Expression Pattern of Tumor Necrosis Factor-α-Induced Protein 8-Like 2 in Acute Rejection of Cardiac Allograft.

BACKGROUND: Tumor necrosis factor-α-induced protein-8 like-2 (TIPE2) is a negative regulator of innate immunity and cellular immunity, yet the expression pattern of TIPE2 in acute rejection of cardiac allograft remain enigmatic. METHODS: We developed cardiac transplantation models and divided into 3 groups: a naive group, a syngeneic group, and an allogeneic group. Then, we detected the messenger RNA and protein of TIPE2 in cardiac allografts. Real-time polymerase chain reaction showed expression of CD4 and CD8 in the donor heart, and immunofluorescence assay revealed the association between T cells and TIPE2. RESULTS: In our study, we first found that the expression of TIPE2 in cardiac allografts is upregulated compared with the syngeneic control, and increases in a time-dependent manner. The immunocytochemistry of heart grafts revealed a strong expression of TIPE2 in the inflammatory cells, but not in the cardiomyocytes. Finally, we proved that CD4+ and CD8+ T cells infiltrated cardiac allografts abundantly, which express ample TIPE2. CONCLUSIONS: The upregulated expression of TIPE2 in cardiac allografts, mainly came from T cells, which infiltrated the donor heart. This finding indicates that there may be an association between TIPE2 and acute cardiac allograft rejection.

Regulation of bone-related genes expression by bone-like apatite in MC3T3-E1 cells.

Bone-like apatite on HA/TCP ceramics sintered at 1,100 degrees C (HT1) and 1,200 degrees C (HT2) could be obtained via immersing substrates into simulated body fluid (SBF) for 3 days. When MC3T3-E1 preosteoblastic cells cultured on the surface of the bone-like apatite for 3 days, SEM observations revealed cell membrane features with secreted crystals very similar to in vivo bone formation during intramembranous ossification with a direct bone apposition on the ceramics. According to semi-quantitative RT-PCR method, mRNA expressions of osteocalcin (marker of late-stage differentiation) and type 1 collagen were increased in cultures with HT1S and HT2S when compared to HT1 and HT2 after cultured for 6 days. The results indicated that bone-like apatite had the ability to support the growth of osteoblast-like cells in vitro and to promote osteoblast differentiation by stimulating the expression of major phenotypic markers. Taken together, our findings will be helpful in understanding the mechanism of osteoinductivity of calcium phosphate ceramics and in constructing more appropriate biomimetic substrate.

Amniotic fluid soluble fas levels in intra-amniotic infection.

OBJECTIVE: Membrane Fas can induce apoptosis in sensitive cells. It has been reported that soluble Fas (sFas) is elevated in septicemia. We examined amniotic fluid (AF) sFas levels in patients with and without intra-amniotic infection. METHODS: Forty-two AF specimens were studied. Intra-amniotic infection was defined as the presence of a positive AF culture. Twenty-one specimens were from patients with intra-amniotic infection and 21 were from patients without intra-ammotic infection. Amniotic fluid sFas was determined by an enzyme immunoassay and normalized by AF creatinine levels. The Mann-Whitney U test, contingency table method, and Spearman's rank correlation test were used for statistical analyses. Data were expressed as median with ranges. RESULTS: There were no significant differences in maternal age, gestational age, parity, and race between the groups. The median AF sFas was significantly higher with intra-amniotic infection than without it (5.07 U/mL, range 0.32-13. 25 compared with 1.95 U/mL, range 0.01-5.35; P =.004). After normalizing to AF creatinine, infected fluids also had significantly higher median sFas/creatinine than uninfected amniotic fluids (289.1 U/mg creatinine, range 16.6-920.5 compared with 126.8 U/mg creatinine, range 0.5-546.2; P =.01). Amniotic fluid sFas and sFas/creatinine were positively correlated with AF leukocytes and negatively correlated with AF glucose. CONCLUSION: Elevated AF sFas is associated with intra-amniotic infection. High production of AF sFas in intra-amniotic infection may play a role in the inhibition of apoptosis of AF leukocytes, leading to the persistence of inflammation.

