新生儿ABO 溶血病相关危险因素的临床分析

目的:分析新生儿ABO溶血病的临床相关危险因素,提高对新生儿ABO溶血病的防治水平。方法:选择ABO血型不合的孕妇433例,根据以上产妇产前IgG抗A(B)效价、产妇妊娠次数和产妇年龄分别分组,分析各组产妇间发生新生儿溶血病(HDN)的差异及临床相关性。结果:产妇产前IgG抗A(B)效价、产妇妊娠次数和产妇年龄均与HDN发生率呈正相关性(P<0.05);IgG抗A(B)效价>256时,HDN发生率将显著提高(P<0.01);产妇妊娠次数和年龄增加后,HDN发生率将显著增加(P<0.01)。结论:夫妻血型不合的产妇进行产前保健时,应进行IgG抗A(B)效价检查,当IgG抗A(B)效价>64或IgG抗A(B)效价进行性增加时,应及早做好HDN的干预措施;通过减少意外妊娠及高龄产妇数量,可减少HDN的发生。     

Immunoadsorption for removal of anti-A and anti-B blood group antibodies in ABO-incompatible bone marrow transplantation

About 10-15 percent of all patients undergoing allogeneic bone marrow transplantation have a major ABO-incompatibility with their donors. The risk of acute hemolytic reactions due to the infusion of an incompatible donor marrow into the recipient can basically be prevented by recipient antibody depletion or by donor marrow red cell depletion. Nine patients were treated by immunoadsorption using a cartridge with chemically synthesized human blood group A and B antigen as immunoadsorbent for antibody depletion. Within a four-hour-procedure about 2-4 times the patient plasma volume could be processed, thus lowering the anti-A and -B hemagglutinins by 2 to 3 tubes. There was a tendency of better IgG removal when titers initially were high, showing a high antibody clearing capacity. There was no significant correlation between starting titer or amount of plasma volume processed and titer reduction. No decrease in titers were observed in one case. We propose repeated immunoadsorption procedures over 2-3 consecutive days before BMT. The procedure is largely safe and without serious side effects. A major advantage is the avoidance of nonautologous human blood products compared to the conventional plasma exchange. All 8 patients surviving long enough had prompt and stable engraftment of all three cell lines post BMT. No late serological complications occurred when patients were regularly monitored and in vivo adsorption was used when titers increased.     

Do ABO Blood Group Antigens Hamper the Therapeutic Efficacy of Mesenchymal Stromal Cells?

Investigation into predictors for treatment outcome is essential to improve the clinical efficacy of therapeutic multipotent mesenchymal stromal cells (MSCs). We therefore studied the possible harmful impact of immunogenic ABO blood groups antigens – genetically governed antigenic determinants – at all given steps of MSC-therapy, from cell isolation and preparation for clinical use, to final recipient outcome.We found that clinical MSCs do not inherently express or upregulate ABO blood group antigens after inflammatory challenge or in vitro differentiation. Although antigen adsorption from standard culture supplements was minimal, MSCs adsorbed small quantities of ABO antigen from fresh human AB plasma (ABP), dependent on antigen concentration and adsorption time. Compared to cells washed in non-immunogenic human serum albumin (HSA), MSCs washed with ABP elicited stronger blood responses after exposure to blood from healthy O donors in vitro, containing high titers of ABO antibodies. Clinical evaluation of hematopoietic stem cell transplant (HSCT) recipients found only very low titers of anti-A/B agglutination in these strongly immunocompromised patients at the time of MSC treatment. Patient analysis revealed a trend for lower clinical response in blood group O recipients treated with ABP-exposed MSC products, but not with HSA-exposed products.We conclude, that clinical grade MSCs are ABO-neutral, but the ABP used for washing and infusion of MSCs can contaminate the cells with immunogenic ABO substance and should therefore be substituted by non-immunogenic HSA, particularly when cells are given to immunocompentent individuals.     

Protective activity of preparations of blood plasma from donors with a high titer of natural antibodies to Re-glycolipid of gram-negative bacteria

The screening of the preparations of blood plasma obtained from 1,608 donors made it possible to establish the presence of high titers of natural antibodies to Re-glycolipid in 3% of the donors. Donor plasma containing antibodies to Re-glycolipid in a titer of 1:128 ensured a high level of protection for mice in experimental fecal peritonitis. The treatment of 10 patients having commonly occurring forms of peritonitis, caused by Gram-negative bacteria, with the use of such plasma yielded a positive clinical effect. The presence of correlation between the titers of antibodies to Re-glycolipid in blood plasma preparations and the content of high-density lipoproteids, expressed in per cent, was noted.     

