Major Mismatch Research Articles

Drug Testing for Chronic Kidney Disease and Diabetes in Animals versus Humans: A Comparative Analysis of Study Designs and Reporting Qualities

Introduction: Diabetes is a highly prevalent accelerator or even cause of chronic kidney disease imposing a large unmet medical need at the global scale. Massive research activities continue to be in search of a cure but the yield of the classical bench-to-bedside research approach has been low. We speculated that a significant mismatch in design and quality of animal and clinical studies in this domain is a hurdle for translation. Methods: We performed a meta-analysis of matched pairs of animal and human studies that tested the efficacy of distinct drug interventions for diabetic kidney disease (DKD). We reviewed study designs and reporting quality of such studies over the last decade according to the standards listed in the CONSORT and ARRIVE recommendations, respectively. Results: We noted a wide diversity in the study designs of animal studies in terms of diabetes induction. Major mismatches with the respective human studies referred to age and sex distribution, comorbidities, stage of the kidney disease, and selection of primary endpoints. Usually, treatment was initiated before onset of kidney disease without any standard of care as a background therapy. The reporting quality of animal studies was poor for randomization procedures, blinding, sample size calculation for a prespecified primary endpoint or the safety analysis. Reporting quality of clinical studies had deficits in trial design-, recruitment-, allocation-, and outcome-related aspects. Conclusion: Bench-to-bedside translation in the domain of DKD suffers from major deficits in the design of experimental studies in view of the projected clinical trials as well as from significant deficits in the reporting quality in preclinical and clinical studies.

ABO incompatibility does not affect transfusion requirements or clinical outcomes of unrelated cord blood transplantation after myeloablative conditioning for haematological malignancies.

The effects of ABO incompatibility on cord blood transplantation (CBT) have not been confirmed. We retrospectively investigated the effect of ABO incompatibility on the clinical outcomes and changes of isoagglutinin titres of 261 consecutive patients who underwent CBT in a single centre. We studied patients with haematological malignancies undergoing unrelated CBT following myeloablative conditioning. There were 80 matched, 72 major mismatched, 72 minor mismatched, and 37 bidirectional mismatched transplants. Risk factors that could potentially influence the patients' outcomes were evaluated. Immunoglobulin M (IgM) isohaemagglutinin antibody (IHA) titres were determined 1 day before and 2, 4, 6 and 8 weeks after the transplant. ABO mismatches did not influence engraftment, transfusion requirements, event-free survival or overall survival following CBT. The anti-donor IgM serum IHA titres fell to ≤1:8 at week 8 after CBT in all patients with ABO major and bidirectional mismatches. The percentages of patients requiring platelet and red blood cell transfusions in the period 31-61 days after CBT were markedly lower than in the period 0-30 days after CBT, being 15 vs 99% for platelets and 23 vs 78% for red blood cells, respectively. Of the 69 recipients of minor mismatched CBT tested, only three with AB blood type developed low titres of anti-recipient IHA after 5 months. In this study ABO incompatibility did not affect clinical outcomes after CBT. A higher number of CD34+ cells infused was correlated with earlier engraftment. Severe acute graft-versus-host disease was associated with poor overall survival. As the IHA titre decreased, so did the number of patients requiring blood transfusion. Rapidly decreasing anti-donor IHA titres and the non-production of donor anti-recipient A and/or B antibodies might contribute to a good outcome of ABO-incompatible CBT with myeloablative conditioning for haematological malignancies.

