Immunochemical characterization of mouse brain-associated theta (Thy-1) antigen.

The Thy-1 antigens or rat brain and thymus have been isolated and chemically characterized, but those of mice have not been identified. Moreover, it is uncertain whether the antigens are glycolipids or glycoproteins. This study with highly purified preparations of gangliosides GM₁, ¹GD_1a, GD_1b and GT_1b from bovine brain and several ganglioside fractions from mouse brain showed that Thy-1 activity does not reside in gangliosides, but rather in the chloroform-methanol-insoluble residue of brain remaining after extraction of gangliosides. The antigen could be solubilized from this residue with a non-ionic detergent. The antigenic activity of the solubilized preparation was heat-labile but resistant to periodate. The chemical properties of the Thy-1 antigen of mouse brain are discussed.     

Expression of nonclassical MHC class Ib genes

Peptide binding proteins of the major histocompatibility complex consist of the “classical” class Ia and “nonclassical” class Ib genes. The gene organization and structure/function relationship of the various exons comprising class I proteins are very similar among the class Ia and class Ib genes. Although the tissue-specific patterns of expression of these two gene families are overlapping, many class Ib genes are distinguished by relative low abundance and/or limited tissue distribution. Further, many of the class Ib genes serve specialized roles in immune responses. Given that the coding sequences of the class Ia and class Ib genes are highly homologous we sought to examine the promoter regions of the various class Ib genes by comparison to the well characterized promoter elements regulating expression of the class Ia genes. This analysis revealed a surprising complexity of promoter structures among all class I genes and few instances of conservation of class Ia promoter regulatory elements among the class Ib genes.     

Preferential apoptosis of CD56dim natural killer cell subset in patients with cancer

Natural killer (NK) cells (CD56(+)/CD3(-)) in the circulation of cancer patients were reported to have low NK activity and undergo spontaneous apoptosis. A possible relationship between apoptosis and impaired NK activity was studied by Annexin V-binding and NK-cell assays performed with peripheral blood mononuclear cells of patients with head and neck cancer (HNC), breast cancer (BC) and normal controls (NC). Cells stained with Annexin V (Anx) and antibodies to CD56, CD3, CD95, CD25, CD122 or CD132 were examined by flow cytometry. NK activity was tested against K562 targets in 4-h (51)Cr-release assays. The ratio of CD56(dim)/CD56(bright) NK cells was significantly different in patients vs. controls (10 vs. 16; p<0.01). A significantly greater percentage of CD56(dim) NK cells bound Anx in HNC patients (27+/-17%, median +/- SD) or BC (46+/-18%) than in NC (15+/-18%, p<0.04 and p<0.0002, respectively). CD56(dim) NK cells were preferentially targeted for apoptosis. NK activity was significantly lower in patients with HNC and BC than in NC (p<0.009). An inverse correlation between NK activity and the percent of Anx(+)CD56(dim) NK cells was observed in cancer patients (p =0.002) but not in NC. In patients, circulating CD56(dim) NK cells were targeted for apoptosis, leading to low levels of NK activity.     

Effect of intravenous immunoglobulin treatment on the Th1/Th2 balance in women with recurrent spontaneous abortions

PROBLEM: The way by which intravenous immunoglobulin (IvIg) acts to prevent immunlogically mediated recurrent spontaneous abortions (RSA) has not been clarified. In the present study, a possible effect of IvIg on the T helper cell (Th1/Th2) balance was investigated in abortions of either alloimmune or autoimmune abnormalities. METHOD OF STUDY: The study included 21 women treated with IvIg before conception because of a history of RSA characterized by alloimmune abnormalities (n = 15) or associated with anti-phospholipid antibodies (APA) (n = 6). Peripheral blood samples, collected before and 5 days after the first IvIg infusion, were stimulated, and Th1 and Th2 cells were detected by flow-cytometric analysis using a combination of monoclonal antibodies against T-cell surface markers and intracellular interferon (IFN)-gamma and interleukin (IL)-4. The percentage of IFN-gamma-producing (Th1) and IL-4-producing (Th2) cells and the Th1/Th2 ratio were compared between pre- and post-infusion samples. RESULTS: A decrease of Th1 percentage in 66.6% of the cases and a concurrent Th2 percentage increase (47.61%) resulted in a decrease in the Th1/Th2 ratio in most of the cases (76.1%) (p < 0.01). Similar results were found in Group A (Th1/Th2 decreased in 60% of the cases, p < 0.05), while in Group B the effect of IvIg was not clear (Th1/Th2 increased in three and decreased in another three cases). CONCLUSION: Our finding suggests that IvIg administration in women with alloimmune RSA enhances Th2 polarization. This is not always the case with APA-associated abortions.     

