Ground-Based Mobile Measurements to Track Urban Methane Emissions from Natural Gas in 12 Cities across Eight Countries.

To mitigate methane emission from urban natural gas distribution systems, it is crucial to understand local leak rates and occurrence rates. To explore urban methane emissions in cities outside the U.S., where significant emissions were found previously, mobile measurements were performed in 12 cities across eight countries. The surveyed cities range from medium size, like Groningen, NL, to large size, like Toronto, CA, and London, UK. Furthermore, this survey spanned across European regions from Barcelona, ES, to Bucharest, RO. The joint analysis of all data allows us to focus on general emission behavior for cities with different infrastructure and environmental conditions. We find that all cities have a spectrum of small, medium, and large methane sources in their domain. The emission rates found follow a heavy-tailed distribution, and the top 10% of emitters account for 60-80% of total emissions, which implies that strategic repair planning could help reduce emissions quickly. Furthermore, we compare our findings with inventory estimates for urban natural gas-related methane emissions from this sector in Europe. While cities with larger reported emissions were found to generally also have larger observed emissions, we find clear discrepancies between observation-based and inventory-based emission estimates for our 12 cities.

Comparative validation of computer programs for haplotype frequency estimation from donor registry data.

Estimation of human leukocyte antigen (HLA) haplotype frequencies from unrelated stem cell donor registries presents a challenge because of large sample sizes and heterogeneity of HLA typing data. For the 14th International HLA and Immunogenetics Workshop, five bioinformatics groups initiated the 'Registry Diversity Component' aiming to cross-validate and improve current haplotype estimation tools. Five datasets were derived from different donor registries and then used as input for five different computer programs for haplotype frequency estimation. Because of issues related to heterogeneity and complexity of HLA typing data identified in the initial phase, the same five implementations, and two new ones, were used on simulated datasets in a controlled experiment where the correct results were known a priori. These datasets contained various fractions of missing HLA-DR modeled after European haplotype frequencies. We measured the contribution of sampling fluctuation and estimation error to the deviation of the frequencies from their true values, finding equivalent contributions of each for the chosen samples. Because of patient-directed activities, selective prospective typing strategies and the variety and evolution of typing technology, some donors have more complete and better HLA data. In this setting, we show that restricting estimation to fully typed individuals introduces biases that could be overcome by including all donors in frequency estimation. Our study underlines the importance of critical review and validation of tools in registry-related activity and provides a sustainable framework for validating the computational tools used. Accurate frequencies are essential for match prediction to improve registry operations and to help more patients identify suitably matched donors.

16(th) IHIW: global analysis of registry HLA haplotypes from 20 million individuals: report from the IHIW Registry Diversity Group.

This project has the goal to validate bioinformatics methods and tools for HLA haplotype frequency analysis specifically addressing unique issues of haematopoietic stem cell registry data sets. In addition to generating new methods and tools for the analysis of registry data sets, the intent is to produce a comprehensive analysis of HLA data from 20 million donors from the Bone Marrow Donors Worldwide (BMDW) database. This report summarizes the activity on this project as of the 16IHIW meeting in Liverpool.

Thirty-six novel HLA alleles: 7 HLA-A, 11 HLA-B, 15 HLA-C and 3 HLA-DRB1.

Thirty-six novel human leukocyte antigen (HLA) alleles are described in this article: A*9225N, A*9234, A*030106, A*0337, A*2317, A*2480, A*3023; B*070206, B*0759, B*0761, B*0765, B*150106, B*1827, B*352002, B*3585, B*3943, B*4082, B*5151; Cw*0342, Cw*0343, Cw*0344, Cw*0428, Cw*0430, Cw*0433, Cw*050104, Cw*0519, Cw*060203, Cw*070109, Cw*070202, Cw*0750, Cw*0815, Cw*120306, Cw*1409; DRB1*0336, DRB1*0473 and DRB1*1382.

Eleven million donors in Bone Marrow Donors Worldwide! Time for reassessment?