Nonnative Teachers’ and Observers’ Evaluation of Teachers’ Teaching Quality: Perceptual Convergences and Divergences in Postobservation Conferences

One of the major purposes of observation is the improvement of instruction. In fact, observers evaluate teachers’ teaching quality and negotiate their critical comments with teachers in postobservation conferences. This paper was an attempt to investigate and compare Iranian EFL teachers’ and observers’ evaluations of teachers’ teaching quality. Specifically, we attempted to figure out any possible perceptual matches and/or mismatches between teachers and observers concerning teachers’ teaching quality. To this end, postobservation conferences between three Iranian EFL observers (one female and two male) and 11 Iranian EFL teachers who worked at a language institute in Iran were audio-recorded, transcribed, and analyzed. Our findings indicated that two major themes were recurrent vis-à-vis observers’ evaluations; that is, their evaluations were mainly based on the specific teaching methodology of the institute as well as teachers’ and learners’ pronunciation and intonation. Three perceptual matches were found: teachers’ agreement with the observers’ comments regarding teaching methodology, pronunciation and stress patterns, as well as teachers’ general information. Furthermore, three major perceptual mismatches were found when teachers disagreed with the observers’ comments regarding teaching methodology, pronunciation and stress patterns, and teachers’ time management. Finally, our findings indicated that most of such perceptual mismatches were resolved through the dialogic interaction that occurred between the observers and teachers. A practical implication is that holding such postobservation conferences is of utmost importance as they might be mutually beneficial for both observers and teachers.

Integrating STEM in to TVET Education Programs in QATAR: Issues, Concerns and Prospects

Qatar TVET education system faces unique challenges in that the percentage of vocational secondary school students constitute only 1.4% compared to world average of 15% due to the poor perception of TVET as lesser pathway than other academic-based education. This low perception is associated with sociocultural, economic and institutional factors. Another challenge is the poor link between vocational and general education and the link to labor market. Further challenge is how TVET institutions can develop new curricula, which can respond to the needs of the 21st century skills. This paper will discuss how STEM can help promote TVET education and what are the possible changes required to overcome those challenges. A survey on “Improving and enriching the Human Capital of the State of Qatar through Identification and Development of 21st Century Skills”, explored perceptions of both employers and TVET program leaders toward the skills needed for economic and social developments in a changing world by Meeting Human Capital Needs through 21st Century Skills including the perceptions on needed STEM and cognitive skills. A total of 85 managers and professionals (from more than forty establishments) completed the survey, together with 35 TVET program leaders located in one national university and six government TVET institutions together with 32 semi structured interviews. Descriptive statistics analysis showed a major mismatch between the perceptions of TVET program leaders and employers’ managers and professionals in many aspects., employers perceive the social skills as more important while TVET consider mathematical reasoning as more important employers perceive technological skills such as digital literacy as more important than what TVET leaders perceive. This presentation will identify several approaches to integration and discuss the advantages and disadvantages of the approaches employed. The presentation addresses the various planning approaches and resources required to effectively integrate STEM in TVET programs and curricula.

Abundance and diversity of n-alkane and PAH-degrading bacteria and their functional genes – Potential for use in detection of marine oil pollution

Monitoring environmental status through molecular investigation of microorganisms in the marine environment is suggested as a potentially very effective method for biomonitoring, with great potential for automation. There are several hurdles to that approach with regards to primer design, variability across geographical locations, seasons, and type of environmental pollution. Here, qPCR analysis of genes involved in the initial activation of aliphatic and aromatic hydrocarbons were used in a laboratory setup mimicking realistic oil leakage at sea. Seawater incubation experiments were carried out under two different seasons with two different oil types. Degenerate primers targeting initial oxygenases (alkane 1-monooxygenase; alkB and aromatic-ring hydroxylating dioxygenase; ARHD) were employed in qPCR assays to quantify the abundance of genes essential for oil degradation. Shotgun metagenomics was used to map the overall community dynamics and the diversity of alkB and ARHD genes represented in the microbial community. The amplicons generated through the qPCR assays were sequenced to reveal the diversity of oil-degradation related genes captured by the degenerate primers. We identified a major mismatch between the taxonomic diversity of alkB and ARHD genes amplified by the degenerate primers and those identified through shotgun metagenomics. More specifically, the designed primers did not amplify the alkB genes of the two most abundant alkane degraders that bloomed in the experiments, Oceanobacter and Oleispira. The relative abundance of alkB sequences from shotgun metagenomics and 16S rRNA-based Oleispira-specific qPCR assay were better signals for oil in water than the tested qPCR alkB assay. The ARHD assay showed a good agreement with PAHs degradation despite covering only 25% of the top 100 ARHD genes and missing several abundant Cycloclasticus sequences that were present in the metagenome. We conclude that further improvement of the degenerate primer approach is needed to rely on the use of oxygenase-related qPCR assays for oil leakage detection.