Isoimmune anti-A blood group reagents in pigs

Immunization or reimmunization of A-negative pigs with red blood cells (RBC) from A-positive donors yielded anti-A antibodies reacting in high titres with pheno-type A(Ac) RBC and, in some cases, in low dilutions, with phenotype Aw(Ap) RBC also. An attempt to raise the anti-A level by immunization with saliva which contained A substance was likewise successful.
Repeated immunization of A-negative recipients with the RBC of A-positive donors (compatible in all other factors), with the aid of adjuvant, is recommended as the best way of obtaining Aw typing reagents.     

Monoclonal anti-A antibody removal by synthetic A antigen immobilized on specific antibody filters

Removal of blood group anti-A and anti-B antibodies can prevent hyperacute organ rejection in ABO-incompatible transplantation. We are developing an extracorporeal-specific antibody filter (SAF) as an immunoadsorption device for direct removal of ABO blood group antibodies from whole blood, without the need for plasma separation and plasma exchange. A hollow fiber-based small scale SAF (mini-SAF) device was fabricated and synthetic A antigen, Atrisaccharide (Atri) conjugated to activated polyacrylic acid, was immobilized on the fiber lumen surface. Monoclonal antibody anti-A IgM were specifically removed up to 70% of initial antibodies using mini-SAF device. The monoclonal anti-A capture experiments on mini-SAF indicated that antibody removal relative to the initial concentration is independent of inlet concentration in the beginning; however, as the surface starts saturating with bound antibodies, removal becomes dependent on inlet concentration. No significant effect of flow rate on removal rate was observed. The radial diffusion and axial convection-based mathematical model developed for unsteady state antibody removal was in good agreement with the experimental data and showed that the antibody removal rate can be maximized by increasing the antibody-binding capacity of the SAF.     

Isoimmune anti-A blood group reagents in pigs.

Immunization or reimmunization of A-negative pigs with red blood cells (RBC) from A-positive donors yielded anti-A antibodies reacting in high titres with phenotype A(Ac) RBC and, in some cases, in low dilutions, with phenotype Aw(Ap) RBC also. An attempt to raise the anti-A level by immunization with saliva which contained A substance was likewise successful. Repeated immunization of A-negative recipients with the RBC of A-positive donors (compatible in all other factors), with the aid of adjuvant, is recommended as the best way of obtaining Aw typing reagents.     

A Novel Assay for Antibody-Dependent Cell-Mediated Cytotoxicity against HIV-1- or SIV-Infected Cells Reveals Incomplete Overlap with Antibodies Measured by Neutralization and Binding Assays

The resistance of human immunodeficiency virus type 1 (HIV-1) to antibody-mediated immunity often prevents the detection of antibodies that neutralize primary isolates of HIV-1. However, conventional assays for antibody functions other than neutralization are suboptimal. Current methods for measuring the killing of virus-infected cells by antibody-dependent cell-mediated cytotoxicity (ADCC) are limited by the number of natural killer (NK) cells obtainable from individual donors, donor-to-donor variation, and the use of nonphysiological targets. We therefore developed an ADCC assay based on NK cell lines that express human or macaque CD16 and a CD4⁺ T-cell line that expresses luciferase from a Tat-inducible promoter upon HIV-1 or simian immunodeficiency virus (SIV) infection. NK cells and virus-infected targets are mixed in the presence of serial plasma dilutions, and ADCC is measured as the dose-dependent loss of luciferase activity. Using this approach, ADCC titers were measured in plasma samples from HIV-infected human donors and SIV-infected macaques. For the same plasma samples paired with the same test viruses, this assay was approximately 2 orders of magnitude more sensitive than optimized assays for neutralizing antibodies—frequently allowing the measurement of ADCC in the absence of detectable neutralization. Although ADCC correlated with other measures of Env-specific antibodies, neutralizing and gp120 binding titers did not consistently predict ADCC activity. Hence, this assay affords a sensitive method for measuring antibodies capable of directing ADCC against HIV- or SIV-infected cells expressing native conformations of the viral envelope glycoprotein and reveals incomplete overlap of the antibodies that direct ADCC and those measured in neutralization and binding assays.     

