, the cold-type autoantibody was characterized as having high affinity for autologous RBCs, but its specificity was unclassified. The antigens corresponding to the cold-type autoantibodies were not located by immunoblotting and immunoprecipitation. These serological and immunochemical approaches to autoantibodies in mixed-type AIHA revealed that the warm and cold components recognized the different antigens.

Aged↗
PubMed1995

Hand-arm vibration in tropical rain forestry workers.

Working conditions and health hazards including vibration syndrome related to forestry work using chain-saws were studied in Papua New Guinea and Indonesia. The subjects comprised 291 workers including 97 chain-saw operators. The health examination consisted of peripheral circulatory and sensory tests in the upper extremities. The vibration spectrum measured at the handle of the chain-saw indicated that these acceleration levels would lead to a moderately high risk of hand-arm vibration syndrome (HAVS). The peripheral circulatory function tests revealed dysfunction after more than five years vibration exposure. However, in general, the results of the function tests and subjective complaints showed fewer health problems compared to those of Japanese forestry workers. The reason of such differences of vibration effects seem to be the following: (1) warmer climate (more than 25 degrees C throughout the year), (2) young workers and short work experience. (3) short time vibration exposures on working days in the natural forests, (4) seasonal changes in logging work (5) healthy workers effects. Thus, we found no clear evidence that the workers of our study suffered from HAVS. A principal component analysis was applied. The factor score of the components of the reactive dynamics of peripheral circulation differed significantly after more than five years' exposure. On the other hand, we cannot deny the possibility that subclinical dysfunction of peripheral circulation may be caused by chain-saw operation in the tropics in future. Further investigations on the HAVS among forestry workers in the tropic environment are needed.

Adult↗
PubMedDate not supplied

Heavy metal determination in aquatic species for food purposes.

New analytical procedures and sample mineralizations are proposed regarding the determination of arsenic, selenium, copper, lead, cadmium, zinc and mercury in matrices involved in food chain as mussel, clams and fishes. As regard As, Se, Cu, Pb, Cd and Zn determinations, H2SO4-HNO3 acidic mixture is used for the digestion of each matrix. In the case of Hg the sample digestion is performed using a concentrated suprapure H2SO4-K2Cr2O7 mixture and the results are compared with those from other conventional methods. Differential pulse cathodic (DPCSV) and anodic stripping voltammetry (DPASV) are employed for determining simultaneously selenium, arsenic and copper, lead, cadmium, zinc, respectively, while mercury determination is carried out by the cold vapour atomic absorption spectrometry (CV-AAS) with reduction with SnCl2. The voltammetric measurements were performed using a conventional three-electrode cell and the ammonia-ammonium chloride buffer (pH 9.3) as supporting electrolyte. For all the elements, in addition to the detection limits, precision and accuracy data are also reported: the former, expressed as relative standard deviation (Sr), and the latter, expressed as relative error (e), are in all cases between 3 to 6%.

Animals↗
PubMedDate not supplied

Acute hemolysis and renal failure with rifampicin-dependent antibodies after discontinuous administration.

Acute hemolysis as a reaction to rifampicin is extremely rare; case reports number less than 15. We recently evaluated a 65-year-old Cambodian refugee who self-regulated the use of rifampicin and isoniazid for pulmonary tuberculosis. Fifteen minutes after a single discontinuous oral dose, he developed flank pain, chills, rigors, vomiting, diarrhea, fever, and brown turbid urine. Laboratory tests at presentation showed acute intravascular hemolysis. Nonoliguric renal failure ensured, and he was transferred to our institution 2 days later. The patient was group A, Rh (D) positive, P1 negative with a cold autoantibody and cold anti-P1 alloantibody. The direct antiglobulin test was negative at the time of transfer. To evaluate the hemolysis, studies were done to test for rifampicin- or isoniazid-dependent antibodies. Rifampicin-dependent antibodies were detected in the antiglobulin phase with broad spectrum anti-human globulin, monospecific anti-gamma chain, and anti-complement antisera. Agglutination titers did not change after dithiothreitol reduction of the patient's serum. We conclude that this patient developed rifampicin-dependent IgG antibodies with complement-fixing capability. The presence of rifampicin-dependent antibodies should be suspected in a patient with hemolysis and/or renal failure taking rifampicin.

Acute Kidney Injury↗
PubMed1992

The hemoglobins of Notothenia angustata, a temperate fish belonging to a family largely endemic to the Antarctic Ocean.