Major ABO Incompatibility Significantly Influences the Survival and Outcomes after Allogeneic Hematopoietic Cell Transplantation in Leukemia - CIBMTR Analysis

Background: Allogeneic hematopoietic cell transplantation (allo-HCT) is a potentially curative treatment for acute myeloid leukemia (AML) and acute lymphoblastic leukemia (ALL). While many factors influence the outcomes of allo-HCT in these leukemias, the independent impact of ABO mismatching between the patient and donor remains unclear.Methods: Using the CIBMTR database, we identified adults aged ≥18 years with AML or ALL who underwent allo-HCT from HLA-matched sibling donor (MSD) or 8/8 matched unrelated donor (MUD) between 2008-2018. We excluded patients who underwent transplant from a mismatched donor source or who received an ex-vivo T-cell depleted graft. Patients were stratified into cohorts based on ABO status (match, minor mismatch, major mismatch, bidirectional mismatch). Outcomes such as overall survival (OS), disease free survival (DFS), non-relapse mortality (NRM), relapse, incidence of acute graft versus host disease (GVHD), chronic GVHD, neutrophil engraftment, platelet engraftment and graft failure were evaluated. Survival analysis was done using Kaplan-Meier method and significant predictors were evaluated using Cox-proportional hazard regression method. Multivariate regression model included main effect (ABO status) and covariates (patient age, gender match, disease type, disease status, HCT-CI, Karnofsky performance status, donor type, conditioning intensity/use of TBI, stem cell source, GVHD prophylaxis, ATG/alemtuzumab use, transplant year).Results: Of 4946 patients who met the study criteria, 2741 patients (55.4%) were ABO matched, 1030 patients (20.8%) had minor ABO mismatch, 899 patients (18.1%) had major ABO mismatch and 276 patients (5.6%) had bidirectional ABO mismatch. Graft manipulation for ABO incompatibility was performed in 900 patients (minor ABO mismatch=532; major ABO mismatch=226; bidirectional mismatch=142), however, the information on individual graft manipulation techniques was limited. In multivariable analysis (table 1), major ABO mismatch significantly affected the OS, platelet engraftment and primary graft failure. Compared to ABO matched allo-HCT, major ABO mismatched allo-HCT was associated with worse OS (major mismatch - HR 1.16, 95% CI 1.05-1.29; p=0.005), inferior platelet engraftment (HR 0.83, 95% CI 0.77-0.90; p=<0.001), and higher risk of primary graft failure [4.5% (major mismatch) vs. 3.2% (ABO matched) - HR 1.60, 95% CI 1.12-2.30, p=0.01]. There was a significant interaction between the ABO status and graft type (peripheral blood vs. bone marrow) for the acute GVHD grades 2-4 model, and they are presented separately (table 1). Bidirectional ABO mismatch also significantly impacted the outcomes such as acute GVHD grades 2-4 (in bone marrow stem cell subgroup, HR 0.50, 95%CI 0.27-0.93, p=0.02) in addition to a trend towards inferior survival and NRM (p-value not significant). Other outcomes such as relapse (p=0.41), acute GVHD grades 3-4 (p=0.13), and chronic GVHD (p=0.30) were not significantly influenced by the ABO status.Conclusions: Our study demonstrates that pre-transplant ABO status is an independent predictor of survival and other post-transplant outcomes in a large cohort of patients with AML and ALL undergoing allo-HCT in the recent era. This demonstrates the importance of considering ABO status in the donor selection algorithms and potential strategies to mitigate its adverse impact. Due to the limited information available on graft manipulation strategies, the impact of graft manipulation techniques on the outcomes could not be evaluated and needs to be investigated in future studies. [Display omitted] DisclosuresGuru Murthy: Techspert: Consultancy; Guidepoint: Consultancy; Cancerexpertnow: Honoraria; Qessential: Consultancy; TG therapeutics: Other: Advisory board; Cardinal Health Inc.: Honoraria. Devine: Magenta Therapeutics: Current Employment, Research Funding; Tmunity: Current Employment, Research Funding; Sanofi: Consultancy, Research Funding; Johnsonand Johnson: Consultancy, Research Funding; Orca Bio: Consultancy, Research Funding; Be the Match: Current Employment; Vor Bio: Research Funding; Kiadis: Consultancy, Research Funding. Farhadfar: Incyte: Consultancy. Sharma: Vertex Pharmaceuticals/CRISPR Therapeutics: Other: Salary support paid to institution; Novartis: Other: Salary support paid to institution; Spotlight Therapeutics: Consultancy; Medexus Inc: Consultancy; CRISPR Therapeutics: Other, Research Funding; Vindico Medical Education: Honoraria. Stefanski: Novartis: Honoraria. Pulsipher: Equillium: Membership on an entity's Board of Directors or advisory committees; Adaptive: Research Funding; Jasper Therapeutics: Honoraria. Shaw: mallinkrodt: Other: payments; Orca bio: Consultancy.