On 16 November 2005, we celebrated the milestone when 10 million donors had been registered in Bone Marrow Donors Worldwide (BMDW). Since then another million donors have been added in little more than a year. It seems an appropriate time for reassessment and to ask whether we are on the right track or not. We will do so by discussing the strength, weaknesses, opportunities and threats of the unrelated stem cell donor operation.

World Marrow Donor Association guidelines for use of HLA nomenclature and its validation in the data exchange among hematopoietic stem cell donor registries and cord blood banks.

Since the advent of the European Marrow Donor Information System in the first half of the last decade, fully automated data exchange between registry computer systems has been playing an ever-increasing role in the international search for unrelated donors of blood progenitor cells. This exchange, however, was hampered by different local conventions used to present HLA data and complicated by the need to extend the official WHO nomenclature to accommodate the registries' information systems and to cross-validate HLA data obtained with different methods and/or at different loci. The guidelines presented here have been developed by the World Marrow Donor Association to standardize the nomenclature to be used and the validation checks to be applied in the international electronic exchange of HLA-typing data among unrelated volunteer hematopoietic stem cell donor registries and umbilical cord blood banks. Two reference web sites have been designated to maintain and update the approved HLA nomenclature and all the ancillary information needed by the conventions described here.

Highly diverged MHC class I mismatches are acceptable for haematopoietic stem cell transplantation.

A fully major histocompatilbility complex (MHC) matched donor is not available for the majority of patients in need of a haematopoietic stem cell transplantation (SCT), which illustrates the need for a tool to define acceptable MHC disparities. Previously, we noticed that a variety of single MHC class I mismatched allogeneic donor-recipient pairs did not elicit an allogeneic cytotoxic-lymphocyte (CTL) response in vitro if the MHC amino-acid sequences had five or more differences in the alpha-helices plus five or more differences in the beta-sheet (> or =5alpha5beta) (7). To address the clinical relevance of this observation, we analysed CTL precursor (CTLp) assay outcome and SCT outcome in 53 Dutch recipients of a single MHC class I mismatched graft from an unrelated donor. Overall patient survival was 44% after 4 years. In multivariate analysis, recipients of a > or =5alpha5beta mismatched graft with negative CTLp frequencies in vitro before transplantation demonstrated superior survival: survival at 4 years was 80% as compared to 47% in recipients of other mismatched grafts with negative CTLp frequencies (hazard ratio=0.131; 95% CI=(0.03-0.61); P=0.009). This option of acceptable mismatches may enlarge the pool of potentially acceptable stem cell donors.

Hematopoietic stem cell donor selection: the Europdonor experience.

International collaboration is essential for the optimal selection of unrelated hematopoietic stem cell donors. This review focuses on the benefit of a joint worldwide donor file called Bone Marrow Donors Worldwide and the experience of the Europdonor Foundation in selecting strategies to identify the best human leukocyte antigen-matched donor in the shortest time.

Glutathione-dependent biotransformation of the alkylating drug thiotepa and transport of its metabolite monoglutathionylthiotepa in human MCF-7 breast cancer cells.

