A2 Antigen Research Articles

Генетическая структура крупного рогатого скота разных пород по антигенным факторам крови

Aim. The study of the genetic polymorphism of erythrocyte antigens in Kalmyk, Kazakh white-headed, Hereford and Simmental cattle, to evaluate their genetic structure, and to compare the blood groups of Kalmyk cattle in the Republic of Kalmykia and the Astrakhan region. Materials and Methods. A comparative characterization of the genetic structure by blood groups of Kalmyk cattle of the Republic of Kalmykia with cattle of the Kazakh Whitehead, Hereford, Simmental and Kalmyk breeds of the Astrakhan region was carried out. Blood group testing was conducted according to standard methods (Neimann-Sorensen) using monospecific serum (Nei M. and Li W.H., 1979) produced by JSC "Samarskoye" for breeding work. Results. Immunogenetic testing of animals was performed according to standard methods (Neimann-Sorensen) using reagents – monospecific serum produced by JSC "Samarskoye" for breeding work. Analysis of immunogenetic testing of Kalmyk cattle in the Republic of Kalmykia established the highest frequency of B2 (69%), E'3 (82%), C1 (88%), W (67%), Z (71%) antigens and the lowest frequency of B' (15%), Q (10%), C2 (17%), F (19%). Blood group testing of Kazakh white-headed breed showed that A1, I', C1, W, L, F antigens had a high frequency index and were 63; 79; 73; 66; 91; 87%, respectively. The antigens B2 (20%), G3 (11%), Y2 (2%), F' (11%), C2 (5%), X2 (5%), U' (12%) had a low frequency rate. Antigens such as O2 and H'' were not found at all. The highest frequency of occurrence of the Hereford antigens was in G2 (69%), O3 (65%), O4 (58%), Y2 (67%), Q (61%), C2 (72%), X2 (59%), L (71%), V (68%), the lowest in B2 (15%), I1 (14%), I' (17%), R2 (13%). The highest frequency of the Simmental breed was observed for the A2 antigen at 66%, Y2 at 77%, C1 at 91%, F at 90%, and H' at 65%. The lowest frequency was observed for the B2 antigen – 5%, I1 – 12%, F' – 11%. Antigens A'2, I', W and U' did not occur at all. Blood group testing of Kalmyk breeds of Astrakhan cattle showed the highest frequency of antigens A2 – 100%, I1 – 70%, A'2 – 90%, O4 – 70%, E'3 – 92%, F' – 70%, R2 – 94%, L – 90%, F – 71% and Z – 100%. Antigens with minimal frequency are O2 (1%), I' (2%), C2 (11%), S1 (1%). Antigens such as O3 and Q were not observed at all. Conclusion. It has been established that the cattle of the Kalmyk breed bred in Kalmykia differs from the animals bred in another zone, providing further improvement of the breed. The genetic status of the breeding stock of the Kalmyk cattle of the Republic of Kalmykia and the Kazakh white-headed, Hereford, Simmental and Kalmyk of the Astrakhan region indicates genetic diversity, multidirectional selection and breeding work in the herds.

Immunogenetic status and breeding value of Holstein cows

In the genealogical structure of the breeding stock, the highest specific weight is occupied by the Reflection Sovering and Vis bek Aydiala lines, accounting for 56.8 and 39.9% of animals, respectively. The study results of Holstein cattle blood-group factors can be used to monitor its gene pool, genetic intrabreed differentiation, assess selection parameters, and conduct targeted selection of parental pairs. It was determined that in EAA blood group system in Holstein cows of reproduction from the USA, the frequency of A1 antigen is 58.7%, and in domestic reproduction animals, this antigen is found with a lower frequency of 50.7%. At the same time, in the specified system of blood groups in both groups of experimental animals, antigen “a” is characterized by the highest frequency, the lowest occurrence is A2 antigen, and animals with A1 antigen took an intermediate position. In EAB system, the US Holstein cows have a higher frequency of such antigens as Е3/, О3 и I1 compared to the reproductive animals of Agro-Soyuz LLC. The EAF blood group system is represented by two antigens, where F antigen frequency in both groups of experimental animals is 100%. Of all the blood group systems, the greatest differences between compared groups of animals were detected for the antigens of EAZ blood group system.

