The alpha chemokine, interleukin 8, inhibits the antiviral action of interferon alpha.

Interferon (IFN) exhibits a potent antiviral activity in vitro and plays a major role in the early defense against viruses. Like IFN, the proinflammatory chemokine, interleukin (IL)-8, is induced by viruses and appears in circulation during viral infections. In an in vitro cytopathic effect assay for IFN, we found that IL-8 can inhibit IFN-alpha activity in a dose-dependent manner. This action was reversed by specific monoclonal antibodies to IL-8. The chemokine was able to attenuate the IFN-mediated inhibition of viral replication as determined by measuring infectious virus yield. IL-8 also diminished the ability of IFN to inhibit an early stage of viral replication since IL-8 attenuated the inhibition of the formation of viral proteins. It appeared that IL-8 interfered with a late rather than an early step of IFN-mediated pathway such as early gene expression. The IL-8 inhibitory action on IFN-alpha antiviral activity was associated with reduced 2',5'-A oligoadenylate synthetase activity, a pathway well correlative with the anti- encephalomyocarditis virus action of IFN-alpha. Understanding pathways that antagonize IFN action may lead to novel approaches to potentiate endogenous and therapeutic IFN.

Monomeric human IgE evokes a transient calcium rise in individual human neutrophils.

Digital fluorescence calcium imaging was used to investigate and identify the primary biological responses of human neutrophils to monomeric immunoglobulin E (IgE). Treatment of neutrophils with IgE caused a transient rise in the level of intracellular calcium that was inhibited by pertussis toxin. The calcium rise was due mainly to release from an intracellular membrane-enclosed store that is also sensitive to the chemotactic peptide formyl-Met-Leu-Phe. The IgE-induced calcium transient was independent of Fc gamma receptors and of Fc epsilon receptor ligation. Our data suggest that the mere binding of IgE to neutrophils is sufficient to evoke a biological response without the need for IgE/receptor cross-linking.

FHIT gene abnormalities in both benign and malignant thyroid tumours.

FHIT, a candidate tumour suppressor gene, has recently been identified at chromosomal region 3p14.2, and deletions of the gene have been reported in many types of human cancers. Loss of heterozygosity (LOH) at this region has also been found frequently in follicular thyroid carcinoma (FTC). To investigate the potential role of FHIT in thyroid tumorigenesis, we examined 57 thyroid tumour specimens (eight benign adenomas, 40 papillary, four follicular and five anaplastic carcinomas), and two thyroid carcinoma cell lines (NPA, SW579) for genetic alterations by using reverse transcription-polymerase chain reaction (RT-PCR), PCR product sequencing, single-strand conformation polymorphism (SSCP) and Southern blot analysis. Two cervical carcinoma cell lines (C-33A, HeLa) were included as positive controls. We detected truncated FHIT transcripts in three of eight (38%) benign adenomas, nine of 40 (23%) papillary, and two of five (40%) anaplastic carcinomas, and in three cell lines (SW579, C-33A, HeLa). Most of the truncated transcripts lacked exons 4 or 5 to 7 or 8 of the gene and were presumably non-functional as the translation start site is located in exon 5. SSCP analysis of the coding exons failed to detect any point mutations among the samples without abnormal FHIT transcripts. Southern blot analysis demonstrated either loss or reduced intensity of major Bam HI restriction fragments in the three cell lines found to have abnormal FHIT transcripts, indicating, respectively, either intragenic homozygous or heterozygous deletions of the FHIT gene. Intragenic homozygous deletions were also found in two papillary thyroid carcinoma specimens: one was missing a 13 kb Bam HI fragment which contains exon 4, the other had deletions of 15.5, 13 and 4.2 kb fragments which contain exons 2 and 9, 4, and 5, respectively. The absence of a defective FHIT gene in FTC indicates that an additional tumour suppressor gene may reside in this region and be involved in the development of FTC. Given that defective FHIT genes were found in both benign and malignant thyroid tumours, the inactivation of this putative tumour suppressor gene is likely to be an early event in the pathogenesis of some forms of thyroid neoplasms.

Activation of coagulation and fibrinolysis in heatstroke.

