Endothelial dysfunction induced by triglycerides is not restored by exenatide in rat conduit arteries ex vivo

Exenatide (synthetic exendin-4) is a stable analogue of glucagon-like peptide 1 (GLP-1) and has recently been approved for clinical use against type 2 diabetes. Exenatide is believed to exert its effects via the GLP-1 receptor with almost the same potency as GLP-1 in terms of lowering blood glucose. Short term exenatide treatment normalizes the altered vascular tone in type 2 diabetic rats, probably due to the reduction in glycemia. The aim of this study was to investigate whether exenatide directly protects against triglyceride-induced endothelial dysfunction in rat femoral arterial rings ex vivo. Short term pre-incubation with Intralipid^® (0.5 and 2%) was found to dose-dependently induce endothelial dysfunction, in that it elicited a significant reduction in ACh-induced vasorelaxation by 29% and 35%, respectively. Paradoxically, this occurred with a concomitant increase in endothelial nitric oxide synthase (eNOS) activity. No such reduction in vasorelaxation by Intralipid^® was seen in response to the NO donor sodium nitroprusside (SNP), revealing an endothelium-dependent vascular dysfunction by Intralipid^®. However, exenatide did not protect against Intralipid^®-induced endothelial dysfunction. More surprisingly, the maximum vasorelaxation induced by exenatide (without Intralipid^®) was only 3 ± 2%, compared to the 23 ± 4%, 38 ± 4%, 79 ± 3% and 97 ± 4% relaxations induced by GLP-1, GLP-1 (9-36), ACh and SNP, respectively. This unexpected finding prompted us to ascertain that the exenatide preparation was biologically active, and both exenatide (10^− 11 mol/l) and GLP-1 (10^− 9 mol/l) significantly increased insulin secretion in pancreatic β-cells from ob/ob mice in vitro. In conclusion, exenatide could neither confer any acute protective effects against triglyceride-induced endothelial dysfunction nor exert any significant vasorelaxant actions in this model of rat conduit arteries ex vivo.     

Application of soil slurry respirometry to optimise and subsequently monitor ex situ bioremediation of hydrocarbon-contaminated soils

A protocol to monitor respiration as O₂ consumption in soil slurries using the Strathtox^® respirometer was developed and tested on four soils from brownfield sites. Respiration rates (mg l⁻¹ h⁻¹) of soil slurries in the Strathtox^® were compared with rates (μl min⁻¹) of field moist soils analysed using the Columbus Oxymax^® ER10 respirometer. One of the soils (99612B), historically contaminated with diesel, was further studied by monitoring the effect of inorganic NH₄NO₃ liquid nutrient on enhancing respiration rate. Soil microcosms were monitored continuously on the Oxymax^® or sampled at 24, 48 and 72 h intervals, prepared as soil slurries, and analysed on the Strathtox^®. On the full-scale remediation project (6000 m³) soil 99612B was treated as a biopile with spent mushroom compost (SMC) amendment and respiration rates monitored in samples over an 8-week period. In the laboratory microcosm experiment and full-scale bioremediation treatment described, correlation was found for respiration rates between the two respirometry systems.     

Effects of Fermenten supplementation to beef cattle

Two experiments were conducted to evaluate a commercially available supplemental N source for beef cattle (Fermenten^®; Church & Dwight Co., Inc., Princeton, NJ, USA). The first experiment evaluated kinetics of in vitro NH₃-N release using batch cultures of rumen fluid incubated with: control (no N added), soybean meal, urea, and Fermenten^®. Ammonia-N was measured at 0, 0.5, 2, 4, 6, 8, 12 and 24 h after incubation began. A treatment by time interaction (P<0.01) occurred in which, during the initial 2 h, Fermenten^® cultures had the highest (P<0.01) NH₃-N but, from 4 to 24 h, the highest (P<0.01) NH₃-N concentrations were with urea-incubated cultures. The total increase in NH₃-N concentrations from 0 to 24 h of incubation was less for Fermenten^® (P<0.01) than for the soybean meal and urea. The second experiment assessed effects of Fermenten^® supplementation on growth, blood parameters, voluntary forage intake and reproductive performance of beef heifers. Sixty heifers, stratified by initial body weight (BW), were randomly allocated to one of two treatments that consisted of iso-nitrogenous grain-based supplements containing either Fermenten^® (72 g/kg, as-fed) or urea (9.7 g/kg, as-fed). Supplements were offered three times weekly at a rate of 2.4 kg of dry matter per heifer daily. Shrunk BW was measured on days 0 and 112 for calculation of daily body weight gain. Body volume measurements were completed on days 0, 28, 56, 84 and 112, whereas pelvic area was assessed on days 0, 56 and 112. Blood samples were collected on days 28, 56, 84 and 112 for analysis of metabolites and hormones. On day 56, 2 heifers, which were randomly selected from each pasture, were placed in individual feeding stations for 26 days to determine treatment effects on voluntary forage intake. On day 112, all heifers were grouped by treatment and exposed to bulls for 60 days. Fewer heifers offered the Fermenten^® supplement attained puberty (P<0.05) and became pregnant during the study compared to heifers fed urea (0.60 and 0.93, respectively; P<0.01). Addition of Fermenten^® to batch cultures of rumen fluid rapidly increased NH₃-N concentrations, whereas further increases occurred in a slower and steady rate. Beef heifers fed a supplement containing Fermenten^® had similar growth and development, but inferior reproductive performance, than heifers fed a supplement containing urea.     

