David Gallardo

Integrating Y-Chromosome, Mitochondrial, and Autosomal Data to Analyze the Origin of Pig Breeds

We have investigated the origin of swine breeds through the joint analysis of mitochondrial, microsatellite, and Y-chromosome polymorphisms in a sample of pigs and wild boars with a worldwide distribution. Genetic differentiation between pigs and wild boars was remarkably weak, likely as a consequence of a sustained gene flow between both populations. The analysis of nuclear markers evidenced the existence of a close genetic relationship between Near Eastern and European wild boars making it difficult to infer their relative contributions to the gene pool of modern European breeds. Moreover, we have shown that European and Far Eastern pig populations have contributed maternal and paternal lineages to the foundation of African and South American breeds. Although West African pigs from Nigeria and Benin exclusively harbored European alleles, Far Eastern and European genetic signatures of similar intensity were detected in swine breeds from Eastern Africa. This region seems to have been a major point of entry of livestock species in the African continent as a result of the Indian Ocean trade. Finally, South American creole breeds had essentially a European ancestry although Asian Y-chromosome and mitochondrial haplotypes were found in a few Nicaraguan pigs. The existence of Spanish and Portuguese commercial routes linking Asia with America might have favored the introduction of Far Eastern breeds into this continent.

Disparity for the minor histocompatibility antigen HA-1 is associated with an increased risk of acute graft-versus-host disease (GvHD) but it does not affect chronic GvHD incidence, disease-free survival or overall survival after allogeneic human leucocyte antigen-identical sibling donor transplantation

Disparity for the minor histocompatibility antigen HA‐1 between patient and donor has been associated with an increased risk of acute graft‐versus‐host disease (GvHD) after allogeneic human leucocyte antigen (HLA)‐identical sibling donor stem cell transplantation (SCT). However, no data concerning the impact of such disparity on chronic GvHD, relapse or overall survival are available. A retrospective multicentre study was performed on 215 HLA‐A2‐positive patients who received an HLA‐identical sibling SCT, in order to determine the differences in acute and chronic GvHD incidence on the basis of the presence or absence of the HA‐1 antigen mismatch. Disease‐free survival and overall survival were also analysed. We detected 34 patient–donor pairs mismatched for HA‐1 antigen (15·8%). Grades II–IV acute GvHD occurred in 51·6% of the HA‐1‐mismatched pairs compared with 37·1% of the non‐mismatched. The multivariate logistic regression model showed statistical significance (P: 0·035, OR: 2·96, 95% CI: 1·07–8·14). No differences were observed between the two groups for grades III–IV acute GvHD, chronic GvHD, disease‐free survival or overall survival. These results confirmed the association between HA‐1 mismatch and risk of mild acute GvHD, but HA‐1 mismatch was not associated with an increased incidence of chronic GvHD and did not affect relapse or overall survival.

Mapping of quantitative trait loci for cholesterol, LDL, HDL, and triglyceride serum concentrations in pigs

The fine mapping of polymorphisms influencing cholesterol (CT), triglyceride (TG), and lipoprotein serum levels in human and mouse has provided a wealth of knowledge about the complex genetic architecture of these traits. The extension of these genetic analyses to pigs would be of utmost importance since they constitute a valuable biological and clinical model for the study of coronary artery disease and myocardial infarction. In the present work, we performed a whole genome scan for serum lipid traits in a half-sib Duroc pig population of 350 individuals. Phenotypic registers included total CT, TG, and low (LDL)- and high (HDL)-density lipoprotein serum concentrations at 45 and 190 days of age. This approach allowed us to identify two genomewide significant quantitative trait loci (QTL) for HDL-to-LDL ratio at 45 days (SSC6, 84 cM) and for TG at 190 days (SSC4, 23 cM) as well as a number of chromosomewide significant QTL. The comparison of QTL locations at 45 and 190 days revealed a notable lack of concordance at these two time points, suggesting that the effects of these QTL are age specific. Moreover, we have observed a considerable level of correspondence among the locations of the most significant porcine lipid QTL and those identified in humans. This finding might suggest that, in mammals, diverse polymorphisms located in a common set of genes are involved in the genetic variation of serum lipid levels.

CTLA-4 genotype and relapse incidence in patients with acute myeloid leukemia in first complete remission after induction chemotherapy

The recently described single-nucleotide polymorphism CT60, located in the 3′-untranslated region of the CTLA4 (cytotoxic T-lymphocyte antigen 4 ) gene, has been associated with susceptibility to several autoimmune diseases and has also been shown to be involved in immune responses following allogeneic stem cell transplantation (SCT). However, the contribution of the CTLA4 genotype to the control of minimal residual disease in patients with acute myeloid leukemia (AML) has yet to be explored. We investigated the association between the CTLA4 CT60 A/G genotype and the incidence of leukemic relapse in 143 adult patients with AML in first complete remission after the same chemotherapy protocol (CETLAM LAM’03). The CT60 AA genotype was associated with a higher rate of leukemic relapse (56.4 vs 35.6%, P=0.004; hazard ratio (HR)=2.64, 95% confidence interval (CI)=1.36–5.14) and lower overall survival at 3 years (39.4 vs 68.4%, P=0.004; HR=2.80, 95% CI=1.39–5.64). This is the first study to report an association between polymorphisms at CTLA-4 and AML relapse.