Effects of temperature and temperature-steps on circadian locomotor rhythmicity in the blow fly Calliphora vicina.

The free-running period (in darkness) of the locomotor activity rhythm in adult blow flies (Calliphora vicina) was temperature-compensated between 15 and 25 degrees C, showing Q(10) values between 0.98 and 1.04. Single steps-up (20 to 25 degrees C) or steps-down (20 to 15 degrees C) in temperature caused stable phase shifts of the activity rhythm, giving rise to temperature-step phase response curves (PRCs) with both advances and delays. Phase advances, however, were dominant for steps-up, and phase delays for steps-down; the two PRCs were almost "mirror images" of each other. Following protocols introduced by Zimmerman et al. [(1968) Temperature compensation of the circadian oscillation in Drosophila pseudoobscura and its entrainment by temperature cycles, Journal of Insect Physiology, 14, 669-684] for the rhythm of pupal eclosion in Drosophila pseudoobscura, the steps-up and steps-down PRCs for C. vicina were used to compute a theoretical PRC for a 6 h low temperature pulse, and from this a theoretical steady-state phase relationship of the locomotor activity rhythm to a train of such pulses making up a temperature cycle (18 h at 20 degrees and 6 h at 15 degrees C).

Comparative physical and genetic maps of the virulence plasmids of Salmonella enterica serovars typhimurium, enteritidis, choleraesuis, and dublin.

Using fragment profiling, PCR, and Southern hybridization, we found that Salmonella enterica serovar Choleraesuis harbored virulence plasmids of various sizes, whereas serovars Typhimurium, Enteritidis, and Dublin carried a plasmid of a unique size. Also, the virulence plasmid of Typhimurium contained genes in the same order detected in the other three plasmids, all of which contained deletions.

Elevated amniotic fluid levels of leukemia inhibitory factor, interleukin 6, and interleukin 8 in intra-amniotic infection.

OBJECTIVE: The study's objective was to determine and correlate amniotic fluid levels of leukemia inhibitory factor, interleukin 6, and interleukin 8 in patients with and without intra-amniotic infection. STUDY DESIGN: Amniocentesis was performed on 41 pregnant women with preterm contractions, labor, or premature rupture of membranes. Intra-amniotic infection was defined as the presence of a positive amniotic fluid culture result. Amniotic fluid tests for Gram stain, glucose, leukocyte counts, creatinine level, pH, and specific gravity were performed. Amniotic fluid levels of leukemia inhibitory factor, interleukin 6, and interleukin 8 were measured by an enzyme-linked immunoassay. Unlike in previous reports, cytokines were normalized by amniotic fluid creatinine levels. RESULTS: Fifteen patients had intra-amniotic infection and 26 did not. Amniotic fluid median levels of leukemia inhibitory factor, interleukin 6, and interleukin 8 were significantly higher in pregnant women with intra-amniotic infection than in those without intra-amniotic infection (leukemia inhibitory factor median 3912 pg/mg creatinine, range 0.0-199314, vs 56 pg/mg creatinine, range 0. 0-12148, P =.01; interleukin 6 median 2005 ng/mg creatinine, range 27-4071, vs 990 ng/mg creatinine, range 7.5-3409, P =.005; interleukin 8: median 4933 ng/mg creatinine, range 0.0-55058, vs 61 ng/mg creatinine, range 0.0-2399, P =.005). Amniotic fluid levels of leukemia inhibitory factor, interleukin 6, and interleukin 8 were positively correlated. CONCLUSIONS: The data indicate that leukemia inhibitory factor plays an important role in the pathogenesis of intra-amniotic infection. In addition, significant elevations of and correlations among amniotic fluid levels of leukemia inhibitory factor, interleukin 6, and interleukin 8 suggest that measurements of these cytokines in amniotic fluid may be of diagnostic and prognostic importance.

Glycosylated hemoglobin in insulin-dependent diabetes mellitus related to preeclampsia.