In this study, the role of glutathione S-transferase (GST) P1-1, the cellular reduced glutathione (GSH) status, and ATP-dependent efflux pumps in the cellular glutathione-dependent biotransformation of thiotepa and transport of the main metabolite monoglutathionylthiotepa in relation to cytotoxicity was studied in control and GST-P1-1-transfected MCF-7 cell lines. It was demonstrated that an enhanced cellular level of GST-P1-1 leads to an enhanced formation of monoglutathionylthiotepa, which is transported out of the cell into the medium. Monoglutathionylthiotepa was able to reversibly inhibit the activity of purified GST-P1-1, but only at nonphysiological concentrations, indicating that feedback inhibition of GST by its metabolites is not a relevant process in vivo. The GST activity, cellular GSH level, and/or ATP-dependent efflux of monoglutathionylthiotepa were modulated using ethacrynic acid, D,L-buthionine-S,R-sulfoximine, probenecid, and verapamil to understand the interplay between GSTs, glutathione conjugation, and efflux of glutathione conjugates in more detail. Inhibition of the GSH biosynthesis by D,L-buthionine-R,S-sulfoximine, a specific inhibitor of gamma-glutamylcysteine synthetase, significantly reduced the glutathione conjugation of thiotepa and potentiated the cytotoxicity of thiotepa. Pretreatment of cells with ethacrynic acid resulted in decreased formation of monoglutathionylthiotepa as a result of inhibition of GST in the GST-P1-1 transfectant. In addition, the intracellular amount of monoglutathionylthiotepa increased in both of the cell lines on exposure to ethacrynic acid, indicating that transport of the glutathione conjugate was partially inhibited by the glutathione conjugate of ethacrynic acid. Transport activity of monoglutathionylthiotepa could also be inhibited by probenecid and verapamil, inhibitors of organic anion transport, without influencing the biotransformation capacity of the cells. It was demonstrated that inhibition of glutathione conjugate efflux by probenecid and verapamil leads to enhanced cytotoxicity, which indicates that besides thiotepa, monoglutathionylthiotepa is also cytotoxic for the cells. Only enhanced biotransformation and subsequent transport of the glutathione conjugate into the medium (which occurs with the GST-P1-1 transfectant) results in enhanced viability. Therefore, it was concluded that only enhanced biotransformation of thiotepa represents a real detoxification pathway when the resulting conjugate is transported out of the cells. Altogether, the results indicate that it is not the overexpression of GST per se but the interplay between GSH/GST and glutathione conjugate efflux pumps that results in increased resistance to alkylating anticancer drugs such as thiotepa.

A comparative reference study for the validation of HLA-matching algorithms in the search for allogeneic hematopoietic stem cell donors and cord blood units.

The accuracy of human leukocyte antigen (HLA)-matching algorithms is a prerequisite for the correct and efficient identification of optimal unrelated donors for patients requiring hematopoietic stem cell transplantation. The goal of this World Marrow Donor Association study was to validate established matching algorithms from different international donor registries by challenging them with simulated input data and subsequently comparing the output. This experiment addressed three specific aspects of HLA matching using different data sets for tasks of increasing complexity. The first two tasks targeted the traditional matching approach identifying discrepancies between patient and donor HLA genotypes by counting antigen and allele differences. Contemporary matching procedures predicting the probability for HLA identity using haplotype frequencies were addressed by the third task. In each task, the identified disparities between the results of the participating computer programs were analyzed, classified and quantified. This study led to a deep understanding of the algorithms participating and finally produced virtually identical results. The unresolved discrepancies total to less than 1%, 4% and 2% for the three tasks and are mostly because of individual decisions in the design of the programs. Based on these findings, reference results for the three input data sets were compiled that can be used to validate future matching algorithms and thus improve the quality of the global donor search process.

The importance of identifying a back-up donor for unrelated stem cell transplantation.

The importance of identifying a back-up donor, once a primary suitable unrelated stem cell donor has been found, is often underestimated. Transplant centres erroneously count on the unrelated volunteer donors to be willing, available and medically fit for actual donation. According to our data, which includes 502 unrelated donor work-up procedures performed for 425 Dutch patients between 1987 and 2002, one of 11 work-ups ended in the primary requested donor failing to donate. Of all donor-related cancellations (N=46), 78% of the procedures were deferred due to medical reasons and 22% due to nonmedical reasons. Most of the donors deferred for medical reasons were female (P=0.005). In 50% of the cases for which a back-up donor was already identified, the patients were transplanted with a delay of less then 2 weeks; when no back-up donor was available, the median delay increased to 18 weeks. We strongly encourage implementing a search for at least one back-up donor in the primary search. Identifying a back-up donor can save precious time and complicated logistic rescheduling.

How to improve the search for an unrelated haematopoietic stem cell donor. Faster is better than more!