Occurrence of Stereoisomers of 1-(2′-Pyrrolidinethione-3′-yl)-1,2,3,4- tetrahydro-β-carboline-3-carboxylic Acid in Fermented Radish Roots and Their Different Mutagenic Properties

Stereoisomers of the tetrahydro-β-carboline derivative, 1-(2′-pyrrolidinethione-3′-yl)-1,2,3,4-tetrahydro-β-carboline-3-carboxylic acid (PTCC), were formed from L-tryptophan with 4-methylthio-3-butenyl isothiocyanate, and their mutagenic properties and contents in different types of the radish products were studied. The isomers were identified as (1S ^*, 3S ^*, 3′R ^*)- and (1R ^*, 3S ^*, 3′R ^*)-PTCCs; the former was found as the major compound but had no mutagenic activity, while the latter was mutagenic toward Salmonella typhimurium TA 98 in the presence of a rat microsomal fraction. Both (1S ^*, 3S ^*, 3′R ^*)- and (1R ^*, 3S ^*, 3′R ^*)-PTCC were detected in a ratio of about 4:1 in a product fermented for 8 months, but only a trace was apparent in products manufactured within a few weeks.     

Inhibition of plant and microbial ureases by phosphoroamides

Enzyme kinetic studies of inhibition of plant (jackbean) and microbial (Bacillus pasteurii) ureases by eight phosphoroamides [phenylphosphorodiamidate, 4-chlorophenylphosphorodiamidate, phosphoric triamide, N-(diaminophosphinyl)benzamide, N-(diaminophosphinyl)benzeneacetamide, 4-chloro-N-(diaminophosphinyl)benzamide, N-(4-nitrophenyl)phosphoric triamide, N-(diaminophosphinyl)-3-pyridinecarboxamide] demonstrated that these compounds are slow, tight-binding inhibitors of urease enzymes. Measurement of the dissociation constants (Ki^*) of the enzyme-inhibitor complexes (E · I^*) formed by interaction of the ureases and phosphoroamide inhibitors studied showed that these inhibitors had a much higher affinity (i.e., a lower Ki^*) for plant urease than for microbial urease. Measurement of rate constants for formation (k_on) and decay (k_off) of E · I^* showed that, whereas k_on varied greatly with the different inhibitors and ureases, k_off was constant for the phosphoroamides tested and had a characteristic value for each urease. The half-life of E · I^* (30°C; pH 7 THAM buffer) for the plant urease was much longer than that for the microbial urease, and this difference largely accounted for the much higher values of Ki^* (k_off/k_on) observed with microbial urease.     

Selective oxidation of carbohydrates by 4-AcNH-TEMPO/peracid systems

Starch, amylopectin, inulin, pullulan and methyl α- -glucopyranoside (Me α-Glcp) were oxidised by 4-acetamido-2,2,6,6-tetramethylpiperidine-1-oxyl (4-AcNH-TEMPO) as the mediator and peracetic acid or monoperoxysulfate (Oxone^®) as the regenerating oxidant. The conversion of primary alcohol groups to the corresponding carboxyl groups proceeded with high yield and selectivity, provided that sodium bromide was added as co-catalyst.The mass molecular distributions of the oxidised polysaccharides indicated that no major depolymerisation occurred during oxidation. Oxone appeared to be the most efficient oxidant as the reaction rate was 25 times higher than that of peracetic acid in the oxidation of Me α-Glcp. On the other hand, oxone produces a larger amount of waste as by-product than peracetic acid.     