Non-Genetic Determinants of Clonotypic T Cell Expansion Following Allogeneic Stem Cell Transplant

Background:The immunologic basis of acute GVHD fundamentally involves alloreactive donor T cells that recognize foreign major histocompatibility complex (MHC)-peptide structures derived from both major and minor antigen mismatches with the host. Within this paradigm, the relationship between the donor and recipient genetics represents a closed system that dictates the potential ability of any given T cell receptor (TCR) to expand, raising the question of whether there are predictable aspects of TCR reconstitution at a clonal level. Take a hypothetical example - if genetically identical twins were to receive allogeneic grafts from the same donor and both recipients develop GVHD, would one expect similar TCRs to be clonally expanded? It has been challenging to rigorously explore this phenomenon, however, because of the vast combinatorial diversity of αβ TCRs, the high prevalence of low copy number TCRs, and sampling constraints - all of which render tracking and comparing TCR expansion between the donor and host difficult.Methods:We address these challenges in order to better understand the predictability TCR clonal dynamics through an analysis platform utilizing 1) a series of matched and mismatched murine transplant experiments where genetically identical littermates receive T cells from the same polyclonal donor pool, thus creating multiple transplant replicates simulating the twin transplant system describe above (Fig 1), and 2) probabilistic modeling of individual TCR frequencies to account for partitioning stochasticity (variation in how low copy number TCRs are distributed from donor to recipient). We conduct high-throughput DNA-based TCR amplicon sequencing for both donor and post-transplant recipient samples to generate over 20 million TCRs and model the expansion rates of all identifiable TCRs in each transplant system using a Bayesian approach.Results:While overall V and J gene usage were similar amongst identical recipients (Fig 2), we find that a small fraction of TCR clonotypes appears to have widely disparate clone counts amongst identical recipients receiving the same donor T cell pool. For example, we saw 9,739, 129 and 0 copies of a particular TCR in 3 different recipients in our B6->B6D2F1 system (Fig 3). In order to distinguish whether TCR count discrepancies seen across identical recipients is simply a reflection of donor partitioning stochasticity or true differential expansion (Fig 4), we apply a Bayesian algorithm to identify differential expanders, which represent TCRs that are asymmetrically expanded between recipients of a genetically identical pair (Fig 5). These TCRs can be generated from both memory and naïve T cell compartments.The presence of these differentially expanded clones amongst identical recipients suggests that non-genetic dependent mechanisms may influence which TCRs expand post-transplant. We next show that broad gut decontamination of microbiota with peri-transplant vancomycin, gentamicin, cefoxitin and metronidazole dramatically reduced the fraction of differential expanders (p<0.0001). However, the change in inflammation from microbiome depletion did not appear to drive this difference, as 1) MyD88/TRIF double knockout recipients (deficient TLR signaling) did not show a reduction in differential expanders, and 2) altering conditioning intensity (900cGy to 1300 cGy TBI) also did not change the fraction of differential expanders. Rather, the difference is likely antigenically driven, as differential expanders are enriched in antigen specificity compared to other TCR sequences (p<0.0001) based on published algorithm that identify TCRs with similar amino acid sequence overlap.Conclusions:These results refine our current understanding of clonal T cell selection and expansion after allogeneic BMT and suggests that for a given transplant system, individual TCR selection is not solely dictated by genetic donor and recipient major and/or minor histocompatibility disparities. Rather, microbiota-derived molecules appear to behave as minor antigens to direct systemic clonal TCR selection. These data suggest a novel mechanism by which the microbiome may modulate transplant outcome, challenging current paradigms suggesting the microbiota primarily drive inflammation via their PAMP activities. [Display omitted] DisclosuresHill: Applied Molecular Transport: Research Funding; Syndax Pharmaceuticals: Research Funding; Compass Therapeutics: Research Funding; NapaJun Pharma: Consultancy; Generon corporation: Consultancy; iTeos Therapeutics: Consultancy, Research Funding; Neoleukin Therapeutics: Consultancy; Roche: Research Funding.