Is mobilized peripheral blood comparable with bone marrow as a source of hematopoietic stem cells for allogeneic transplantation from HLA-identical sibling donors? A case-control study

The question of the relative efficacy of stem cell sources (bone marrow vs peripheral blood) for sibling allografts still remains, particularly in relation to quality of life. A study of a relatively homogeneous population has confirmed similar outcomes in terms of overall survival, transplant-related mortality or relapse incidence. However acute and chronic graft-vs-host disease showed increases in the peripheral blood group. Possibly as a consequence, although global quality of life did not differ, there was also a significant impairment of role and social functioning in this group. Background Granulocyte colony-stimulating factor mobilized peripheral blood stem cells are increasingly used instead of bone marrow as a stem cell source for transplantation. Whereas this change is almost complete for autologous transplantation, there are some concerns when considering allogeneic transplants. Design and Methods We performed a retrospective case-control study including 820 adult patients who had received an allogeneic stem cell transplant from an HLA-identical sibling donor. Quality of life (QoL) was assessed in 150 patients using the EORTC Quality of Life Questionnaire C30 (QLQ-C30). Results There were no statistically significant differences in overall survival at ten years (bone marrow: 48.9% vs. peripheral blood stem cells: 39.8%; p=0.621), transplant-related mortality (bone marrow: 28.9% vs. peripheral blood stem cells: 34.4%; p=0.682) or relapse incidence at 9 years (29.4% vs. 35.2%, respectively; p=0.688). Similar outcomes were maintained independently of the phase of the disease. However, multivariate analysis identified a higher incidence of acute graft-versus-host disease grades II-IV (p: 0.023; Hazard ratio [HR]: 1.41; 95% confidence interval [CI]: 1.05–1.89) and grades III-IV (p: 0.006; HR: 1.89; 95% CI: 1.20–2.98), in the peripheral blood stem cells-stem cell transplant group. As previously described, extensive chronic graft-versus-host disease was also more frequent in the peripheral blood stem cells group (28% vs. 15.6%; p<0.001). Patients transplanted with peripheral blood stem cells had significant impairment of role and social functioning. Conclusions Although overall survival was not affected by the stem cell source, peripheral blood stem cell transplants were associated with a higher risk of both acute and chronic GvHD. Global quality of life was similar in both groups, but patients transplanted with peripheral blood stem cells showed worse role and social functioning scores, probably related to the increased incidence of chronic graft-versus-host disease.

Porcine intramuscular fat content and composition are regulated by quantitative trait loci with muscle-specific effects.

Intramuscular fat (IMF) storage is a biological process with a strong impact on nutritional and technological properties of meat and also with relevant consequences on human health. The genetic architecture of IMF content and composition phenotypes has been thoroughly studied in pigs through the identification of QTL and the estimation of genetic parameters. A question that has not been elucidated yet is if the genetic determinants of IMF-related phenotypes are muscle specific or, conversely, have broad effects on the whole skeletal muscle compartment. We have addressed this question by generating lipid QTL maps for 2 muscles with a high commercial value, gluteus medius (GM) and longissimus thoracis et lumborum (LTL), in a Duroc commercial population (n = 350). Our data support a lack of concordance between the GM and LTL QTL maps, suggesting that the effects of polymorphisms influencing IMF, cholesterol, and fatty acid contents are modulated to some extent by complex spatial factors related to muscle location, metabolism, and function. These results have important implications on the implementation of genomic selection schemes aimed to improve the lipid profile of swine meat.

MicroRNA expression at diagnosis adds relevant prognostic information to molecular categorization in patients with intermediate-risk cytogenetic acute myeloid leukemia

Acute myeloid leukemia (AML) is a heterogeneous disease, and optimal treatment varies according to cytogenetic risk factors and molecular markers. Several studies have demonstrated the prognostic importance of microRNAs (miRNAs) in AML. Here we report a potential association between miRNA expression and clinical outcome in 238 intermediate-risk cytogenetic AML (IR-AML) patients from 16 institutions in the CETLAM cooperative group. We first profiled 670 miRNAs in a subset of 85 IR-AML patients from a single institution and identified 10 outcome-related miRNAs. We then validated these 10 miRNAs by individual assays in the total cohort and confirmed the prognostic impact of 4 miRNAs. High levels of miR-196b and miR-644 were independently associated with shorter overall survival, and low levels of miR-135a and miR-409-3p with a higher risk of relapse. Interestingly, miR-135a and miR-409-3p maintained their independent prognostic value within the unfavorable molecular subcategory (wild-type NPM1 and CEBPA and/or FLT3-ITD), and miR-644 retained its value within the favorable molecular subcategory. miR-409-3p, miR-135a, miR-196b and mir-644 arose as prognostic markers for IR-AML, both overall and within specific molecular subgroups.