Silymarin BIO-C, an extract from Silybum marianum fruits, induces hyperprolactinemia in intact female rats

Breastfeeding is widely acknowledged to have important health benefits for infants and mothers. Milk thistle (Silybum marianum fruits) has been recently proposed to be used by nursing mothers for stimulating milk production; however, the mode of action of this herbal drug is still unknown. In this paper, we have evaluated the effect of a micronized standardized extract of S. marianum (Silymarin BIO-C^®=Piùlatte^®) on the serum levels of prolactin in female rats. A 14-day treatment with Silymarin BIO-C^® (25–200 mg/kg, given orally) increased, in a dose dependent manner, the serum prolactin levels. Moreover, after a 66-day discontinuation of Silymarin BIO-C^® treatment, prolactin levels were still significantly elevated although we observed a trend to decrease that was counteracted by a further 7-day treatment with Silymarin BIO-C^®. Bromocriptine, a dopamine D₂ receptor agonist, (1–10 mg/kg, os) significantly and in a dose dependent manner, reduced the serum prolactin levels; bromocriptine, at the dose of 1 mg/kg, significantly reduced the high serum prolactin levels induced by Silymarin BIO-C^®. In conclusion, we have shown that an extract from S. marianum fruits significantly increases circulating prolactin levels in female rats; this effect seems to involve, at least in part, dopamine D₂ receptors.     

The prostacyclin analogue beraprost sodium prevents development of arterial stiffness in elderly patients with cerebral infarction

Prostacyclin (PGI(2)) inhibits platelet aggregation, smooth muscle cell proliferation, and vasoconstriction. Arterial stiffness assessed by pulse wave velocity (PWV) predicts mortality in various cardiovascular diseases. To study the preventive effects of a prostacyclin analogue, beraprost sodium, on arterial PWV values in elderly patients with cerebral infarction. Forty-four patients with a history of cerebral infarction received beraprost sodium (120 microg/day p.o.) or no beraprost sodium (control) for 3 months. Arterial PWV and ankle brachial indices (ABI) were determined prior to starting the medication and after 3 months of medication. Initially, there were no differences in age, blood pressure, and body mass index. Further, PWV or ABI did not differ between the beraprost sodium group (n = 22) and the control group (n = 22). After 3 months, PWV in beraprost sodium group was significantly reduced (-123 +/- 282) when compared with the control group (147 +/- 274)(P = 0.006). ABI was not significantly different when comparing the two groups at 3 months. Long-term administration of beraprost sodium prevents the decline in arterial biomechanics in elderly patients with cerebral infarction.     

The methodological approach for the generation of humandendritic cells from monocytes affects the maturation state of the resultant dendritic cells

Dendritic cells (DCs) are effective as antigen-presenting cells in the immune system and are present at two functional stages depending on their maturation state. For experimental investigation of this concept, CD14⁺ monocytes from blood are isolated and cultured to generate in vitro the DCs needed for functional analysis. For positive selection of CD14⁺ monocytes we compared two immunomagnetic bead technologies: MACS^® Separation, created by Miltenyi Biotec, and EasySep^® Selection, created by StemCell Technologies. The monocytes provided dendritic cells for their functional analysis. Lipopolysaccharide was added to cultured DCs to induce maturation. Although both systems generated DCs from the positively selected CD14⁺ cells, there were certain differences between them. Morphological, phenotypic, and functional analysis showed that MACS^®-selection provided DCs that have typical features corresponding to day 6 or 7 of maturation. EasySep^®–DCs exist in a partially-mature state from day 6 onward, even without the addition of a maturation stimulus. The reason behind this partial maturation is possibly based on the dextran-coated beads that are associated with the EasySep^® product. Both methods provide pure and viable DCs, but we would recommend using the MACS^® system for obtaining DCs suitable for functional studies.     

Acid dissociation constants and pH values for standard “bes” and “tricine” buffer solutions in 30, 40, and 50 mass% dimethyl sulfoxide/water between 25 and −25 °C

Information is given concerning two standard buffer solutions suitable as pH references in 30, 40, and 50 mass% dimethyl sulfoxide (DMSO)/H₂O mixed solvents at subzero temperatures from −20 to 0 °C, with the intention of establishing a pH (designated pH^*) scale. The two buffers selected were the ampholytes N,N-bis(2-hydroxyethyl)-2-aminoethane sulfonic acid (“bes”) and N-tris(hydroxymethyl)methylglycine (“tricine”), and the reference standard consisted of equal molal quantities of the buffer and its respective sodium salt. The assignment of pH^* values was based on measurements of the emf of cells without liquid junction of the type: Pt;H₂(g,1 atm) ¦Bes, Na Besate, NaCl ¦ AgCl;Ag and Pt;H₂(g,1 atm) ¦Tricine, Na Tricinate, NaCl ¦AgCl;Ag and the pH^* was derived from a determination of K₂, the equilibrium constant for the dissociation process (Buffer)^±/ai (Buffer)⁻ + H⁺.     