Hemorrhagic diathesis and widespread microthrombosis are common in heatstroke. To assess the early stages of coagulopathy in heatstroke, thrombin-antithrombin III (TAT), fibrin monomers, plasmin-alpha 2-antiplasmin (PAP), plasminogen and D-Dimer were measured in 16 heatstroke patients (means +/- SE rectal temperature 42.3 +/- 0.2 degrees C) pre- and postcooling and compared with 8 heatstressed and 23 normal controls. Comparing heatstroke patients with normal controls, TAT, fibrin monomers, PAP and D-Dimer were elevated to (median (range)) 16.5 (4-1000) versus 3.5 (2-7.2) micrograms/l p < 0.001, 16 (4-113) versus 2 (2-9) nM p < 0.001; 3300 (1000-36500) versus 255 (136-462) micrograms/l p < 0.001 and 0.72 (0.22-64.8) versus 0.15 (0.05-0.25) microgram/ml p < 0.01 respectively. Plasminogen decreased to 81% (34-106); PAP, TAT and D-Dimer correlated significantly with hyperthermia (r = 0.577, p = 0.02; r = 0.635, p = 0.01; r = 0.76, p = 0.003). Postcooling PAP decreased to 545 (260-850) micrograms/l p < 0.005, TAT 10 (6-70) micrograms/l, and fibrin monomers 22 (18-86) nM remained unchanged. Heatstressed controls showed mild but significant increase in all markers. Activation of coagulation and fibrinolysis occurs early and is profound and sustained in heatstroke. Cooling seems to attenuate the activation of fibrinolysis only, however, this requires confirmation in a larger study population.

A molecular study of EBV DNA and p53 mutations in nasopharyngeal carcinoma of Saudi Arab patients.

Tumor biopsies obtained from 25 Saudi Arab patients with nasopharyngeal carcinoma (NPC) were examined for the presence of Epstein-Barr virus (EBV) DNA detected by the polymerase chain reaction (PCR) and for the incidence of p53 mutations screened by a combination of PCR, single strand conformation polymorphism (SSCP) and PCR-restriction fragment length polymorphism (PCR-RFLP). DNA sequencing was carried out to confirm the occurrence of p53 mutation. While 92% of the tumor specimens were found to carry EBV DNA, only 1/25 showed the incidence of a homozygous mutation at codon 248 of the p53 gene. The data showed that despite a high association of EBV infection with Saudi NPC, the frequency of p53 mutations was very low. Our results are consistent with the worldwide observation of infrequent p53 mutations in NPC.

Elevated pyrogenic cytokines in heatstroke.

STUDY OBJECTIVES: Heatstroke, characterized by hyperthermia and neurologic abnormalities, can cause shock, adult respiratory distress syndrome, and multiorgan failure culminating in death. The mediation of metabolic changes and tissue damage is not fully understood. Recent evidence suggests the involvement of endotoxin, tumor necrosis factor alpha (TNF-alpha), and interleukin 1 alpha (IL-1 alpha) and we hypothesized that other pyrogenic cytokines may be implicated. DESIGN: Prospective analysis. SETTING: Heatstroke Center in Makkah (Mecca), Saudi Arabia. MEASUREMENTS AND RESULTS: We measured plasma IL-1 beta, IL-6, and interferon gamma (INF-gamma) concentrations by enzyme-linked immunosorbent assay in 28 heatstroke patients at the time of hospital admission (precooling) and after complete cooling (postcooling), and in 10 normal control subjects. We measured C-reactive protein (CRP) as a marker of acute phase response and calculated severity of illness using the simplified acute physiology score. Twenty-five male and 3 female subjects had mean (+/- SEM) rectal temperature of 41.2 +/- 0.2 degrees C. IL-6, IL-1 beta, and INF-gamma concentrations were elevated in 100 percent, 39 percent, and 50 percent of patients to (mean +/- SEM) 220 +/- 44 pg/ml, 42 +/- 14 pg/ml, and 1,180 +/- 879 pg/ml, respectively (normal control values: < 3.5 pg/ml, < 4.5 pg/ml, < 20 pg/ml). The CRP value was elevated in 72 percent of patients to 152 +/- 40 mg/L (control value: 0 to 17 mg/L). The IL-6 concentrations correlated with severity of illness (r = 0.516, p = 0.03); two patients with the highest concentrations died. There was no significant correlation between circulating levels of IL-6, IL-1 beta, INF-gamma, and temperature, or between IL-6, IL-1 beta, and CRP. Postcooling, IL-6, and IL-1 beta were still above normal control values; INF-gamma could be detected in one patient only. CONCLUSION: Our findings of elevated circulating IL-6, IL-1 beta, and INF-gamma in the presence of acute phase response, and correlation with severity of illness, suggest that these cytokines have a role in the pathogenesis of heatstroke, which could lead to new therapeutic strategies.