Repeat A2 Into B Kidney Transplantation After Failed Prior A2 Into B Transplant: A Case Report

Kidneys from donors with blood type A2 can be successfully transplanted into blood type B and O recipients without the need for desensitization if the recipient's starting anti-A hemagglutinin titer is within an acceptable range. National kidney allocation policy now offers priority for eligible B recipients to receive A2 or A2B deceased donor kidneys, and therefore, the frequency with which A2 or A2B to B transplants will occur is expected to increase. The precise mechanisms by which antibody-mediated rejection is averted in these cases despite the presence of both circulating anti-A antibody and expression of the A2 antigen on the graft endothelium are not known. Whether this process mirrors proposed mechanisms of accommodation, which can occur in recipients of ABO incompatible transplants, is also not known. Repeated exposure to mismatched antigens after retransplantation could elicit memory responses resulting in antibody rebound and accelerated antibody-mediated rejection. Whether this would occur in the setting of repeated A2 donor exposure was uncertain. Here we report the case of a patient with history of a prior A2 to B transplant which failed owing to nonimmunologic reasons; the patient successfully underwent a repeat A2 to B transplant. Neither rebound in anti-A2 antibody nor clinical evidence of antibody-mediated rejection were observed after the transplant. Current kidney allocation will likely enable more such transplants in the future, and this may provide a unique patient population in whom the molecular mechanisms of incompatible graft accommodation may be investigated.

Is It Time to Reconsider the Concepts of "Universal Donor" and "ABO Compatible" Transfusions?

Is It Time to Reconsider the Concepts of "Universal Donor" and "ABO Compatible" Transfusions?

Field randomized trial to evaluate the efficacy of the Leish-Tec® vaccine against canine visceral leishmaniasis in an endemic area of Brazil

BackgroundA canine vaccine remains a promising approach for effective control of visceral leishmaniasis (VL), given its complex epidemiology in areas where zoonotic VL is prevalent. Leish-Tec® is a recombinant vaccine, based on the Leishmania A2 antigen, against canine VL (CVL). It is, since 2014, the single commercial vaccine licensed in Brazil. Here, Leish-Tec® efficacy was estimated through a randomized field trial (RFT), in a highly VL endemic area. MethodsThe RFT was conducted from 2008 to 2010 in an endemic area of southeastern Brazil, presenting a CVL seroprevalence of 41.9%. Eight hundred forty-seven seronegative dogs were randomly selected to receive Leish-Tec® (n=429) or placebo (n=418). Animals were followed up by clinical, serological, and parasitological exams for 18 months. The CVL incidence in both groups was compared through proportion analysis. ResultsA significant reduction in the number of cases of CVL was observed in the vaccine group, as compared with the placebo group, whether efficacy was estimated according to parasitological results (71.4%; 95% CI: 34.9–87.3%; p=0.001; risk ratio=0.287), by adding results of xenodiagnosis and parasitological exams (58.1%; 95% CI: 26.0–76.3%; p=0.002; risk ratio=0.419). Among the animals that converted to a positive anti-A2 serology, efficacy reached 80.8% (95% CI: 37.6–94.1%, p=0.001; risk ratio=0.192). Xenodiagnosis has detected a reduction of 46.6% (p=0.05) in transmission to sand flies from vaccinated animals presenting anti-A2 positive serology. ConclusionThe Leish-Tec® vaccine proved significantly effective for prophylaxis of CVL, after natural challenge assured by transmission of Leishmania parasites, in a highly endemic area. Noteworthy, this report has unveiled the complexity of performing a RFT for anti-CVL vaccines in Brazil, which may be helpful for designing of future studies.

First Report on the OPTN National Variance: Allocation of A2 /A2 B Deceased Donor Kidneys to Blood Group B Increases Minority Transplantation.