Many patients do not reach haematopoietic stem cell transplantation. Shortage of unrelated donors (UDs) is still seen as the main cause. However, with a worldwide UD pool containing more than 8 million donors, it is possible that other impediments are becoming more important. We analysed 549 UD searches for Dutch patients, performed between 1987 and 2000, in order to find the reasons for failure or success to reach transplantation. Between 1996 and 2000, 59% of the patients of Northwest European origin received a graft from an UD with a median time span of 4.4 months from the start of the search. In all, 11% of the patients lacked a compatible donor, while 30% became medically unfit for transplantation. This is in contrast to the patients of non-Northwest European origin for whom UD shortage is still the most important impediment; only 32% were transplanted while 50% lacked a compatible donor. We conclude that the shortage of donors is no longer the biggest constraint in unrelated stem cell transplantation for patients of Northwest European origin. It may be more effective to optimize the chance on transplantation by making the search process more efficient.

HLA-DR4 is associated with a diminished risk of the development of chronic myeloid leukemia (CML). Chronic Leukemia Working Party of the European Blood and Marrow Transplant Registry.

CML is characterized by the chromosomal translocation t(9;22) (q34;q11) resulting in the chimeric bcr-abl oncogene that encodes P210 fusion proteins with novel amino acid sequences in the breakpoint region. If these peptides derived from P210 are presented by HLA molecules on the cell membrane of leukemic cells an immunological response may occur. Recent studies using synthetic peptides identical to the bcr-abl fusion region revealed that some peptides are capable of binding to the class I molecules HLA-A2,-A3,-A11 and -B8 and the class II molecules HLA-DR1, -DR2, -DR3, -DR4 and -DR11. Moreover T cell responses have been induced against bcr-abl-derived synthetic peptides bound to some of these HLA molecules. For HLA class I, we have previously shown that individuals expressing HLA-A3 and -B8 have a diminished risk of development of CML. To assess a similar protective effect of class II molecules we performed a large multi-center study. This study compared the frequencies of HLA-DR1, -DR2, -DR3, -DR4 and -DR11 of patients with CML from the database of the EBMT (n = 1462) with unaffected individuals from the registry of Bone Marrow Donors Worldwide (n = 500 596). Patients and controls were matched per country. This analysis yielded significantly lower odds ratios (ORs) of 0.86 (95% CI 0.75-0.98) for HLA-DR3 and of 0.80 (95% CI 0.71-0.89) for HLA-DR4. The OR was 0.91 (95% CI 0.80-1.04) for HLA-DR1, 1.05 (95% CI 0.94-1.18) for HLA-DR2 and 0.87 (95% CI 0.74-1.01) for HLA-DR11. To assess a possible effect of the linkage disequilibrium between HLA-B8 and HLA-DR3 we found that coexpression of HLA-B8 and HLA-DR3 gave an OR of 0.84 (95% CI 0.72-0.98), whereas HLA-DR3 positive/HLA-B8 negative individuals showed an OR of 1.02 (95% CI 0.84-1.24). This means that the protective effect of HLA-DR3 of the development of CML was probably caused by its linkage disequilibrium with HLA-B8. In contrast, as there is no linkage disequilibrium of HLA-DR4 with HLA-A3 or HLA-B8, the results indicate that HLA-DR4 expression itself is associated with a diminished incidence of CML possibly by the presentation of bcr-abl breakpoint peptides in these HLA molecules on the membrane of the leukemic cells.

Physiochemical disparity of mismatched HLA class I alloantigens and risk of acute GVHD following HSCT.