Endotoxemia and release of tumor necrosis factor and interleukin 1 alpha in acute heatstroke.

To determine whether endotoxemia and release of tumor necrosis factor (TNF-alpha) and/or interleukin 1 alpha (IL-1 alpha) are involved in the pathogenesis of heatstroke, 17 adult patients with a mean rectal temperature of 42.1 +/- 0.2 degrees C were studied. Blood samples were taken on admission and after cooling was completed. TNF-alpha and IL-1 alpha levels were measured by enzyme-linked immunosorbent assay, and lipopolysaccharide (LPS) content was measured by the chromogenic substrate modification of the Limulus amebocyte lysate. TNF-alpha, IL-1 alpha, and LPS were elevated in all patients [199 +/- 25 (SE) pg/ml, 480.5 +/- 68.3 pg/ml, and 8.60 +/- 1.19 ng/ml, respectively, compared with normal control values of 31.4 +/- 8.4 pg/ml, 53.7 +/- 5.32 pg/ml, and less than 9 pg/ml]. There was no significant correlation between temperature and the circulating concentration of TNF-alpha, IL-1 alpha, and LPS. Postcooling TNF-alpha, IL-1 alpha, and LPS concentrations were significantly decreased but still above normal control values. The findings suggest that these mediators may have a role in the pathogenesis of heatstroke that could change the strategy of management.

Lymphocyte subsets and adhesion molecules expression in heatstroke and heat stress.

We examined the specificity of the recently reported alterations in circulating lymphocytes in heatstroke by determining lymphocyte subsets in 14 consecutive heatstroke patients before and after cooling and in 7 heat-stressed controls using single- or two-color immunofluorescence flow cytometry. The relationship with catecholamine levels was also studied. In heatstroke, percentages of T (CD3(+)/CD19(-)), T-helper (CD4(+)/CD8(-)), T-inactive [CD3(+)/human leukocyte antigen-DR-], CD11a+, CD11c+, and CD44(+) lymphocytes were significantly decreased, whereas percentages of T-suppressor-cytotoxic (CD8(+)/CD4(-)), natural killer (NK; CD3(-)/CD16(+) or CD56(+)), CD3(+)/CD16(+) or CD56(+), and CD54(+) lymphocytes were significantly increased, compared with 11 normal controls. The changes in the absolute numbers of lymphocyte subsets were in the same direction and were significant for T-helper, T-suppressor-cytotoxic, NK, CD3(+)/CD16(+) or CD56(+), and CD11c+ lymphocytes. Milder but significant changes in percentages of T-helper, T-suppressor-cytotoxic, CD11c+, and CD44(+) lymphocytes were seen in heat stress. Cooling was associated with partial or complete normalization, further derangement (CD11a+, CD11c+), or overcorrection (NK, T-suppressor-cytotoxic, CD11b+) of abnormal percentages of lymphocyte subsets. Norepinephrine levels were significantly elevated in heatstroke (4.7-fold) and heat stress (3.2-fold), but did not significantly correlate with lymphocyte subsets. We conclude that heatstroke is associated with significant changes in percentages and in absolute numbers of a wide range of circulating lymphocyte subsets that are not related to elevated catecholamine levels or totally normalized by cooling. Similar, albeit milder, changes are seen in heat stress, suggesting that the two syndromes represent a continuum.

Distribution of peripheral blood leukocytes in acute heatstroke.