In 2002, the Organ Procurement and Transplantation Network (OPTN) Minority Affairs Committee (MAC) implemented a national, prospective, "variance of practice" to allow deceased donor, ABO blood group incompatible, A2 antigen, kidney transplantation into blood group B recipients; outcomes of this cohort were compared to ABO compatible recipients. The goal of the variance was to increase the number of transplants to B candidates without negatively impacting survival or compromising system equity. Only B recipients with low anti-A IgG titers (<1:8) were eligible to receive these kidneys. Across eight participating Donation Service Areas (DSA), there were 101 A2 /A2 B to B transplants through 12/31/11, of which the majority of the recipients (61%) were ethnic minorities. At 12, 24, and 36 months, Kaplan-Meier graft survival rates for the B recipients of A2 /A2 B kidneys were 95.0%, 90.6%, and 85.4%, respectively, comparable to outcomes for B recipients of B kidneys, 92.6%, 87.9%, and 82.5%, respectively (p-value = 0.48). Five DSAs increased the proportion of B transplants during 41 months postvariance, with a lesser proportional decrease in blood group A transplants. The data support the proposition that this allocation algorithm may provide a robust mechanism to increase access of blood group B minority candidates to kidney transplantation.

Immunological comparison of DNA vaccination using two delivery systems against canine leishmaniasis

Visceral leishmaniasis (VL) is a fatal disease caused by the intracellular protozoan parasite Leishmania infantum. Dogs are the primary reservoirs of this parasite, and vaccination of dogs could be an effective method to reduce its transfer to humans. In order to develop a vaccine against VL (apart from the choice of immunogenic candidate antigens), it is necessary to use an appropriate delivery system to promote a proper antigen-specific immune response. In this study, we compared two vaccine delivery systems, namely electroporation and cationic solid–lipid nanoparticle (cSLN) formulation, to administer a DNA vaccine containing the Leishmania donovani A2 antigen, and L. infantum cysteine proteinases of type I (CPA) and II (CPB) without its unusual C-terminal extension. The protective potencies of these two vaccine delivery systems were evaluated against L. infantum challenge in outbred dogs. Our results show that the administration of pcDNA-A2-CPA-CPB−CTEGFP vaccine as a prime-boost by either electroporation or cSLN formulation protects the dogs against L. infantum infection. Partial protection in vaccinated dogs is associated with significantly (p<0.05) higher levels of IgG2, IFN-γ, and TNF-α and with low levels of IgG1 and IL-10 as compared to the control group. Protection was also correlated with a low parasite burden and a strong delayed-type hypersensitivity (DTH) response. This study demonstrates that both electroporation and cSLN formulation can be used as efficient vaccine delivery systems against visceral leishmaniasis.

Evaluation of Live Recombinant Nonpathogenic Leishmania tarentolae Expressing Cysteine Proteinase and A2 Genes as a Candidate Vaccine against Experimental Canine Visceral Leishmaniasis.

Canine Visceral Leishmaniasis (CVL) is a major veterinary and public health problem caused by Leishmania infantum (L. infantum) in many endemic countries. It is a severe chronic disease with generalized parasite spread to the reticuloendothelial system, such as spleen, liver and bone marrow and is often fatal when left untreated. Control of VL in dogs would dramatically decrease infection pressure of L. infantum for humans, since dogs are the main domestic reservoir. In the past decade, various subunits and DNA antigens have been identified as potential vaccine candidates in experimental animal models, but none has been approved for human use so far. In this study, we vaccinated outbreed dogs with a prime-boost regimen based on recombinant L. tarentolae expressing the L. donovani A2 antigen along with cysteine proteinase genes (CPA and CPB without its unusual C-terminal extension (CPB-CTE) and evaluated its immunogenicity and protective immunity against L. infantum infectious challenge. We showed that vaccinated animals produced significantly higher levels of IgG2, but not IgG1, and also IFN-γ and TNF-α, but low IL-10 levels, before and after challenge as compared to control animals. Protection in dogs was also correlated with a strong DTH response and low parasite burden in the vaccinated group. Altogether, immunization with recombinant L. tarentolae A2-CPA-CPB-CTE was proven to be immunogenic and induced partial protection in dogs, hence representing a promising live vaccine candidate against CVL.