Assessment by immunoenzyme analysis of the immune status of donors with reference to the viruses of rubella, measles and herpes simplex

The results of testing the blood sera obtained from donors at a blood transfusion center in Moscow for the presence of antibodies to rubella, measles and herpes simplex viruses, carried out by means of the enzyme immunoassay with the use of the corresponding test systems, are presented. Antibodies to rubella, measles and herpes simplex viruses have been detected, respectively, in 81.5, 96.7 and 100% of blood sera. The proportion of sera with low, medium and high antibody titers has proved to be virtually the same with respect to antibodies to rubella and herpes simplex viruses, the sera with medium antibody titers constituting 59%. At the same time tests for measles antibodies have shown the prevalence of sera with low titers (49.2%) with the highest percentage of seronegative donors (18.5%, as compared with 3.3% in rubella and the absence of negative sera in herpes simplex).     

Carboxyethylpyrrole protein adducts and autoantibodies,biomarkers for age-related macular degeneration

Age-related macular degeneration (AMD) is a slow, progressive disease with both genetic and environmental risk factors. Free radical-induced oxidation of docosahexaenoate (DHA)-containing lipids generates omega-(2-carboxyethyl)pyrrole (CEP) protein adducts that are more abundant in ocular tissues from AMD than normal human donors. To understand better the role of oxidative damage in AMD, we have synthesized CEP-modified proteins, produced anti-CEP antibodies, and initiated analysis of CEP immunoreactivity and autoantibodies in human plasma. A highly selective rabbit polyclonal anti-CEP antibody was raised that binds CEP 1000 times more strongly than carboxypropylpyrrole, a close structural analogue. The CEP adduct uniquely indicates oxidative modification from DHA derivatives because CEP protein modifications cannot arise from any other common polyunsaturated fatty acid. Immunocytochemistry localized CEP to photoreceptor rod outer segments and retinal pigment epithelium in mouse retina and demonstrated more intense CEP immunoreactivity in photoreceptors from a human AMD donor compared with healthy human retina. The mean level of anti-CEP immunoreactivity in AMD human plasma (n = 19 donors) was 1.5-fold higher (p = 0.004) than in age-matched controls (n = 19 donors). Sera from AMD patients demonstrated mean titers of anti-CEP autoantibody 2.3-fold higher than controls (p = 0.02). Of individuals (n = 13) exhibiting both antigen and autoantibody levels above the mean for non-AMD controls, 92% had AMD. These results suggest that together CEP immunoreactivity and autoantibody titer may have diagnostic utility in predicting AMD susceptibility.     

In Vivo Electroporation of a New Gene Vaccine Encoding Ten Repeats of Aβ3-10 Prevents Brain Aβ Deposition and Delays Cognitive Impairment in Young Tg-APPswe/PSEN1dE9 Mice

Active immunization holds great promise for the treatment of Alzheimer's disease but the infiltration of T-lymphocytes and associated meningoencephalitis observed in clinical trials needs to be overcome. To avoid this toxicity, previous studies have used synthetic truncated derivatives of Aβ to promote humoral immunity. In this study, we developed a novel vaccine [p(Aβ3-10)10-MT] that expresses ten repeats of Aβ3-10 with melatonin (MT) as an adjuvant, and administered it intramuscularly in three-month-old Tg-APPswe/PSEN1dE9 (Tg) mice by in vivo electroporation. The p(Aβ3-10)10-MT vaccine induced high titers of anti-Aβ antibodies, which in turn reduced Aβ deposits in the mouse brains and decreased cognitive impairment. Immunoglobulin isotyping revealed a predominantly IgG1 response, indicating a Th2 anti-inflammatory response. Ex vivo cultured splenocytes exhibited a low IFN-γ and high IL-4 response. Immunohistochemical analysis revealed that glial cell activation was also attenuated. These results indicate that p(Aβ3-10)10-MT may potentially be an effective vaccine to reduce accumulated Aβ and attenuate cognitive deficits.     

Maternal IgG anti-A and anti-B titres predict outcome in ABO incompatibility in the neonate

Hemolytic disease of newborn (HDN) is an alloimmune hemolytic disease which occurs due to red blood type incompatibility between mother and fetus. An AB blood type neonate was admitted to Shengjing hospital with severe anemia. Major crossmatch incompatibility was found with some random donors. Serological tests were administered to the neonate and his parents. The mother was B blood type, while the father was AB blood type. The neonate's direct antiglobulin test (DAT) was negative, but the elution test was positive with A₁ cell and negative with A₂ cell. Titers 64 anti-A₁ and 2 anti-A in the mother's serum were detected after treated by dithiothreitol (DTT). The mother's red cells showed a weak agglutination with anti-A under microscopy. The neonate was diagnosed with HDN. After phototherapy and A₂B red blood cell (RBC) transfusion, the neonate was discharged with a recovery of his hemoglobin and physiological index. This study describes a rare case of HDN caused by anti-A1 allo-antibodies.     