We examined 11 heatstroke patients (mean rectal temperature 41.4 +/- 0.3 degrees C) and 40 healthy subjects to determine the effects of hyperthermia on peripheral blood leukocyte distribution. Precooling samples were taken on admission. Whole blood was incubated with conjugated monoclonal antibodies, and erythrocytes were eliminated by FACS lysing solution. Lymphocyte subsets were detected by specific mouse monoclonal antibodies: Leu-4/CD3+ (T-cells), Leu-3a/CD4+ (T-helper cells), Leu-2a/CD8+ (T-suppressor-cytotoxic cells), Leu-11/19/CD16+/CD56+ (natural killer cells), and Leu-12/CD19+ (B-cells). Immunofluorescence was measured with a flow cytometer. The number of circulating leukocytes and lymphocytes was significantly increased in heatstroke patients. This lymphocytosis was mainly due to an increase in T-suppressor-cytotoxic cells and natural killer cells. The absolute number of lymphocytes and T-suppressor-cytotoxic cells significantly correlated with the degree of hyperthermia (r = 0.62, P = 0.04; r = 0.751, P = 0.007, respectively). There was a significant decrease in the percentages of T-, B-, and T-helper cells and increase in T-suppressor-cytotoxic and natural killer cells, giving a marked decrease in the ratio of T-helper to T-suppressor-cytotoxic cells. We conclude that heatstroke is associated with leukocytosis and significant alteration in absolute number and percentage of circulating lymphocyte subpopulations.

Antibacterial activity of lomefloxacin against Brucella melitensis.

The in vitro activity of lomefloxacin was tested against 114 clinical isolates of Brucella melitensis. Comparison was made with ciprofloxacin, tetracycline, gentamicin, rifampin, and trimethoprim-sulfamethoxazole. Lomefloxacin inhibited 113 (99.1%) of the 114 strains tested at less than or equal to 0.5 microgram/ml. It was comparable to ciprofloxacin, tetracycline and gentamicin in antimicrobial potency. One strain that was previously susceptible to ciprofloxacin and had become resistant after the patient was treated with ciprofloxacin showed cross-resistance to lomefloxacin.

Reduced induction of P53 protein by gamma-irradiation in ataxia telangiectasia cells without constitutional mutations in exons 5, 6, 7, and 8 of the p53 gene.

Ataxia telangiectasia (AT) is an autosomal recessive disease of childhood with several phenotypic characteristics. One of the hallmarks of this syndrome is its hypersensitivity to ionizing radiation, which is believed to be due to defects in DNA repair/processing. In addition to radio-resistant DNA synthesis, both fibroblasts and lymphoblastoid cell lines derived from these patients have been shown to have an impaired G1 arrest and prolonged G2 accumulation of cells indicating a defect in the regulation of cell cycle after irradiation. Since the (tumor suppressor) p53 protein has been reported to participate in the regulation of G1 arrest after irradiation, the possibility of p53 gene mutation and deregulating cell cycle in AT needed to be examined. We used the PCR amplification and DNA sequencing methods to detect mutations in the hypermutable exons (5-8) of germline p53 in fibroblast cells from 3 AT homozygotes. No mutation was found in any of these exons. In order to determine the role of the p53 protein in G1 arrest, its levels were measured before and after gamma-irradiation by flow cytometry in both AT and normal cells. Radiation-induced p53 protein levels in the AT cells varied from 6 to 60% compared to the normal cells, indicating a reduced induction of the protein in AT. These results suggest that mutation in the AT gene affects the p53 induction by irradiation and may, thus, alter the cell cycle regulation in the AT patients.

Prediction of successful embryo implantation by measuring interleukin-1-alpha and immunosuppressive factor(s) in preimplantation embryo culture fluid.