We determined whether assessment of the immunogenicity of individual donor-recipient HLA mismatches based on differences in their amino-acid sequence and physiochemical properties predicts clinical outcome following haematopoietic SCT (HSCT). We examined patients transplanted with 9/10 single HLA class I-mismatched grafts (n=171) and 10/10 HLA-A-, -B-, -C-, -DRB1- and -DQB1-matched grafts (n=168). A computer algorithm was used to determine the physiochemical disparity (electrostatic mismatch score (EMS) and hydrophobic mismatch score (HMS)) of mismatched HLA class I specificities in the graft-versus-host direction. Patients transplanted with HLA-mismatched grafts with high EMS/HMS had increased incidence of ⩾grade II acute GVHD (aGVHD) compared with patients transplanted with low EMS/HMS grafts; patients transplanted with low and medium EMS/HMS grafts had similar incidence of aGVHD to patients transplanted with 10/10 HLA-matched grafts. Mortality was higher following single HLA-mismatched HSCT but was not correlated with HLA physiochemical disparity. Assessment of donor-recipient HLA incompatibility based on physiochemical HLA disparity may enable better selection of HLA-mismatched donors in HSCT.

Cross-reactive group matching does not lead to a better allocation and survival of donor kidneys.

BACKGROUND: In cadaveric renal transplantation HLA-A, -B, -DR matching of donor and recipient is beneficial for graft survival. However, allocation based on HLA matching seems to favor recipients with more frequently occurring HLA antigens. In this study we investigated whether matching on the basis of cross-reactive groups (CREGs), defined according to the United Network for Organ Sharing (UNOS), would be a good alternative for the allocation of kidneys without negatively influencing graft survival. Theoretically, this approach would provide more recipients with an immunologically well-matched donor organ. METHODS: The influence of CREG matching on graft survival was studied in univariate analyses using the Eurotransplant database. RESULTS: No beneficial effect of CREG matching was observed, whereas a significant HLA matching effect was observed in the 0 CREG mismatched donor/ recipient combinations. Only in the small subgroup with 1 MM for HLA-A, -B and 0 MM for HLA-DR, a significantly better survival was observed, when this mismatch belonged to the 0 or 1 MM CREG group versus two or more MM CREG group. However, this subgroup concerns only 8% of the transplants performed. CONCLUSIONS: In contrast to other reports, our study showed that HLA matching is by far more beneficial than CREG matching. In the homogenous Eurotransplant population, adjusting the matching criteria toward CREG matching would not lead to an improved graft survival.

Polymorphism of DRw52 and its association with DRw11 and DRw12 in South African blacks (Negroes) and individuals of mixed ancestry (Cape coloreds).

The HLA-DRB3 gene, which encodes the supertypic HLA-DRw52 antigen, has been shown to have limited polymorphism. The alleles at this locus are also in linkage disequilibrium with the alleles at the DRB1 locus. We have studied 16 DRw11 and three DRw12 haplotypes in the South African populations. Five of the DRw11,DQw7 haplotypes were associated with a TaqI restriction fragment length polymorphism which has not been previously described and which correlated with the DRB3 gene. This new variant, which has been called DRw52d, is confined to individuals of black or mixed ancestry. Two of the DRw11,DQw7 haplotypes were also associated with DRw52a or DRw52c and not with DRw52b as has always been observed in white populations. The less common DRw11,DQw6 haplotype, observed in four individuals, also revealed different allelic associations with the DRB3 gene, together with an unusual DQA association. None of the three DRw12,DQw7 haplotypes had the usual association with the DRw52b allele and also demonstrated two distinct DQA associations. The pattern of linkage disequilibrium of the HLA-D region loci in the South African black populations is more complex than in other populations. These findings may be of significance for the matching of unrelated donors for organ transplantation, as well as the study of disease association with HLA.

Unusual HLA-DR,DQ haplotypes found in South African families of black, Asian Indian, and mixed ancestral origin.

Four non-Caucasoid families with the unusual HLA-DR,DQ haplotypes DRw17,DQw7; DR9,DQw2; DR4,DQw2; and DR4,DQw5 were studied. All four haplotypes showed identical serological patterns to those seen with the equivalent Caucasoid antigens, but no HLA-Dw specificities could be assigned. TaqI restriction fragment length polymorphism (RFLP) patterns observed using DRB, DQB, and DQA probes showed that the DRw17,DQw7 haplotype may have originated from a homologous crossover between a DRw17,DQw2 haplotype and a haplotype with DQw7. The results obtained for the DR9,DQw2 and DR4,DQw2 haplotypes suggest that these could have resulted from recombination events with an ancestral "black" DQw2 haplotype. From the RFLP data, it is difficult to postulate the origin of the DR4,DQw5 haplotype being from a single recombination event.