Canine Leishmania vaccines: Still a long way to go

Dogs are the main reservoir host for zoonotic visceral leishmaniasis, a sand fly-borne disease caused by Leishmania infantum. In endemic areas, “susceptible” dogs suffer from a severe disease characterized by chronic polymorphic viscerocutaneous signs that manifest several months from the exposure, whereas “resistant” dogs can remain subclinically infected for years or lifelong. The protective immune response to Leishmania is cell-mediated; for visceralizing Leishmania species a mixed T helper (Th)1/Th2 response with a dominant Th1 profile is required for protection. The activation of the adaptive immune system in naturally resistant dogs is revealed by parasite-specific lymphoproliferation, delayed-type hypersensitivity, the production of interferon-γ and tumour necrosis factor-α cytokines, and enhanced macrophage leishmanicidal activity via nitric oxide. Hence, an effective canine Leishmania vaccine should induce strong and long-lasting Th1-dominated immunity to control both infection progression and the parasite transmissibility via the vector. Preclinical research in rodent models has evaluated the efficacy of several categories of Leishmania antigens including killed parasites, cell purified fractions, parasite protein components or subunits, single or multiple chimeric recombinant proteins, plasmid DNA and viral particles encoding parasite virulence factors. Promising antigen(s)/adjuvant combinations from each of the above categories have also been tested in dogs; they mostly resulted in limited or no protection in Phase I–II studies (designed to test vaccine safety, immunogenicity and laboratory-induced protection) in which vaccinated dogs were challenged by the artificial intravenous injection of high-load L. infantum promastigotes. The recombinant A2 antigen plus saponin conferred about 40% protection against infection by this challenge system and has been registered in Brazil as a canine vaccine (LeishTec®). An increasing number of efficacy studies have privileged the use of natural challenge consisting in the long-term exposure of vaccinated dogs in endemic settings (Phase III). A 2-year field model including regular assessments by a set of standard diagnostic markers useful for an accurate infection staging has been developed. Again, most of the vaccines tested by this system, which included several antigen categories and adjuvants, failed to protect against infection and disease. Only two vaccines, consisting of parasite purified fractions with saponin derivative adjuvants, showed to confer significant protection against disease and death under natural conditions, and have been registered as canine vaccines: FML-QuilA (Leishmune®) in Brazil, and LiESP/QA-21 (CaniLeish®) in Europe.

Experimental prestorage filtration removes antibodies and decreases lipids in RBC supernatants mitigating TRALI in vivo

AbstractTransfusion-related acute lung injury (TRALI) remains a significant cause of transfusion-related mortality with red cell transfusion. We hypothesize that prestorage filtration may reduce proinflammatory activity in the red blood cell (RBC) supernatant and prevent TRALI. Filters were manufactured for both small volumes and RBC units. Plasma containing antibodies to human lymphocyte antigen (HLA)-A2 or human neutrophil antigen (HNA)-3a was filtered, and immunoglobulins and specific HNA-3a and HLA-2a neutrophil (PMN) priming activity were measured. Antibodies to OX27 were added to plasma, and filtration was evaluated in a 2-event animal model of TRALI. RBC units from 31 donors known to have antibodies against HLA antigens and from 16 antibody-negative controls were filtered. Furthermore, 4 RBC units were drawn and underwent standard leukoreduction. Immunoglobulins, HLA antibodies, PMN priming activity, and the ability to induce TRALI in an animal model were measured. Small-volume filtration of plasma removed >96% of IgG, antibodies to HLA-A2 and HNA-3a, and their respective priming activity, as well as mitigating antibody-mediated in vivo TRALI. In RBC units, experimental filtration removed antibodies to HLA antigens and inhibited the accumulation of lipid priming activity and lipid-mediated TRALI. We conclude that filtration removes proinflammatory activity and the ability to induce TRALI from RBCs and may represent a TRALI mitigation step.