To find a better predictor of pregnancy after in vitro fertilization (IVF), supernatant fluids from embryo culture media were analyzed after 24 hours and 48 hours for the presence of interleukin-1-alpha (IL-1), interleukin-2, and the percent of immunosuppression. The measurements were performed on 108 consecutive IVF cycles between June 1989 and October 1989. The IL-1 level +/- SD in the 24-hour aliquots of the supernatant of embryo culture fluid was 66.2 +/- 10.2 pg/mL in all viable pregnancy cycles and 35.4 +/- 9.01 pg/mL in unsuccessful cycles. The percent of immunosuppression after 24 hours was 22.06% +/- 4.5% in viable pregnancy cycles and 7.3 +/- 5.5% in unsuccessful cycles. The percent of immunosuppression 48 hours after ovum pick-up was generally decreased in all embryo culture fluid, showing 17.5% +/- 4.4% in viable pregnancy cycles and 3.8% +/- 3.6% in unsuccessful cycles. Interleukin-1 levels in the 48-hour aliquots were moderately decreased, being 39.0 +/- 6.3 pg/mL in viable pregnancy cycles and 34.3 +/- 4.7 pg/mL in the unsuccessful cycles. In 24 hours, embryo culture aliquots IL-1 level greater than 60 pg/mL was seen in 17 of 21 (80.9%) pregnancy cycles, and the combined data of IL-1 level greater than 60 pg/mL and/or greater than 20 percent of immunosuppression predicted 21 of 21 (100%) pregnancy cycles.

Anti-tumor cytotoxic potential and effect on human bone marrow GM-CFU of human LAK cells generated in response to various cytokines.

The effects of various recombinant cytokines i.e. IL-1 alpha, IL-3, IL-4, IL-6, IFN-gamma, TNF-alpha and GM-CSF used either alone or in combination with IL-2, were investigated in this study. First, their capacity to induce killer cells from human PBL was examined by evaluating the degree of killing of human NK-sensitive K562 or NK-resistant Daudi cells. Second the effects of these cytokines, LAK cells (at 1/1, 2/1, 4/1 ratio LAK effectors/bone marrow cell targets) and of the supernatants from washed killer cell cultures, were examined on the colony forming ability of human bone marrow for GM-CFU in vitro. Various degrees of NK activity against K562 was observed in PBL stimulated with the cytokines, whereas LAK activity was found only with IL-2 alone. Culture of PBL with IL-2 + IL-1 alpha or IL-2 + IL-6 or IL-2 + GM-CSF resulted in the highest LAK killing. However, addition of TNF-alpha, or IFN-gamma to IL-2 in cultures resulted in a significant suppression of LAK cell activity. Addition of IL-1 alpha, IL-2, IL-3, and IL-4 to BM cultures had little or no effect on day 14 GM-CFU, whereas addition of IL-6 and GM-CSF resulted in a stimulatory effect. LAK cells induced with IL-2 alone had no significant suppressive effects on GM-CFU.(ABSTRACT TRUNCATED AT 250 WORDS)

The effect of antibodies against cell-surface adhesion molecules (LFA-1 alpha and ICAM-1) on the migration and localization of 99mTc-labeled leukocytes in acute infection.

The deficiency of adhesion molecules on leukocytes could severely impair their ability to migrate and perform effective immunological functions leading to clinical situation such as LAD (leukocyte adhesion deficiency) syndrome. We investigated the effects of blocking anti-LFA-1 alpha and ICAM-1 antibody-treated 99mTc-labeled leukocytes on the migration and localization to the site of E. coli-induced acute infection in CBA/J mice. A significant inhibition of migration and localization of antibody-treated leukocytes to the site of infection was observed, reaffirming the vital role of these adhesion molecules, especially during scintigraphic examination of patients for deep infections or abscess using labeled leukocytes.

Changes in peripheral blood lymphocyte subsets associated with marathon running.