The use of blood plasma with elevated titers of antibodies to Re-chemotype glycolipid in peritonitis

The article analyzes the possibility of immunotherapy of septic complications in cases of peritonitis caused by gram-negative bacteria. A strictly inverse correlation between the severity of intoxication and the level of antibodies to glycolipid Re in blood serum has been established. About 5% of healthy nonimmunized donors have elevated Re-antibody titers (1:128 and higher) in their blood plasma. Screening of blood preparations from more than 1000 donors permitted the creation of the blood plasma bank used for the treatment of peritonitis patients. Immunotherapy of such patients has made it possible to decrease almost two-fold the death rate and to reduce the severity and duration of the intoxication syndrome in patients, as well as to improve the results of the treatment of peritonitis.     

A new symmetry: A anti-B is anti-(B anti-A), and reverse enhancement

Immune system network theory leads to a new symmetry, namely that the antibodies produced in an allogeneic A anti-B immune response (where A and B are, say, two different mouse strains), should have complementary shapes to the antibodies in a B anti-A response. That is, A anti-B is anti-(B anti-A). This symmetry is due to the existence of two readily separable populations of antibodies that are present in alloantisera: anti-foreign and anti-anti-self antibodies. The theoretical basis for the symmetry is described, and results indicating the presence of anti-anti-self antibodies in each of 12 alloantisera (six made in B10-congenic strains, and six made with the unrelated chains CBA, SJL, and C57BL/6) are reported. The finding that hyperimmune alloantisera routinely contain anti-anti-self antibodies suggests that network regulation plays an important role in maintaining self-tolerance during responses to allogeneic cells. We further show that A anti-B serum absorbed against B can specifically prolong the survival of A grafts in a B strain animal. We suggest that this result can be interpreted as being due to A anti-(B anti-A) antibodies preventing B anti-A cells from rejecting the A grafts. We call this phenomenon "reverse enhancement" because it involves the converse antiserum to that used in conventional enhancement of graft survival by specific antibodies.     

Plasma exchange for removal of anti-A and anti-B group antibodies in AB0 major-incompatible bone marrow transplantation

Between 1980-1986 seven patients underwent AB0-major incompatible bone marrow transplantation. Incompatible anti-A and anti-B antibodies could be decreased by the plasma exchange from titers 8-256 down to 0-8. Our practice combined with additional depletion of erythrocytes from marrow allowed to transplant all patients successfully, however, one patient demonstrated an acute haemolytic reaction during infusion of marrow. Plasma exchange was fairly well tolerated, however, 4 patients developed slight urticarial reactions. Both thrombocytopenic patients overcompensated the platelet loss caused by the exchange.     

Production of Anti-Amyloid β Antibodies in Mice Fed Rice Expressing Amyloid β

The main signs of Alzheimer’s disease (AD) are cognitive impairment and senile plaques composed of amyloid beta (Aβ) observed in patients’ brains. Therefore, therapy for AD focuses on the removal of Aβ. We developed an “edible vaccine” that employs intestinal immunity with little to no side effects. Rice was utilized as an edible vaccine. It expressed GFP-Aβ42. Aβ rice was administered orally to wild-type (WT) mice causing production of anti-Aβ antibodies. Since Aβ rice was mixed with the cholera toxin B subunit (CTB), antibody against the rice seed protein was also produced. Then, mice were caused to develop immune tolerance against the rice seed protein by oral administration of Aβ rice mixed with CTB. The results indicated that only anti-Aβ antibodies were produced.     

Production of anti-amyloid β antibodies in mice fed rice expressing amyloid β

The main signs of Alzheimer's disease (AD) are cognitive impairment and senile plaques composed of amyloid beta (Aβ) observed in patients' brains. Therefore, therapy for AD focuses on the removal of Aβ. We developed an "edible vaccine" that employs intestinal immunity with little to no side effects. Rice was utilized as an edible vaccine. It expressed GFP-Aβ42. Aβ rice was administered orally to wild-type (WT) mice causing production of anti-Aβ antibodies. Since Aβ rice was mixed with the cholera toxin B subunit (CTB), antibody against the rice seed protein was also produced. Then, mice were caused to develop immune tolerance against the rice seed protein by oral administration of Aβ rice mixed with CTB. The results indicated that only anti-Aβ antibodies were produced.     