Preeclampsia may occur due to failure of the second wave of trophoblastic invasion between 16 to 20 weeks of gestation. We studied whether the mean value of glycosylated hemoglobin (HbA1c) during this gestational period was associated with an increased incidence of preeclampsia in insulin-dependent diabetes mellitus. We conducted a retrospective study of 131 insulin-dependent diabetes mellitus (IDDM) pregnancies with HbA1c values available in medical records over the past 10 years. The correlation between high mean HbA1c (>8%) before or after 20 weeks and the incidence of preeclampsia was determined. Mean HbA1c values before 20 weeks were further divided into 10- to 15- and 16- to 20-week intervals. Mean values of HbA1c in these different gestational periods were analyzed by chi(2) or Fisher's exact tests. P < 0.05 was considered statistically significant. We found that IDDM pregnant women with high mean values of HbA1c (>8%) had a significantly higher rate of preeclampsia than those with normal mean values (45 vs. 24%, p = 0.018). However, we found that IDDM pregnant women with an elevated mean HbA1c value at 16-20 weeks', but not 10-15 weeks' or after 20 weeks', gestation had a significantly higher incidence of preeclampsia than those with normal mean HbA1c value (chi(2) = 4.49, p = 0.03). We conclude that a significant association between elevated mean HbA1c values at 16-20 weeks' gestation and a high frequency of preeclampsia in IDDM pregnancies suggests that glycosylated hemoglobin may play an important role in the pathogenesis of preeclampsia in IDDM pregnant women.

Elevated amniotic fluid nitric oxide metabolites and interleukin-6 in intra-amniotic infection.

OBJECTIVE: To compare amniotic fluid nitric oxide metabolites and interleukin-6 (IL-6) concentrations in patients with and without intra-amniotic infection. METHODS: Amniotic fluid nitric oxide metabolites, IL-6, Gram stains, glucose, leukocyte counts, leukocyte esterase activity, creatinine, pH, and specific gravity were determined in 14 patients with intra-amniotic infection and 26 patients without intra-amniotic infection. Intra-amniotic infection was defined as the presence of a positive amniotic fluid culture. The nitric oxide metabolites, nitrate and nitrite (NOx), were measured using Greiss reagent after reduction of nitrate to nitrite with aspergillus nitrate reductase. Interleukin-6 was measured by a two-site, enzyme-linked immunosorbent assay. Amniotic fluid nitric oxide metabolites and IL-6 concentrations were normalized by amniotic fluid creatinine levels. The Mann-Whitney U test, contingency table method, and Spearman's rank correlation test were used for statistical analyses. RESULTS: Amniotic fluid NOx and IL-6 levels were significantly higher in patients with intra-amniotic infection than in those without intra-amniotic infection (NOx: median = 2.06 mumol/mg creatinine, range = 0.74-6.81 versus 1.35 mumol/mg creatinine, range = 0.99-1.60, P = .01, IL-6: median = 2.00 micrograms/mg creatinine, range = 0.026-4.07 versus median = 0.04 micrograms/mg creatinine, range = 0.004-3.210, P = .0009, respectively). Patients with intra-amniotic infection had significantly elevated leukocyte counts, leukocyte esterase activity, Gram positive stains, and significantly lower amniotic fluid glucose levels compared with those without intra-amniotic infection. There were no differences in gestational age, maternal age, parity, race, pH, or specific gravity between the two groups. Amniotic fluid NOx was significantly correlated with IL-6 (r = .4, P = .02). Both amniotic fluid NOx and IL-6 were also positively correlated with amniotic fluid leukocyte counts, leukocyte esterase activity and Gram stains, and negatively correlated with glucose levels. CONCLUSIONS: Amniotic fluid NOx and IL-6 are significantly elevated and positively correlated during intra-amniotic infection. Both increased amniotic fluid IL-6 and nitric oxide may exert cytotoxic and cytostatic effects on the target cells. We suggest that measurements of amniotic fluid NOx and IL-6 may serve as useful clinical markers in patients with intra-amniotic infection.

Dual roles of amniotic fluid nitric oxide and prostaglandin E2 in preterm labor with intra-amniotic infection.