The percentage of peripheral blood mononuclear leukocytes that reacted with monoclonal antibodies specific for T-lymphocytes (CD3 cells), the helper/inducer subsets (CD4 cells), and cytotoxic/suppressor subsets (CD8 cells) of T-lymphocytes, and cells with NK activity (CD16 cells) were enumerated by fluorescence-activated flow cytometry for samples obtained immediately before and after the marathon running. It was found that long-term physical exercise resulted in a significant (P < 0.04 for relative and P < 0.008 for absolute lymphocytes) reduction in CD3 cells. A significant (P < 0.009) percentage change was also observed in B lymphocytes (CD19 cells) right after the marathon. The number of NK (CD16 cells) lymphocyte subsets was significantly (P < 0.05 for relative and P < 0.03 for absolute lymphocytes) changed. No significant changes were recorded for CD4, CD8, or CD4/CD8 ratios after the marathon run. A marked leukocytosis was noticed after the endurance exercise and the mean white blood cell (WBC number was increased from 7.8 +/- 2.6 to 22.9 +/- 2.8 x 10(9) cells x 1-1) count was changed by a factor of 2.9. The mean serum cortisol was significantly (P < 0.0001) increased. No hematocrit change was recorded in subjects pre- to post-run. The results of this study demonstrated that long-term physical exercise (marathon running) influenced the T-cell subsets remarkably and produced leukocytosis that was stress dependent and correlated with the increased serum cortisol levels and not the hemoconcentration.

Chronic gamma-irradiation results in increased cell killing and chromosomal aberration with specific breakpoints in fibroblast cell strains derived from non-Hodgkin's lymphoma patients.

Cultured skin fibroblast cells from 6 patients with non-Hodgkin's lymphoma (NHL) and 2 clinically normal subjects were compared for cell survival and chromosomal aberration after chronic gamma-irradiation. Fibroblasts from an ataxia telangiectasia (AT) homozygote and an AT heterozygote were used as positive controls. Following irradiation, fibroblasts from all 6 NHL patients showed an increase in both cell death and chromosomal aberration (breaks and rearrangements) compared to the normal subjects. The difference in the frequency of chromosomal aberration between the normals and the NHL patients remained virtually unchanged over a period of 24-72 h post irradiation incubation of the cells. Cell cycle analysis by flow cytometry carried out in 1 normal and 1 NHL fibroblast cell strain showed that more cells representing the NHL patient were in G2/M phase compared to the normal at various times of cytogenetic analysis. While the AT homozygote appeared to be the most radiosensitive, the AT heterozygote showed a slightly higher incidence of cell death and chromosomal aberration than the normals. The cellular and chromosomal radiosensitivity of fibroblast cell lines from the NHL patients differed slightly from that of the AT heterozygote but clearly occupied an intermediate position between the AT homozygote and the normal subjects. Cells from 3 of the NHL patients showed radiation-induced specific chromosomal breaks involving chromosomes 1, 2, 6, 8, 10 and 11 which correspond to known fragile sites. Such breakpoints associated with increased radiosensitivity may be indicative of predisposition to malignancy in the patients studied.

Post-irradiation DNA synthesis inhibition and G2 phase delay in radiosensitive body cells from non-Hodgkin's lymphoma patients: an indication of cell cycle defects.

In the present study, both post-irradiation DNA synthesis and G2 phase accumulation were analyzed in lymphoblastoid cell lines (LCLs) and fibroblast cell strains derived from (Saudi) patients with non-Hodgkin's lymphoma (NHL), ataxia telangiectasia (AT), AT heterozygotes and normal subjects. A comparison of the percent DNA synthesis inhibition (assayed by 3H-thymidine uptake 30 min after irradiation), and a 24 h post-irradiation G2 phase accumulation determined by flow cytometry placed the AT heterozygotes and the NHL patients in an intermediate position between the normal subjects (with maximum DNA synthesis inhibition and minimum G2 phase accumulation) and the AT homozygotes (with minimum DNA synthesis inhibition and maximum G2 accumulation). The similarity between AT heterozygotes and the NHL patients with respect to the two parameters studied after irradiation was statistically significant. The data indicating a moderate abnormality in the control of cell cycle progression after irradiation in the LCLs and fibroblasts from NHL patients may explain the enhanced cellular and chromosomal radiosensitivity in these patients reported by us earlier. In addition to demonstrating a link between cell cycle abnormality and radiosensitivity as a possible basis for cancer susceptibility, particularly in the NHL patients, the present studies emphasized the usefulness of the assay for 24 h post-irradiation G2 phase accumulation developed by Lavin et al. (1992) in characterizing AT heterozygote-like cell cycle anomaly in cancer patients irrespective of whether they carried the AT gene or any other affecting the cell cycle.