Cationic solid–lipid nanoparticles are as efficient as electroporation in DNA vaccination against visceral leishmaniasis in mice

The use of an appropriate delivery system has recently emerged as a promising approach for the development of effective vaccination against visceral leishmaniasis (VL). Here, we compare two vaccine delivery systems, namely electroporation and cationic solid-lipid nanoparticle (cSLN) formulation, to administer a DNA vaccine harbouring the L. donovani A2 antigen along with L. infantum cysteine proteinases [CPA and CPB without its unusual C-terminal extension (CPB(-CTE) )] and evaluate their potential against L. infantum challenge. Prime-boost administration of the pcDNA-A2-CPA-CPB(-CTE) delivered by either electroporation or cSLN formulation protects BALB/c mice against L. infantum challenge and that protective immunity is associated with high levels of IFN-γ and lower levels of IL-10 production, leading to a strong Th1 immune response. At all time points, the ratio of IFN-γ: IL-10 induced upon restimulation with rA2-rCPA-rCPB and F/T antigens was significantly higher in vaccinated animals. Moreover, Th2-efficient protection was elicited through a high humoral immune response. Nitric oxide production, parasite burden and histopathological analysis were also in concordance with other findings. Overall, these data indicate that similar to the electroporation delivery system, cSLNs as a nanoscale vehicle of Leishmania antigens could improve immune response, hence indicating the promise of these strategies against visceral leishmaniasis.

Development of Novel Prime-Boost Strategies Based on a Tri-Gene Fusion Recombinant L. tarentolae Vaccine against Experimental Murine Visceral Leishmaniasis

Visceral leishmaniasis (VL) is a vector-borne disease affecting humans and domestic animals that constitutes a serious public health problem in many countries. Although many antigens have been examined so far as protein- or DNA-based vaccines, none of them conferred complete long-term protection. The use of the lizard non-pathogenic to humans Leishmania (L.) tarentolae species as a live vaccine vector to deliver specific Leishmania antigens is a recent approach that needs to be explored further. In this study, we evaluated the effectiveness of live vaccination in protecting BALB/c mice against L. infantum infection using prime-boost regimens, namely Live/Live and DNA/Live. As a live vaccine, we used recombinant L. tarentolae expressing the L. donovani A2 antigen along with cysteine proteinases (CPA and CPB without its unusual C-terminal extension (CPB-CTE)) as a tri-fusion gene. For DNA priming, the tri-fusion gene was encoded in pcDNA formulated with cationic solid lipid nanoparticles (cSLN) acting as an adjuvant. At different time points post-challenge, parasite burden and histopathological changes as well as humoral and cellular immune responses were assessed. Our results showed that immunization with both prime-boost A2-CPA-CPB-CTE-recombinant L. tarentolae protects BALB/c mice against L. infantum challenge. This protective immunity is associated with a Th1-type immune response due to high levels of IFN-γ production prior and after challenge and with lower levels of IL-10 production after challenge, leading to a significantly higher IFN-γ/IL-10 ratio compared to the control groups. Moreover, this immunization elicited high IgG1 and IgG2a humoral immune responses. Protection in mice was also correlated with a high nitric oxide production and low parasite burden. Altogether, these results indicate the promise of the A2-CPA-CPB-CTE-recombinant L. tarentolae as a safe live vaccine candidate against VL.

Rapid detection of human and canine visceral leishmaniasis: Assessment of a latex agglutination test based on the A2 antigen from amastigote forms of Leishmania infantum