Comparative study of expression of hemagglutinins, hemolysins, and enterotoxins by clinical and environmental isolates of non-O1 Vibrio cholerae in relation to their enteropathogenicity

A comparative study was undertaken of clinical and environmental isolates of non-O1 Vibrio cholerae with respect to their hemagglutinating, hemolytic, enterotoxigenic, and enteropathogenic activities. Cell-associated hemagglutinin titers of the clinical and environmental isolates did not differ much, although the clinical isolates displayed higher cell-free hemagglutinin titers compared with those of environmental isolates. Culture supernatants of 61.5% (24 of 39) of clinical isolates showed hemolytic activity (greater than or equal to 10% lysis of rabbit erythrocytes), while only 33.3% (10 to 30) of the environmental group had such activity. Furthermore, hemolytic activities of the clinical isolates showed a good correlation with their cell-associated hemagglutinin titers which was not true for the environmental group. Culture supernatants of 45.8% (11 of 25) of the clinical and 20% (2 of 10) of the environmental isolates exhibited enterotoxigenic activity in the rabbit ileal loop assay. Such activity was mediated mainly by cholera toxin-like substances, although some of the isolates produced fluid-accumulating factors unrelated to cholera toxin. Experimental animal studies demonstrated that the enteropathogenic potential of the environmental isolates was significantly lower than that of the clinical group. Further analysis of our data showed that phenotypic expression of cholera toxin-like products by the non-O1 V. cholerae isolates was accompanied by their enteropathogenicity. The latter effect was also noted with some of the cholera toxin-negative isolates, particularly in those having high hemagglutinating and hemolytic titers.     

Comparative study of expression of hemagglutinins, hemolysins, and enterotoxins by clinical and environmental isolates of non-O1 Vibrio cholerae in relation to their enteropathogenicity.

A comparative study was undertaken of clinical and environmental isolates of non-O1 Vibrio cholerae with respect to their hemagglutinating, hemolytic, enterotoxigenic, and enteropathogenic activities. Cell-associated hemagglutinin titers of the clinical and environmental isolates did not differ much, although the clinical isolates displayed higher cell-free hemagglutinin titers compared with those of environmental isolates. Culture supernatants of 61.5% (24 of 39) of clinical isolates showed hemolytic activity (greater than or equal to 10% lysis of rabbit erythrocytes), while only 33.3% (10 to 30) of the environmental group had such activity. Furthermore, hemolytic activities of the clinical isolates showed a good correlation with their cell-associated hemagglutinin titers which was not true for the environmental group. Culture supernatants of 45.8% (11 of 25) of the clinical and 20% (2 of 10) of the environmental isolates exhibited enterotoxigenic activity in the rabbit ileal loop assay. Such activity was mediated mainly by cholera toxin-like substances, although some of the isolates produced fluid-accumulating factors unrelated to cholera toxin. Experimental animal studies demonstrated that the enteropathogenic potential of the environmental isolates was significantly lower than that of the clinical group. Further analysis of our data showed that phenotypic expression of cholera toxin-like products by the non-O1 V. cholerae isolates was accompanied by their enteropathogenicity. The latter effect was also noted with some of the cholera toxin-negative isolates, particularly in those having high hemagglutinating and hemolytic titers.     

Effects of a Single Histoplasmin Skin Test on the Serological Diagnosis of Histoplasmosis

Numerous reports have indicated that a single histoplasmin skin test may stimulate humoral antibodies to Histoplasma capsulatum antigens in histoplasmin-hypersensitive individuals. Although these investigations concur that antibody elevations are evoked, they vary in the reported degree of incidence and response induced, and they cast doubt on the interpretation of serological tests in the diagnosis of histoplasmosis. Histoplasmin-hypersensitive subjects (114) were bled prior to administration of the skin test, 2 days later, at the time this test was read, and 15 and 30 days after testing. No significant antibody titers were observed at 2 days. At 15- and 30-day intervals, only 17 (15%) of the subjects demonstrated circulating antibodies. All 17 showed agar gel bands; 5 demonstrated no complement-fixation (CF) titers, 10 produced CF antibodies ranging from 1:8 to 1:16, and 2 demonstrated titers of 1:32. The data suggest that skin testing does not interfere significantly with antibody levels in sera drawn approximately 2 days after administration of antigen. However, since titers as high as 1:32 were obtained at later intervals, such reactions should be evaluated cautiously and only after consideration of clinical findings.