The blood of the teleost Notothenia angustata contains a major hemoglobin (Hb 1, over 95% of the total), accompanied by a minor component (Hb 2). The two hemoglobins have identical beta chains and differ in their alpha chains. The primary structure of both hemoglobins has been established through the elucidation of the complete amino acid sequence of the three chains. The study of the oxygen-binding properties shows that Hb 1 displays the Bohr and Root effects and has high affinity for organic phosphates. N. angustata belongs to the family Nototheniidae, suborder Notothenioidei. Unlike the vast majority of nototheniid species, which live in isolation in the Antarctic Ocean and have developed cold adaptation, N. angustata inhabits the waters of southern New Zealand and is not cold adapted. Although some hematological parameters typically favour oxygen transport in a temperate environment, the hemoglobin multiplicity and structural and functional features closely resemble those of the Antarctic species of the same family and suborder. Thus, N. angustata may be considered as a link between temperate and Antarctic habitats. The hypothetical separation history of N. angustata from the Antarctic species of the same family is discussed in the light of the present findings.

Acclimatization↗
PubMed1990

A promising approach for improving the recovery of heart transplants. Prevention of free radical injury through iron chelation by deferoxamine.

Iron catalysis is involved in the generation of the highly cytotoxic hydroxyl radical and in the chain reactions of subsequent lipid peroxidation that lead to irreversible membrane damage. Assuming that ischemically stored heart transplants may incur free radical injury at the time of reoxygenation, we assessed the effects of the iron chelator deferoxamine in 70 isolated isovolumic buffer-perfused rat hearts subjected to the following protocol: cardioplegic arrest; cold (2 degrees C) storage for 5 hours; global ischemia at 15 degrees C for 1 hour, intended to simulate the implantation procedure; and normothermic reperfusion for 1 additional hour. During poststorage ischemic arrest, the following techniques of myocardial protection were evaluated: hypothermia alone; high-pressure (60 cm H2O) cardioplegia given at 0, 30, and 55 minutes of arrest; low-pressure (30 cm H2O) cardioplegia given at 0 and 55 minutes of arrest; and low-pressure (30 cm H2O) cardioplegia only given at 55 minutes of arrest. Treated hearts had deferoxamine (6 mumol) added to the cardioplegic solution used throughout the experimental time course. Further, in the treated group subjected to the protocol of single cardioplegic delivery at end ischemia, deferoxamine was given both in the cardioplegic reperfusate and in the Krebs buffer over the 15 initial minutes of reflow. Based on comparisons of postreperfusion ventricular pressure development, maximal rate of rise of ventricular pressure, left ventricular compliance, and coronary flow, the best myocardial protection was afforded by deferoxamine given as an additive to single-dose cardioplegic solution at the end of arrest and to the reperfusate during the initial phase of reoxygenation. As the drug has no inotropic effect, its protective action is most likely related to a decrease in catalytic iron available for free radical production and lipid peroxidation. These results support the hypothesis that oxidative damage may contribute to donor heart failure and demonstrate that this form of damage can be efficiently acted upon by iron chelation. The clinical relevance of these data stems from the fact that deferoxamine is available for human use and might become an effective means of improving donor heart preservation in the setting of clinical heart transplantation.

Animals↗
PubMed1995

The vibrotactile threshold (VT) at the fingertips of chain saw operators.

The vibration sensitivity of the skin at the ten fingertips was tested with 72 chain saw operators examined due to VWF-disease and compared with controls. The vibrotactile perception threshold (frequency f = 125 Hz, load p = 1.3 N) proved to be 3 to 5 times as high for the exposed group compared to controls. There was no correlation between vibration sensitivity and peripheral circulation found in cold provocation test. Two different testing instruments (pallaesthesiometer = vibrotactometer) showed significant differences in results probably due to different technical details of the methods. It may be concluded that a standardization of the instrumentation and method is necessary for testing the vibrotactile perception threshold as a part of the nervous function in a comparable manner.

Adult↗
PubMed2006

Salicylic acid, ethephon, and methyl jasmonate enhance ester regeneration in 1-MCP-treated apple fruit after long-term cold storage.