The diagnosis of visceral leishmaniasis (VL) in humans and animal reservoir hosts is difficult, particularly in rural areas where the disease is endemic and laboratory facilities are limited. This study aimed to develop a latex agglutination test (LAT) for the rapid detection of anti-Leishmania antibodies against the A2 antigen derived from the amastigote form as well as those against crude antigens derived from the promastigote form of an Iranian strain of Leishmania (Leishmania) infantum. The A2 antigen (42-100 kDa) was prepared from the amastigote form of L. infantum, purified with electroelution and compared with the crude antigen from the promastigote form of L. infantum. Both antigens showed appropriate intensity reactions, were selected using dot blotting of positive and negative pooled sera and used to sensitize 0.9-μm latex beads. The tests were carried out on sera from 43 symptomatic, human patients with VL confirmed by parasitological examination and direct agglutination test (DAT), 30 healthy controls and 32 patients with other infections but without VL. Canine sera were collected from 63 domestic dogs with VL confirmed using parasitological examinations and DAT and 31 healthy dogs from areas non-endemic for VL. Compared with the controls, human sera from DAT-confirmed patients yielded a sensitivity of 88.4% (95% CI, 82.1-94.5%) and specificity of 93.5% (95% CI, 87.0-99.7%) on A2-LAT (amastigote) when 1:3200 was used as the cut-off titre. A good degree of agreement was found between A2-LAT and DAT (0.914). LAT required 3-5 min to complete, versus the 12-18 h needed for DAT. Compared with the controls, A2-LAT of canine sera from DAT-confirmed cases yielded a sensitivity of 95.2% (95% CI, 95.0-95.4%) and specificity of 100% (95% CI 100%) when 1:320 was used as the cut-off titre. A good degree of agreement was found between A2-LAT and DAT (0.968). Similarly, the sensitivity and specificity of Pro.-LAT (promastigote) was calculated to be 88.4% and 91.9%, respectively for human sera and 96.8% and 90.3%, respectively for canine sera. No statistically significant differences were observed between A2-LAT and Pro.-LAT for the detection of human and canine L. infantum infections. In conclusion, A2-LAT and Pro.-LAT could be used in parallel to screen for L. infantum infections in humans and dogs in areas endemic for VL in Iran.

A Novel Concept of Phased Desensitization Using Bortezomib and Rituximab for Highly HLA-Sensitized Kidney Transplantation

Introduction: Donor specific alloantibody producing plasma cells (DSA-PCs) appear resistant to conventional immunosuppressive agents. The recent studies demonstrated the impact of pretransplant desensitization with the proteasome inhibitor bortezomib on DSA-PCs in sensitized renal allograft candidates. However, other studies have showed that one cycle of bortezomib alone does not decrease DSA levels in sensitized kidney transplant recipients. We developed a concept of phased desensitization with rituximab and bortezomib, and applied it in to a highly sensitized kidney transplant candidate. Methods: We assessed the activity of bortezomib against various B cell subpopulations and plasma cells and suppressive effect of that to produce anti-blood group antibodies (Abs) DSA in mice. To elicit Abs against allopeptide (DSA), B6 mice were immunized with thymocytes from Balb/c mice. After the immunization, the mice received intravenous injections of bortezomib (0.75 mg/kg body weight) 3 times a week for 1 week. On the basis of the results, we have established a novel concept of desensitization for a sensitized patient. Results: Bortezomib led to a significant reduction in the subpopulations of CD21-/IgMhigh plasmablasts, and CD138+/CD25-/IgG κ-light chain+ plasma cells in the spleen of Balb/c mice, but not in that of CD21+/IgMhigh marginal-zone B cells and the CD21low/IgMlow follicular B cells. Flow cytometry analyses revealed that bortezomib did not reduce the B cells with receptors for either A-carbohydrates. Thus, bortezomib reduced not only IgG producing plasma cells but also IgM producing cells, although their precursor memory B cells were resistant to this reagent. Taken together with a known fact that rituximab is able to deplete memory B cells, we have proposed a concept of phased desensitization using bortezomib and rituximab, and applied it in to a highly sensitized kidney transplant patient. A 56-year-old man with renal graft failure was referred to our hospital. His first renal graft from his brother remained functioning for 16 years but was lost due to chronic rejection. This allograft loss resulted in sensitization of the recipient against HLA antigens. Unfortunately, his wife who volunteered as a second donor shared the same A2 antigen with the first donor, which might be a target of the preformed anti-HLA Abs. Single antigen flow-based DSA testing revealed 11 anti-HLA class I Abs including anti-donor HLA A2 Ab. He received a desensitization regimen consisting of a single dose of rituximab (375 mg/m2) combined with double-filtration plasmapheresis (DFPP) followed by low doses of intravenous immuno-globulin (DFPP/IVIG treatment). Tacrolimus and mycophenolate mofetil were started 2 weeks before the DFPP/IVIG treatment. Despite 3 DFPP/IVIG sessions, the crossmatch remained positive. Thereafter, he was treated with one cycle of bortezomib (1.3mg/m2 × 4 doses). After 3 DFPP/IVIG sessions, the crossmatch turned negative and renal transplantation was successfully performed. The induction quadruple immunosuppression protocol comprised tacrolimus, mycophenolate mofetil and methylprednisolone. After the transplantation, neither Ab-mediated rejection nor severe acute cellular rejection was encountered in this patient. Conclusions: The current study suggests that phased desensitization with rituximab and bortezomib may be needed for highly HLA-sensitized kidney transplant candidates.