High-performance liquid chromatographic determination of paclitaxel in rat serum: application to a toxicokinetic study

A simple and sensitive high-performance liquid chromatographic method is described for the determination of paclitaxel (Taxol^®) at 230 nm using a Nucleosil C₁₈ (5 μm) column and a methanol–water (70:30, v/v) mobile phase following a single-step extraction from serum with dichloromethane. The assay was validated against the classical criteria and was applied to a toxicokinetic study in rats after one or five, one per week) intraperitoneal administrations of 16 mg/kg Taxol^®.     

Determination of the enantiomeric composition of (R*,S*)-1-ethylpropyl 2-methyl-3-hydroxypentanoate, the male-produced aggregation pheromone of Sitophilus granarius

The enantiomeric composition of sitophilate, the granary weevil [Sitophilus granarius (L.)] male-produced aggregation pheromone [(R^*,S^*)-1-ethylpropyl 2-methyl-3-hydroxypentanoate)], was determined by three methods: (1) bioassaying the synthetic (2S,3R) and (2R,3S) enantiomers of the active (R^*,S^*) diastereomer; (2) ¹H NMR spectroscopy of Mosher ester derivatives of the natural pheromone and synthetic (2S,3R)-and (2R,3S)-sitophilate; and (3) capillary GLC comparisons of the retention times of derivatized natural pheromone and the two synthetic enantiomers. The combined methods confirmed the (2S,3R) enantiomer as the active form of sitophilate. Male granary weevils were shown to produce >96% (2S,3R)-sitophilate. No significant attraction of S. granarius by the (2R,3S) enantiomer was observed. Rice and maize weevils [S. oryzae (L.) and S. zeamais Motschulsky] were not attracted by (2S,3R)-sitophilate. S. granarius L. est un déprédateur important des grains stockés. Le (R^*,S^*)-1-éthylpropyl 2-méthyl-3-hydroxypentanoate a été identifié en 1987 comme le principal composé du sitophilate, la phéromone mâle d'agrégation de S. granarius. La composition énantiométrique du sitophilate a été déterminée par 3 méthodes:

In vitro biohydrogenation of four dietary fats

This study was designed to determine in vitro rates of biohydrogenation of dietary unsaturated fatty acids by a mixed population of rumen microbes. The four dietary fats [Alifet High-Energy^® (AHE), Alifet-Repro^® (AR), Megalac^® (MG), and Energy Booster^® (EB)] differ in method of preparation, fatty acid composition, or both of these factors. Dietary fats (20 mg) were incubated with 4 mL strained rumen fluid diluted with 16 mL of medium, 0.8 mL of reducing solution buffer, and 200 mg of a synthetic diet (370 g cellulose, 370 g starch, and 160 g casein per kg DM) at 37 °C. Total contents were collected after 0, 6, 12, 24, or 36 h and change in fatty acid content determined. Disappearance of oleic acid was minimal (0.05–0.20) in AR and MG but moderate (about 0.60) in AHE and EB after 36 h of incubation. Rate of biohydrogenation of linoleic and linolenic acids from AR were similar (0.025 ± 0.009 h⁻¹) and 0.65 of these fatty acids remained intact after 36 h. Rate of biohydrogenation of linoleic acid was four times greater than for oleic acid (0.040 ± 0.013 h⁻¹ versus 0.009 ± 0.002 h⁻¹) in MG. Thus, 0.65 of the linoleic acid but only 0.20 of the oleic acid had disappeared from MG after 36 h. Trans-11 and trans-12 were the predominant trans-isomers in AHE and AR cultures whereas trans-9 and trans-10 were the predominant trans-isomers in EB and MG cultures. None of the dietary fats contained conjugated linoleic acid (CLA) but CLA was present in the incubation inoculum. The amount of CLA decreased with time but this was not affected by source of dietary fat. Most (0.90–0.95) of the long-chain fatty acids eicosapentaenoic (EPA) and docosahexaenoic (DHA) in AR remained after 36 h of incubation. Results demonstrate that biohydrogenation varied among fatty acids and among source of dietary fat and indicate that AR can be used to increase post-ruminal supply of linolenic, EPA and DHA.     