We hypothesized that induction of nitric oxide synthase and cyclo-oxygenase-2 by bacterial products in intra-amniotic infection could increase the production of proinflammatory nitric oxide and prostaglandin E2 (PGE2) and cause preterm labor. Thus, we sought to determine amniotic fluid levels of nitric oxide metabolites (NOx) and PGE2 in preterm labor patients with and without intra-amniotic infection. Amniotic fluid from 13 preterm labor patients with intra-amniotic infection and 24 without intra-amniotic infection were studied. Intra-amniotic infection was defined as the presence of a positive amniotic fluid culture. Amniotic fluid was tested for NOx, PGE2, glucose, leukocyte counts, Gram stains, creatinine, pH, and specific gravity. NOx was determined using Griess reagent after reduction of nitrate to nitrite with aspergillus nitrate reductase. PGE2 was measured by an enzyme-linked immunoassay. Both amniotic fluid NOx and PGE2 were normalized by amniotic fluid creatinine. We found that amniotic fluid concentrations of NOx and PGE2 were significantly higher in preterm labor patients with intra-amniotic infection compared to those without intraamniotic infection (NOx: median 1.8 micromol/mg creatinine, range 0.7 to 6.8 vs. 1.3 micromol/mg creatinine, range 0.9 to 2.1, p=0.03; PGE2: median 33.5 ng/mg creatinine, range 0.0 to 1048.6 vs. 0.0 ng/mg creatinine, range 0.0 to 33.6, p=0.004). In addition, amniotic fluid NOx and PGE2 were positively correlated (r=0.343, p=0.0398). We conclude that there may be an interaction between the nitric oxide and prostaglandin pathways in intraamniotic infection. Increased production of amniotic fluid pro-inflammatory nitric oxide and PGE2 may play an important role in the pathogenesis of preterm labor in patients with intra-amniotic infection.

Strategies for reducing the frequency of preeclampsia in pregnancies with insulin-dependent diabetes mellitus.

The object of this study was whether improving glycemic control and maintaining normal glycosylated hemoglobin (HbA1c) through pregnancy can reduce the frequency of preeclampsia. One hundred and twenty-three complete medical records of pregnant insulin-dependent diabetics (IDDM) managed at Yale-New Haven Hospital from 1983 to 1993 were reviewed. Serial HbA1c measurements and the occurrence of preeclampsia were recorded. Based on the change of HbA1c values through the pregnancy, glycemic control was categorized into four groups: group 1, high to normal; group 2, high to high; group 3, normal to normal; group 4, normal to high. The association between HbA1c change and the incidence of preeclampsia was analyzed by chi-square test and Fisher's exact test. Among 123 IDDM pregnancies, 40 (32.5%) developed preeclampsia. High HbA1c levels at any time in IDDM pregnancies were associated with an increased incidence of preeclampsia (group 1 or 2 or 4 versus group 3). Reducing HbA1c by improving glycemic control both before and during pregnancy resulted in a significantly lower incidence of pre-eclampsia (group 3 versus groups 1+2+4, p < 0.05). The best strategy for reducing the frequency of preeclampsia in IDDM pregnancies is by improving glycemic control before pregnancy.

S-antigen antibody partially blocks entrainment and the effects of constant light on the circadian rhythm of locomotor activity in the adult blow fly, Calliphora vicina.

Injection of S-antigen (arrestin) antibody into the brain of the blow fly, Calliphora vicina, appeared to reduce sensitivity to the photic effects of a light:dark cycle (LD; entrainment) or continuous "bright" light (LL; arrhythmicity). In LD, a proportion of the injected flies evaded entrainment or showed delayed entrainment. In bright LL, flies continued with a free-running rhythm, which remained unchanged (as in continuous darkness) or lengthened (as in "dim" LL). These results focus attention on four groups of arrestin-positive neurons in the fly's brain as potential components of the photoreceptive system.

Thrombomodulin levels in preeclampsia, gestational hypertension, and chronic hypertension.