Serological characteristic and molecular basis of A2 subgroup in the Chinese population

BackgroundA2 phenotype is a common subgroup of blood group A, but the serological characteristic and genetics basis of A2 phenotype currently was rare reported in the Chinese Han population. Here, a large scale study of the serology and genetics of A2 and A2B phenotypes was performed. Methods/materials11263 Chinese individuals with group A and AB phenotypes were determined for A2 antigen with the standard serological method. The full coding region of the ABO gene was sequenced in the individuals with A2 and A2B phenotypes. Some samples including each ABO genotypes were chosen for determining the activity of glycosyltransferase A (GTA) in plasma. Results134 individuals were assigned as A2 and A2B phenotypes in 11263 individuals. There was imbalance in A2 and A2B phenotypes and the proportion of A2B among AB samples was significantly higher than that of A2 in group A samples. All samples of the A2 and A2B phenotypes were classified into A2-related allele group, A1-related allele group and the other group based on kind of the ABO genotype. Four novel A2-related alleles (A217, A218, A219, A220) were identified. The individuals with same genotype showed different agglutination strength with anti-A1 and anti-H on their RBCs. The plasma from individuals with A2-related allele had almost no GTA activity, while plasma from individuals with A1-related allele had some GTA activity. ConclusionA2 and A2B phenotypes could derive from different genotypes and the serological characteristic may be heterogeneity in the Chinese Han population.

Generation and evaluation of A2-expressing Lactococcus lactis live vaccines against Leishmania donovani in BALB/c mice

Leishmaniasis is a parasitic disease affecting over 12 million individuals worldwide. As current treatments are insufficient, the development of an effective vaccine is a priority. This study generated and assessed the efficacy of Leishmania vaccines engineered from the non-colonizing, non-pathogenic Gram-positive bacterium Lactococcus lactis. A truncated, codon-optimized version of the A2 antigen from Leishmania donovani was engineered for expression in Lactococcus lactis in three different subcellular compartments: in the cytoplasm, secreted outside the cell or anchored to the cell wall. These three A2-expressing Lactococcus lactis strains were tested for their ability to generate A2-specific immune responses and as live vaccines against visceral Leishmania donovani infection in BALB/c mice. Subcutaneous immunization with live Lactococcus lactis expressing A2 anchored to the cell wall effectively induced high levels of antigen-specific serum antibodies. It was demonstrated that Lactococcus lactis-based vaccines are a feasible approach in the generation of live vaccines against leishmaniasis. The Lactococcus lactis strains generated in this study provide an excellent foundation for further studies on live bacterial vaccines against leishmaniasis and other pathogens.

Recombinant Leishmania tarentolae expressing the A2 virulence gene as a novel candidate vaccine against visceral leishmaniasis

Visceral leishmaniasis is the most severe form of leishmaniasis. To date, there is no effective vaccine against this disease. Many antigens have been examined so far as protein- or DNA-based vaccines, but none of them conferred complete long-term protection. The use of live attenuated vaccines has recently emerged as a promising vaccination strategy. In this study, we stably expressed the Leishmania donovani A2 antigen in Leishmania tarentolae, a non-pathogenic member of the genus Leishmania, and evaluated its protective efficacy as a live vaccine against L. infantum challenge. Our results show that a single intraperitoneal administration of the A2-recombinant L. tarentolae strain protects BALB/c mice against L. infantum challenge and that protective immunity is associated with high levels of IFN-γ production prior and after challenge. This is accompanied by reduced levels of IL-5 production after challenge, leading to a potent Th1 immune response. In contrast, intravenous injection elicited a Th2 type response, characterized by higher levels of IL-5 and high humoral immune response, resulting in a less efficient protection. All together, these results indicate the promise of A2-expressing L. tarentolae as a safe live vaccine against visceral leishmaniasis.