Pharmacological characteristics of P2 receptors in human uterus

We studied contractile responses of isolated smooth muscles from human uterus induced by P2 receptor agonists. In preparations from pregnant uterus all tested P2 receptor agonists caused smooth muscle contractions. The relative activity of P2 receptor agonists decreased in the following order: a,b-methylene-ATP-uridine triphosphate-ATP. Responses induced by ATP and ADP were similar. The amplitude of contractions induced by alpha,beta-methylene-ATP significantly decreased in the presence of P2 receptor antagonist pyridoxal-phosphate-6-azophenyl-2',4'-disulfonic acid. None of tested P2 receptor agonists induced contractions of isolated myometrium from nonpregnant women. Our results indicate that pregnant human uterus contains P2 receptors mediating the contractile response.     

Ganglioside expression in tissues of mice lacking beta2-microglobulin

This study presents a comparative analysis of gangliosides from lymphoid (spleen and thymus) and other (brain, liver, lungs and muscle) tissues of C57BL/6 mice lacking the gene for beta2-microglobulin (beta2M), a constitutive component of the MHC class I molecule. Ganglioside fractions in the tissues of mice homozygous (beta2M-/-) and heterozygous (beta2M-/+) for the gene deletion were determined by high performance thin-layer chromatography (HPTLC), followed by immunostaining with specific polyclonal antibodies. Ubiquitous gangliosides GM3(Neu5Ac) and GM3(Neu5Gc) were the dominant gangliosides in the lungs of the control beta2M-/+ mice, whereas the homozygous knockout mice had substantially decreased expression of these structures. The lungs of the beta2M-/- mice also had reduced expression of T-lymphocyte-specific GM1b-type gangliosides (GM1b and GalNAc-GM1b). beta2M-deficient mice also had more GM1a and GD1a gangliosides in the liver, and several neolacto-series gangliosides were increased in the brain and lungs. This study provides in vivo evidence that the beta2M molecule can influence the acquisition of a distinct ganglioside assembly in different mouse organs, implicating its non-immunological functions.     

普乐可复对难治性肾病综合征患者血清IL-2、sIL-2R的影响及其临床意义

目的:研究普乐可复(FK506)对难治性肾病综合征患者血清IL-2、sIL-2R的影响及其临床意义。方法:应用酶联免疫吸附法检测难治性肾病综合征患者经普乐可复治疗前后血清IL-2、sIL-2R水平变化,并监测患者24小时尿蛋白、血浆白蛋白、血脂的变化。结果:难治性肾病综合征患者经普乐可复治疗前血清IL-2、sIL-2R水平均显著高于正常对照组(P<0.05)。治疗后血清IL-2、sIL-2R水平较治疗前明显下降(P<0.05)。治疗前24小时尿蛋白水平显著高于正常对照组(P<0.01),血浆白蛋白显著低于正常对照组(P<0.01),血脂水平显著高于正常对照组(P<0.01);与治疗前比较,治疗后24小时尿蛋白水平显著下降(P<0.05),血浆白蛋白水平显著升高(P<0.01),血脂水平显著下降(P<0.05)。治疗后组与正常组比较,除IL-2外,余各项指标均无显著性差异(P>0.05)。结论:在难治性肾病综合征患者体内存在IL-2、sIL-2R的异常,普乐可复对其有明确的抑制作用,从而调节T细胞活性,有效降低24小时尿蛋白,提高血浆白蛋白含量,降血脂,缓解难治性肾病综合征的病情。     

COX-2+ myofibroblasts may play a key role in marked stromal fibrosis in strictured colorectal carcinomas