Effects of endometrium on metabolismof arachidonic acid by bovine blastocysts

Metabolism of radiolabeled arachidonic acid (^*AA) by blastocysts and endometrial slices recovered from five gilts 16 days after detection of estrus was studies . Blastocysts from each gilt were divided into four 216 ± 18 mg, and each portion was placed into a separate petri dish containing 15 ml modified minimum essential medium (MEM)_. The incubates from each gilt received either 25, 50, 100 or 200 μg radioinert arachidonic acid (AA). Endometrium was dissected from each uterin horn, sliced and duplicate 509 ± 3 mg portions from each gilt were placed into petri dishes containing 15 ml MEM and 200 μm AA. All incubates received 5 νCi of ^*AA (either [¹⁴C]-arichidonic acid or [³H]-arichidonic acid). The incubates were rocked at 37°C for 24 h in an atmosphere of 50% n₂:45% O₂:5% CO₂. After incubation, tissues and MEM were separated by centrifugation. Metabolism of ^*AA was assessed in extracts of MEM and tissue homogenates by separating ^*AA and its metabolites on columns of Sephades LH-20. Blastocysts produced compounds that migrated with [³H]-13,14-dihydro-15-keto-PGF_2^α (^*PGFM), [³H]-PGE₂ (^*PGE₂) and [³H]-PGF_2^α (^*PGF_2^α). The greatest (P<.05) proportion (35.7 ± 1.8%) of the radioactivity in blastocyst MEM was recovered as PGE₂. In blastocyst homogenates, most (66.2 ± 3.3%; P<0.05) of the radioactivity was in a nonporal peak assumed to be arachidonate esters. The concentration of AA ni MEM did not alter metabolism of ^*AA by blastocysts. Endometrial slices produced ^*PGFM and ^*PGE₂ but only in small amounts, and they were capable of producing nonpolar, probably esterified, forms of ^*AA. It was concluded that porcine blastocysts produced and metabolized prostaglandins and that they make a contribution to the uterine milieu during early pregnancy.     

Biomonitoring of urine mutagenicity with the Ames test: improvement of the extraction/concentration method

Comparative extraction efficiency of the pre-packed Bakerbond^®-spe-SDB-1 resin and of Amberlite^®-AD2 (XAD-2) resin, for the preparation of urine extracts in biomonitoring studies. Urine extracts were prepared in parallel with the Bakerbond^® column and with the classical XAD-2 resin from urines (1) spiked with mutagenic chemicals, (2) collected from patients after chemotherapy, and (3) from smokers. Mutagenic activities were evaluated on Salmonella typhimurium tester strains TA97a, TA98, TA100 and TA102 with and without S9 mix. Mutagenic activities obtained with Bakerbond^® extracts were almost always higher or at least equivalent to those prepared on XAD-2 resin. Similar results were observed for the three urine sample groups. When fully validated, the use of the pre-packed columns will be more convenient and time-saving for large population studies.     

Genetic evidence of closed life-cycles for some coral reef fishes within Taiaro Lagoon (Tuamotu Archipelago, French Polynesia)

 Taiaro Lagoon has no permanent or regular connection with the ocean (ingress is restricted to episodes of high sea-level and/or strong wave action) raising the question of how fish populations with normally dispersive larvae are maintained inside this lagoon. We compared the genetic population structures of two coral reef fishes, Acanthurus triostegus and Chaetodon ulietensis, present on both sides of the atoll rim to determine whether there is evidence of reproductive isolation. Genetic surveys showed that the lagoonal and oceanic populations were statistically different at five loci (AAT ^* -3, GDA ^*, HPD ^*, MDH ^* and SDH ^*) in A. triostegus and three loci (PGI-2^*, IDH ^* and PGD ^*) in C. ulietensis, producing high F _st values of 0.055 and 0.021, respectively. Our genetic and demographic data on these species suggest that both may be completing their life-cycles inside the lagoon, which leads us to question the common assumption that coral reef fishes require oceanic conditions for larval development. Accepted: 28 August 1997     

Progressively developed myocardial apoptotic cell death during late phase of reperfusion