Prostanoid production in the umbilicoplacental arterial tree relative to impaired glucose tolerance.

The purpose of this study was to investigate prostanoid synthesis in different segments of the umbilicoplacental vascular tree and its relationship to impaired maternal glucose tolerance. Segments from the umbilical artery and vein, allantochorionic artery branches, and the cotyledon artery from 21 women with diabetes or impaired glucose tolerance and 10 healthy women were studied. Production of prostacyclin (PGI2) and thromboxane (TxA2) metabolites was determined. The Mann-Whitney U test, Wilcoxon signed-ranks matched-pairs test, Kruskal-Wallis test, analysis of variance, and simple linear regression analysis were used. A two-tailed P value of < 0.05 was considered statistically significant. From the umbilical artery distal to the cotyledon artery, the PGI2 synthesis decreased and the TxA2 synthesis increased gradually towards the periphery in normal pregnancy. The PGI2/TxA2 ratio was more than 200 times higher in the umbilical artery than in the cotyledon artery. The TxA2 production tended in general to be higher in the diabetic group than in the control group, resulting in significantly lower PGI2/TxA2 ratios in some vessels. The prostanoid production was not significantly correlated to maternal HbA1c or cord C-peptide concentrations. The balance between vascular prostacyclin and thromboxane synthesis in the umbilicoplacental arterial tree changed gradually towards the periphery: the more peripheral, the lower the prostacyclin and the higher the thromboxane production. The physiological role of this phenomenon is unknown, but may be of importance for the equilibration of vascular tone between arteries of different calibers. The altered prostanoid balance found in diabetic pregnancy was not directly attributable to the degree of maternal glycemic control, but may reflect increased free radical activity and peroxide production in diabetes.

Prostanoid production in umbilical vessels and its relation to glucose tolerance and umbilical artery flow resistance.

OBJECTIVE: To study prostanoid synthesis in umbilical vessels relative to maternal glucose tolerance and umbilical artery blood flow resistance. STUDY DESIGN: Umbilical artery pulsatility index was determined by Doppler velocimetry in 21 women with diabetes or impaired glucose tolerance and 10 healthy women. Segments from the umbilical artery and vein were incubated and prostacyclin (PGI2) and thromboxane (TxA2) metabolites determined. Statistical analyses with the Mann-Whitney U test, Kruskal-Wallis test, Wilcoxon signed-ranks matched-pairs test, contingency table analysis, Fisher's exact test, and simple linear regression analysis were used and a two-tailed P value of < 0.05 considered statistically significant. RESULTS: No significant difference in PGI2 or TxA2 production was found in umbilical vessels between the women with diabetes/impaired glucose tolerance and controls, but the PGI2/TxA2 ratio in the vein was significantly lower in the diabetes/impaired glucose tolerance group. The umbilical artery pulsatility index was positively correlated to the PGI2/TxA2 ratio in cord vessel segments and to cord plasma TxA2 concentration. The cord plasma TxA2 concentration was significantly higher in cases with a high umbilical artery pulsatility index. The prostanoid production was not correlated to maternal HbA1c or cord plasma C-peptide concentrations. CONCLUSIONS: In association with diabetes, an increased 'peroxide vascular tone' and an enhanced 'endoperoxide shift' between platelets and vascular endothelium may explain the unexpected positive correlation found between the umbilical artery pulsatility index and the vascular PGI2/TxA2 synthesis ratio.

Modified whole blood microtechnique for measurement of mitogens and allogeneic lymphocyte stimulated proliferation of human lymphocytes.

A modified micro whole blood technique for the measurement of human lymphocyte proliferative response upon stimulation with commonly used mitogens such as phytohemagglutinin (PHA), concanavalin-A (con-A), pokeweed mitogen (PKW) and pooled allogeneic lymphocytes is described. The modified microtechnique results are comparable to the conventional techniques using density gradient separated lympocyte. The advantage of this micro whole blood technique is the capability of measuring response to common additional equipment is needed. It can easily be adopted by laboratories performing lymphocyte proliferative assay.