A study of the HLA-Dw determinants and their relationship to DR and DQ antigens in three South African population groups: South African Caucasoids, South African Negroes, and Cape coloureds.

The HLA-Dw specificities of a group of 177 unrelated randomly selected healthy individuals consisting of 67 South African Negroes (Xhosa), 57 Cape Coloureds, and 53 South African Caucasoids were determined. HLA-Dw specificities were determined in a mixed lymphocyte culture test using HTCs. Antigen and gene frequencies as well as the association between HLA-DR, DQ, and Dw were established in three populations. HLA-Dw gene frequencies in the South African Caucasoids agreed with these frequencies in other Caucasoid groups. The HLA-Dw1 frequency was decreased in the Cape Coloureds and South African Negroes compared to the Caucasoids. The gene frequency of Dw3 was low in the South African Negroes in spite of the fact that DR3 is a common DR antigen in this group. A high frequency of Dw 'blank' was observed in the South African Negroes and Cape Coloureds, suggesting the existence of undefined HLA-Dw specificities in these populations. Data concerning the HLA-Dw, DR, and DQ relationships showed that once a certain Dw specificity was associated with a particular DR and DQ antigen, this association remained a fixed entity in the different population groups.

The HLA genetic constitution of the Bushmen (San).

The HLA-A, -B, and -C antigens of 290 and the DR antigens of 212 !Kung San individuals were characterized. The most frequent antigens were HLA-A30 [gene frequency (gf) = 0.193], Bw58 (gf = 0.303), Cw6 (gf = 0.327), DR4 (gf = 0.273), and DQw3 (gf = 0.553). An unexpected finding was the low frequency of the classic African black antigen Bw42 (gf = 0.004). Marked differences as well as similarities in HLA gene frequencies were observed between the San and the South African Negroes, supporting the view that they had a common origin and were then separated for a very long time. During this period differences developed as a result of selective advantage in the Negroes following the pastoralist-agriculturalist way of life as opposed to the hunter-gatherer way of life. The picture is further complicated by the fact that gene flow, mostly from the San to the southern African Negroes, took place when they met again a few hundred years ago. The data also illustrate HLA haplotypes, linkage disequilibria, and four-locus haplotypes not previously seen in other human populations. The most frequent four-locus haplotype in the San, HLA-Aw43,Cw7,B7,DRw6 was also different from A30, Cw2,Bw42,DR3, the most common among the South African Negroes.

The polymorphism of HLA-DR3 in South African populations.

The polymorphism of HLA-DR3 was investigated in families and unrelated individuals of three population groups: South African (SA) Negroes, Cape Coloureds and SA Caucasoids. Serological and restriction fragment length polymorphism (RFLP) analysis indicated that DR3 could be subdivided into DRw17 (previously DR3.1) and DRw18 (previously DR3.2). In contrast, the two-dimensional (2-D) gel electrophoresis patterns could not distinguish between the DRB1 gene products of the HLA-DRw17 and DRw18 cells. Two DRB3 variants, correlating with the T-cell defined specificities Dw24 and Dw25 were identified at the genomic and product level. Of ten haplotypes studied with the newly defined HLA-DRw18 specificity, all had the DRB3 RFLP pattern associated with Dw24. HLA-DRw17 was found in all three population groups tested, although in the SA Negroes HLA-DRw18 was the prevalent DR3 subgroup. This subgroup was also present in the Cape Coloureds but was absent in the SA Caucasoids tested. HLA-DRw18 forms part of the most characteristic SA Negro haplotype, Bw42, DQw4, Dw"RSH," while HLA-DRw17 is part of the classic Caucasoid haplotype, B8, DQw2, Dw3.