Epitope mapping and protective immunity elicited by adenovirus expressing the Leishmania amastigote specific A2 antigen: Correlation with IFN-γ and cytolytic activity by CD8+ T cells

A2 was identified as an amastigote virulence factor of Leishmania ( Leishmania) donovani and as a candidate antigen for vaccine development against visceral leishmaniasis. Here, predicted hydrophilic, class I and II MHC-binding synthetic peptides were used to define epitopes recognized by A2-specific antibodies, CD8+ T and CD4+ T cells, respectively. Immunization of BALB/c mice with adenovirus expressing A2 (AdA2) resulted in low antibody response, contrasting with high levels of IFN-γ producing CD4+ T and CD8+ T cells specific for A2. Further, A2-specific CD8+ T cells from immunized mice were capable of lysing sensitized target cells in vivo. Finally, we demonstrated an association of A2-specific T cell responses and reduced parasitism in both liver and spleen from mice immunized with AdA2 and challenged with L. ( L.) chagasi.

Hyperacute Rejection After Single Lung Transplantation: A Case Report

Hyperacute rejection is a well-known complication in kidney and heart transplantations. However, its occurrence in lung transplantation is extremely rare, with only 4 cases previously described. A 53-year-old female patient blood type O with end-stage chronic obstructive pulmonary disease underwent left lung transplantation. She had 2 negative pretransplantation evaluations for panel-reactive antibodies. One hour after the vascular clamps were released, progressive hypoxia developed. Fiberoptic bronchoscopy revealed an optimal bronchial anastomosis; an abundant pink frothy fluid was observed on the allograft side. Chest X ray sevealed a completely opacified left lung. Due to the low-compliance of the transplanted lung and the risk for native lung hyperinsufflation, independent mechanical ventilation was employed. Despite all measures, multiple organ failure developed and the patient died 24 hours after the procedure. A necropsy evaluation for confirmed the patency of all anastomoses and no signs of ischemia. Retrospectively, a new evaluation for panel-reactive antibodies was performed, with 24% reactivity. Complement-dependent cytotoxicity crossmatch was negative, however, a flow cytometric analysis was positive for both HLA-I (56%) and HLA-II (45%). Further investigation detected an anti-A2 in the recipient serum and the donor had an A2 antigen. Hyperacute rejection is a rare posttransplantation complication highlighted by its precocity and lethality. With the increased number of lung transplantations performed yearly, it is believed that its incidence will also rise. Therefore, prompt diagnosis and familiarity with management strategies are fundamental.

Skin biopsies in DC vaccines for stage III–IV melanoma patients: Role of neutrophils?

Dendritic cell (DC) vaccines are used for the induction of anti-tumor T cell reaction in melanoma patients. DC are generated in vitro, pulsed with antigen and matured prior to injection. They are supposed to migrate to lymph nodes and to present the processed antigen to naive T cells allowing activation of tumor-specific lymphocytes. It has been suggested that intradermal injection allows a superior migration to the lymph node. Eight HLA-A2 positive patients with stage III or IV melanomas expressing NA 17 antigen were collected. They were included in a pilot trial of vaccination in which they received IL3/INFb DC presenting the NA17 A2 antigen. In each patient, a skin biopsy was performed at the injection site, 24 h after inoculation. The striking features of the biopsies were the presence of a perivascular CD3+/CD8+ T cell infiltrate with a slight population of CD4+ cells and the presence of a massive neutrophilic infiltrate associated with the injected DC still present, realizing a suppurative granuloma. The persistence of DC 24 h after the injection suggests that migration in the lymph node is not necessary for the induction of the immune response. The skin itself could be the location of a reaction starting with a massive recruitment of neutrophils.