Myocardial apoptosis is primarily triggered during reperfusion (R). The aim of this study was to test the hypothesis that R-induced apoptosis develops progressively during the late phase of R, and that R-induced apoptosis is associated with changes in expression of anti- and pro-apoptotic proteins and infiltrated inflammatory cells. Thirty-one dogs were subjected to 60 min of left anterior descending coronary occlusion followed by 6, 24, 48, and 72 h R, respectively. There was no group difference in collateral blood flow, measured by colored microspheres during ischemia. Necrotic cell death (TTC staining) was significantly increased during R, starting at 27 ± 2% at 6 h R and increasing to 41 ± 2% at 24 h R. There was no further change at 48 (37 ± 3%) and 72 (36 ± 6%) h R, respectively. TUNEL positive cells (% total normal nuclei) in the peri-necrotic zone progressively increased from 6 (26 ± 2^*) to 24 (38 ± 1^*), 48 (48 ± 3^*) and 72 (59 ± 4^*) h R, respectively. The number of detected TUNEL positive cells at these time points was consistent with an increased intensity of DNA ladders, identified by agarose gel electrophoresis. Compared with normal tissue, western blot analysis showed persistent reduction in expression of anti-apoptotic protein Bcl-2 from 6 (16 ± 0.8%^*) to 72 h R (78 ± 2%^*), and increase in expression of pro-apoptotic proteins including Bax from 6 (30 ± 3%^*) to 72 h R (66 ± 3%^*), and p53 from 6 (12 ± 1%^*) to 72 h R (91 ± 2%^*), respectively. Immunohistochemical staining revealed that infiltrated neutrophils (mm² myocardium) were significantly correlated with development of necrotic and apoptotic cell death from 6 to 24 h R, respectively (P < 0.05), while large macrophage infiltration seen during 48 to 72 h R were correlated with apoptotic cell death (P < 0.05). These results indicate that 1) necrosis peaked at 24 h R when apoptosis was still progressively developing during later R; 2) changes in Bcl-2 family and p53 proteins may participate in R-induced myocardial apoptosis; 3) inflammatory cells may play a role in triggering cell death during R. ^* P < 0.05 vs. normal nuclei and tissue; P < 0.01 vs. 6 h R.     

Synthesis of (−)-Methyl Shikimate via Enzymatic Resolution of (1S*, 4R*, 5R*)-4-Hydroxy-6-oxabicyclo[3.2.1]oct-2-en-7-one

The synthesis of methyl (?)-shikimate [(?)-2] was achieved via lipase-catalyzed optical resolution of (1S^*,4R^*,5R^*)-4-hydroxy-6-oxabicyclo[3.2.1]oct-2-en-7-one (3). Transesterification of (±)-3 and vinyl acetate with lipase MY and subsequent hydrolysis gave optically pure (?)-3. This compound was converted to (?)-2 in two steps.     

Molecular characterization of HLA-A28*, a novel HLA product,and its relationship to HLA-A28 and HLA-A2

The HLA-A28^* molecule expressed by the B-cell line IDF is serologically distinct and intermediate between HLA-A28 and HLA-A2. Comparative tryptic peptide mapping of biosynthetically labeled HLA-A28^*, A28, and A2 molecules showed that HLA-A28^* is also chemically distinct. Reverse-phase high pressure liquid chromatographic analysis of tryptic peptides labeled with ³H-arginine and ³H-lysine revealed that A28^*. A28, and A2 share 65% of their tryptic peptides. Multiple differences were observed between A28^* and both A28 and A2. No peptides unique to A28^* were detected and 25 peptides were shared with both A28 and A2. These results show that A28^* is a novel HLA product that is closely related to A28 and A2. Tryptic peptide map comparisons of these molecules labeled separately with 11 amino acids confirm these results. The data suggest that HLA-A28 ^* may have arisen from a genetic exchange event involving HLA-A28 and -A2. These data are consistent with the hypothesis that A28^* is identical with A28 in the first extracellular domain ( ₁) and identical with A2 in the second domain ( ₂).Abbreviations used in this paper EDTA ethylenediaminetetraacetic acid - HPLC high-pressure liquid chromatography - MHC major histocompatibility complex - NP40 Nonidet P40 - PMSF phenylmethylsulphonylfluoride - SDS-PAGE sodium dodecyl sulfate-polyacrylamide gel electrophoresis - TPCK L(tosylamido-2-phenyl) ethyl chloromethyl ketone - Tris tris (hydroxymethyl)-aminomethane - A alanine - C cysteine - D aspartic acid - E glutamic acid - G glycine - H histidine - K lysine - L leucine - M methionine - N asparagine - Q glutamine - R arginine - S serine - T threonine - V valine - W tryptophan - Y tyrosine