Polymorphism of the pig acetyl-coenzyme A carboxylase α gene is associated with fatty acid composition in a Duroc commercial line

Acetyl-coenzyme A carboxylase alpha (ACACA) catalyses the first committed step in the biosynthesis of long-chain fatty acids (FA) by converting acetyl-CoA into malonyl-CoA. In pigs, the ACACA gene maps to a chromosome 12 QTL with important effects on FA composition. In the present study, we have sequenced the coding region of the pig ACACA gene in 15 pigs, identifying 21 polymorphic sites that were either synonymous or non-coding. Ten of these SNPs segregated in a Duroc commercial population (n = 350) for which lipid metabolism and meat and carcass quality trait records were available. Significant associations were found between two linked single nucleotide polymorphisms (c.4899G>A and c.5196T>C) and percentages of carcass lean, intramuscular fat, monounsaturated, saturated (myristic, palmitic and stearic) and polyunsaturated (linoleic) FAs in the longissimus thoracis et lumborum muscle, along with serum HDL-cholesterol concentration. The most important allele substitution effects were observed for the polyunsaturated/saturated FA ratio (13-21% of the phenotypic mean) as well as for the percentages of omega-6 and polyunsaturated FAs, especially linoleic acid (7-16% of the phenotypic mean). These results suggest the existence of a causal mutation, mapping to the chromosomal region containing the pig ACACA gene, with marked effects on FA composition of meat.

Bone marrow WT1 levels at diagnosis, post-induction and post-intensification in adult de novo AML.

We retrospectively assessed whether normalized bone marrow WT1 levels could be used for risk stratification in a consecutive series of 584 acute myeloid leukemia (AML) patients. A cutoff value of 5065 copies at diagnosis identified two prognostic groups (overall survival (OS): 44±3 vs 36±3%, P=0.023; leukemia-free survival (LFS): 47±3 vs 36±4%, P=0.038; and cumulative incidence of relapse (CIR): 37±3 vs 47±4%, P=:0.043). Three groups were identified on the basis of WT1 levels post-induction: Group 0 (WT1 between 0 and 17.5 copies, 134 patients, OS: 59±4%, LFS:59±4% and CIR: 26±4%); Group 1 (WT1 between 17.6 and 170.5 copies, 160 patients, OS: 48±5%, LFS:41±4% and CIR: 45±4%); and Group 2 (WT1 >170.5 copies, 71 patients, OS: 23±6%, LFS: 19±7% and CIR: 68±8%) (P<0.001). Post-intensification samples distinguished three groups: patients with WT1 >100 copies (47 patients, 16%); an intermediate group of patients with WT1 between 10 and 100 copies (148 patients, 52%); and a third group with WT1 <10 copies (92 patients, 32%). Outcomes differed significantly in terms of OS (30±7%, 59±4%, 72±5%), LFS (24±7%, 46±4%, 65±5%) and relapse probability (CIR 72±7%, 45±4%, 25±5%), all P<0.001. WT1 levels in bone marrow assayed using the standardized ELN method provide relevant prognostic information in de novo AML.

Predictive factors for poor peripheral blood stem cell mobilization and peak CD34+cell count to guide pre-emptive or immediate rescue mobilization

BACKGROUND AIMS Failure in mobilization of peripheral blood (PB) stem cells is a frequent reason for not performing hematopoietic stem cell transplantation (HSCT). Early identification of poor mobilizers could avoid repeated attempts at mobilization, with the administration of pre-emptive rescue mobilization. METHODS Data from the first mobilization schedule of 397 patients referred consecutively for autologous HSCT between 2000 and 2010 were collected. Poor mobilization was defined as the collection of < 2 × 10(6) CD34(+)cells/kg body weight (BW). RESULTS The median age was 53 years (range 4-70) and 228 (57%) were males. Diagnoses were multiple myeloma in 133 cases, non-Hodgkins lymphoma in 114, acute myeloid leukemia or myelodysplastic syndrome in 81, Hodgkins lymphoma in 42, solid tumors in 17 and acute lymphoblastic leukemia in 10. The mobilization regimen consisted of recombinant human granulocyte-colony-stimulating factor (G-CSF) in 346 patients (87%) and chemotherapy followed by G-CSF (C + G-CSF) in 51 (13%). Poor mobilization occurred in 105 patients (29%), without differences according to mobilization schedule. Diagnosis, previous therapy with purine analogs and three or more previous chemotherapy lines were predictive factors for poor mobilization. A CD34(+)cell count in PB > 13.8/μL was enough to ensure ≥ 2 × 10(6) CD34(+)cells/kg, with high sensitivity (90%) and specificity (91%). CONCLUSIONS The prevalence of poor mobilization was high, being associated with disease type, therapy with purine analogs and multiple chemotherapy regimens. The threshold of CD34(+) cell count in PB identified poor mobilizers, in whom the administration of immediate or pre-emptive plerixafor could be useful to avoid a second mobilization.