AIMS: To elucidate the mechanism of marked stromal fibrosis in strictured colorectal carcinomas (SC) that cause complete ileus. METHODS AND RESULTS: Sixteen cases of SC and 29 cases of non-strictured colorectal carcinoma (NSC) were studied. These carcinomas showed similar clinicopathological features except for bowel stricture. The stricture index (SI) showing the degree of bowel stricture was 59.8 +/- 12.1% in SC versus 20.8 +/- 24.6% in NSC (P < 0.001). The fibrosis index (FI), defined to reflect the extent of stromal fibrosis, was 56.3 +/- 8.8% in SC versus 21.9 +/- 10.6% in NSC (P < 0.001). COX-2+ myofibroblasts were detected in 13 cases (81.3%) in SC versus eight cases (27.6%) in NSC (P < 0.01). The COX-2+ myofibroblast density was 276.7 +/- 181.1 cells/mm(2) in SC versus 26.6 +/- 52.7 cells/mm(2) in NSC (P < 0.001). When all cases were divided into two groups with and without COX-2+ myofibroblasts, the SI was 48.8 +/- 19.1% in those with COX-2+ myofibroblasts versus 24.8 +/- 29.3% in those with COX-2- myofibroblasts (P < 0.001). CONCLUSION: COX-2+ myofibroblasts may play an important role in extensive bowel stricture in colorectal carcinomas.     

Caffeine and histamine-induced oscillations of K(Ca) current in single smooth muscle cells of rabbit cerebral artery

In the present experiment, we characterized the intracellular Ca²⁺ oscillations induced by caffeine (1 mM) or histamine (1–3 M) in voltage-clamped single smooth muscle cells of rabbit cerebral (basilar) artery. Superfusion of caffeine or histamine induced periodic oscillations of large whole-cell K⁺ current with fairly uniform amplitudes and intervals. The oscillatory K⁺ current was abolished by inclusion of ethylenebis(oxonitrilo)tetraacetate (EGTA, 5 mM) in the pipette solution. Caffeine- and histamine-induced periodic activation of the large-conductance Ca²⁺-activated K⁺ [K_(Ca)] channel was recorded in the cell-attached patch mode. These results suggest that the oscillations of K⁺ current are carried by the K_(Ca) channel and reflect the oscillations of intracellular Ca²⁺ concentration ([Ca²⁺]_i). Ryanodine (1–10 M) abolished both caffeine- and histamine-induced oscillations. Caffeine- induced oscillations were abolished by the sarcoplasmic reticulum Ca²⁺-adenosine 5-triphosphatase (Ca²⁺-ATPase) inhibitor, cyclopiazonic acid (10 M), and a high concentration of caffeine (10 mM). Inclusion of heparin (3 mg/ml) in the pipette solution blocked histamine-induced oscillations, but did not block caffeine-induced oscillations. By the removal of extracellular Ca²⁺, but not by the addition of verapamil and Cd²⁺, the caffeine-induced oscillations were abolished. Increasing Ca²⁺ influx rate increased the frequencies of caffeine-induced oscillations. Spontaneous oscillations were also observed in cells that were not superfused with agonists, and had similar characteristics to the caffeine-induced oscillations. From the above results, it is concluded, that in smooth muscle cells of the rabbit cerebral (basilar) artery, ryanodine-sensitive Ca²⁺-induced Ca²⁺ release pools play key roles in the generation of caffeine- and histamine-induced intracellular Ca²⁺ oscillations.     

Mucopolysaccharidosis type II (Hunter disease): Identification and characterization of eight point mutations in the iduronate-2-sulfatase gene in Japanese patients

Mucopolysaccharidosis type II (Hunter disease) is a lysosomal storage disorder caused by a deficiency of the enzyme iduronate-2-sulfatase. Varied clinical phenotypes of this disease have been described. To identify mutations in individual patients and to examine possible correlations between mutations and clinical phenotypes, we analyzed the iduronate-2-sulfatase gene in Japanese patients with different clinical phenotypes. Five missense mutations, S333L (severe), R468Q (severe), R468L (severe), W337R (intermediate), R48P (mild), and three nonsense mutations, W345X (severe), R443X (intermediate), Q531X (mild), were identified by the RT-PCR method. Transient expression in the enzyme-deficient fibroblasts revealed that all five missense mutant enzymes were synthesized as the normal-size precursor (73 kD), and the nonsense mutant enzymes were synthesized as truncated ones (W345X:54 kD, R443X:59 kD, and Q531X:69 kD), although stable mature enzymes (45–56 kD) were not detected by Western blot analysis. Further more, expression of the eight mutant cDNAs resulted in severe reductions of iduronate-2-sulfatase enzyme activity in comparison with a normal cDNA. © 1995 Wiley-Liss, Inc.