OBJECTIVE: To measure the circulating levels of thrombomodulin in women with preeclampsia, gestational hypertension, and chronic hypertension. METHODS: Serum levels of thrombomodulin were measured in 34 women with preeclampsia, 15 with gestational hypertension, 11 with chronic hypertension, and 34 normotensive pregnant women in the third trimester. Preeclampsia, gestational hypertension, and chronic hypertension were defined by ACOG criteria. Soluble thrombomodulin antigen was measured by a two-site enzyme-linked immunosorbent assay. RESULTS: Mean (+/- standard error) serum thrombomodulin levels were significantly higher in patients with preeclampsia (69.7 +/- 6.3 ng/mL) than in those with gestational hypertension (46.0 +/- 3.2 ng/mL) or chronic hypertension (46.2 +/- 3.3 ng/mL), and normotensive controls (50.1 +/- 3.1 ng/mL). There were no significant differences among the gestational hypertension, chronic hypertension, and normotensive control groups. CONCLUSION: Thrombomodulin may serve as a clinically meaningful marker to differentiate preeclampsia from other forms of hypertensive disorders in pregnancy.

Does preeclamptic pregnancy increase fetal-maternal hemorrhage?

Since pathologic changes of the placenta have been found in preeclamptic pregnancies, a relationship between fetal-maternal hemorrhage and preeclampsia was investigated. Sixty-two women with singleton pregnancies in the third trimester were studied. Thirty-one of these women with preeclampsia were matched with 31 normotensive healthy pregnant women. Maternal serum levels of alpha-fetoprotein (MSAFP) and Kleihauer-Betke (KB) test were determined to reflect the degree of fetal-maternal hemorrhage. There were no significant differences in MSAFP levels, MSAFP to creatinine ratio, or the appearance of fetal cells as measured by the KB test between preeclamptic pregnancies and matched controls. Our data do not suggest that fetal-maternal hemorrhage is increased in pregnant women with preeclampsia.

Plasma thrombomodulin levels in women with systemic lupus erythematosus.

Thrombomodulin (TM), an endothelial cell membrane glycoprotein, is released into blood as a soluble TM antigen after inflammation or injury to endothelium. Systemic lupus erythematosus (SLE) is characterized by multisystem inflammation of vascular endothelium. The objective of this study is to determine the circulating TM levels in pregnant and nonpregnant SLE women and the correlation of plasma TM levels with variables used to assess SLE activity. We found that there were no significant differences in plasma TM levels among pregnant SLE, nonpregnant SLE, and normal pregnant patients. However, significantly higher plasma TM levels were found in some SLE women with active disease or preeclampsia.

Elevated urine thrombomodulin/creatinine ratio in severely preeclamptic pregnancies.

A relationship between the urine thrombomodulin/creatinine ratio and preeclampsia was investigated. Preeclamptic women had a higher urine thrombomodulin/creatinine ratio than did matched controls. However, significantly elevated urine thrombomodulin/creatinine ratios were found in severely, but not mildly, preeclamptic pregnancies.

Elevated serum human chorionic gonadotropin as evidence of secretory response in severe preeclampsia.

OBJECTIVE: Because preeclampsia is a trophoblastic disorder and human chorionic gonadotropin is secreted from trophoblast, we sought to determine whether measurement of serum human chorionic gonadotropin might reflect a different trophoblastic secretory response of preeclampsia. STUDY DESIGN: Twenty patients with mild preeclampsia and 12 with severe preeclampsia were matched with 32 healthy, normotensive women in the third trimester with singleton pregnancies. Serum total human chorionic gonadotropin and total human chorionic gonadotropin-beta were measured by a two-site immunoenzymometric assay, and total hCG-alpha was determined by a double-antibody radioimmunoassay. Wilcoxon signed-rank and Mann-Whitney rank-sum tests were used for statistical analysis. RESULTS: Serum total human chorionic gonadotropin, total human chorionic gonadotropin-alpha, and total human chorionic gonadotropin-beta levels were significantly higher in severely preeclamptic women (p < 0.05), but not in those with mild preeclampsia, compared with those in their matched controls. CONCLUSION: Elevated serum human chorionic gonadotropin levels in severely preeclamptic women might reflect a significantly pathologic change and secretory reaction of the placenta.

Elevated circulating thrombomodulin in severe preeclampsia.

A relationship between serum thrombomodulin and preeclampsia was investigated. In women with severely preeclamptic pregnancies, serum thrombomodulin levels were found to be significantly elevated as compared with those of matched control subjects (p < 0.005). Serum thrombomodulin levels correlated positively with serum creatinine (r = 0.854, p < 0.0001) and uric acid levels (r = 0.784, p < 0.001).