Volatile esters, primarily synthesized in peel tissues, are major aromatic components of apple fruits [Malus sylvestris (L.) Mill. var. domestica (Borkh.) Mansf.]. The use of cold storage combined with 1-methylcyclopropene (1-MCP) treatment prolongs the life of apples but represses the regeneration of esters during poststorage ripening. In this study, the regeneration of total esters was significantly increased in apple fruits treated with salicylic acid (SA) and Ethephon (ETH) that had been treated once or twice with 1-MCP. However, methyl jasmonate (MeJA) treatment resulted in regeneration of total esters after a single 1-MCP treatment. To determine the mechanism by which SA, ETH, and MeJA regulate ester regeneration, the apple alcohol acyltransferase gene (MdAAT2) was investigated at the mRNA, protein, and enzyme activity levels. Genes associated with ethylene perception were also investigated by RT-PCR. The results suggest that MdAAT2 controls ester regeneration and that MdETR1 plays a key role in ethylene perception and regulation of downstream MdAAT2 gene expression during poststorage. Ester compounds and concentrations differed in peels treated with different signal molecules, indicating that regulation of the pathway upstream of straight-chain ester biosynthesis depended on the regulation of lipoxygenase (LOX) and alcohol dehydrogenase (ADH) activity by SA, ETH, and MeJA during poststorage ripening.

Acetates↗
PubMed1996

32P-incorporation PCR for the detection of rearrangements at the TCR-gamma locus.

We have adapted and developed a PCR (polymerase chain reaction)-based technique for the T-cell receptor (TCR)-gamma chain gene, which has subsequently been used for routine diagnosis. Variable-region oligonucleotide primers were chosen from subgroups I and II, and the joining region primer was from the J2 segment. The primers were used to perform a 32P-incorporation PCR, and the products were then separated on an 8% denaturing polyacrylamide gel. In our hands, this technique is more reliable than cold methods, when separation is performed on either agarose or nondenaturing polyacrylamide. The radioactive technique was used to look at 102 T-cell proliferations, of which eight of eight T-acute lymphoblastic leukemia (ALL), 24 of 34 T-non-Hodgkin's leukemia (NHL), and 35 of 60 large granular lymphocyte (LGL) expansions were clonal. Of 122 B-cell proliferations investigated, including 72 cases of B-cell lineage ALL, 36 demonstrated a T-cell rearrangement (33 ALLs and three myelomas). Samples from nonlymphoid tumors were tested and produced a normal distribution ladder of PCR products after autoradiography, a pattern also observed with antenatal and preoperative patients. The radiolabel-incorporation method detected an abnormal pattern of a ladder with prominent dark bands in 29 of 122 B-cell and 27 of 102 T-cell cases and in 0 of 49 of the nonlymphoid and normal samples. The abnormal banding patterns obtained in a proportion of the B- and T-cell cases was not readily discernible by nondenaturing-acrylamide or agarose-separation methods.

Blotting, Southern↗
PubMed1994

Hypocomplementemia associated with hepatitis C viremia in sera from voluntary blood donors.

OBJECTIVES: Hepatitis C virus (HCV) infection induces extra-hepatic manifestations, most of which are considered to be mediated by circulating immune complexes. For evaluating this association in a wider perspective, complement activity was determined in sera from apparently healthy individuals, and hypocomplementemia was tested for correlation with HCV viremia. METHODS: Sera from 10,532 voluntary blood donors were stored at 4 degrees C overnight, serially diluted 2-fold, and tested for hemolytic activity by a microtitration method and antibody to HCV (anti-HCV) by passive hemagglutination with recombinant HCV antigens of the second generation. HCV RNA was determined in sera with anti-HCV or hypocomplementemia, or both, by polymerase chain reaction with nested primers deduced from the 5'-noncoding region of the HCV genome. RESULTS: Hypocomplementemia was detected in 53 (0.5%) of 10,532 donations and anti-HCV in 94 (0.9%). Anti-HCV was detected in 48 (91%) of the 53 sera with hypocomplementemia, more frequently than in 46 (0.44%) of 10,479 sera without (p < 0.001). Among 94 sera positive for anti-HCV, HCV RNA was detected in 45 (94%) of 48 sera with hypocomplementemia, more often than in 10 (22%) of 46 sera without (p < 0.001). CONCLUSIONS: A close association of hypocomplementemia with HCV viremia among apparently healthy blood donors would reflect circulating immune complexes which may cause extrahepatic diseases, such as cryoglobulinemia and membranoproliferative glomerulonephritis, in some HCV carriers. The storage of sera from HCV carriers at 4 degrees C before the test would have contributed to a decreased hemolytic activity due to the cold activation of complement by cryoglobulins involving HCV.

Adult↗