Impact of Donor Age and Relationship on Outcomes of Peripheral Blood Haploidentical Hematopoietic Cell Transplantation

Introduction: Haploidentical hematopoietic cell transplantation (haplo-HCT) is an increasingly utilized therapy for a variety of hematologic malignancies. Determining which donor characteristics affect transplant outcomes is of particular interest in haplo-HCT, as there are often multiple donors available for a given patient. A survival benefit with younger donors has been reported in some recent observational studies (DeZern et. al., Blood Advances, March 2021); (Canaani et. al., AJH, Sep. 2017). A decrease in non-relapse mortality (NRM) and increase in relapse with no overall survival difference associated with younger donors has also been observed (Mariotti et. al., Blood Advances, June 2020). These previous studies have utilized populations with bone marrow as the predominant stem cell source. Solomon et al. (BBMT Sep. 2018) observed poorer survival, increased relapse, and worse NRM with parent donors relative to children in a largely peripheral blood population. HLA DR and DP mismatch were noted to be associated with improved survival. Here we describe outcomes in peripheral blood haplo-HCT and their association with potentially selectable donor characteristics including age and relationship to the patient.Patients and Methods: We performed a retrospective review of patients who underwent peripheral blood haplo-HCT with PtCy from July 2009 through May 2021. A total of 323 patients were identified with AML (205), ALL (43), MDS (26), and other (49). Univariate and multivariate analyses (MVA) were conducted examining the effect of donor characteristics on overall survival (OS), NRM, relapse, acute and chronic GVHD. Donor characteristics included age, relationship, ABO status, CMV status, and HLA match grade. We controlled for patient characteristics known to affect outcomes including disease type, DRI, HCT CI, KPS, active disease at transplant, myeloablative conditioning, and prior HCT.Results: Median donor age was 40 (range 15-71) with male predominance (64%). Most were ABO compatible (63%) - 12% had major ABO mismatch, 20% minor, and 4% bidirectional. Donor-recipient CMV status matched in 61% of pairs, 13% were donor positive-recipient negative, 26% donor negative-recipient positive. Most were 5/10 HLA matched (51%) with 20% 6/10 and 13% 7-9/10. Univariate analysis revealed that increasing donor age was associated with higher NRM (HR 2.29, p=0.005 for donors age 30-44; HR 2.06, p=0.012 age > 44) but lower relapse risk (HR 0.56, p=0.012 age 30-44; HR 0.69, p=0.10 age > 44). There were no differences in aGVHD or cGVHD based on donor characteristics in univariate analysis. In MVA, relapse risk was lower in patients with older donors , p=0.046). In contrast, NRM was higher in patients with older donors (HR 1.73 age 30-44, HR 1.69 age > 44, p=0.010). There was no difference in overall survival based on donor age (HR 1.23 age 30-44, HR 1.38 age > 44, p=0.11). We next examined the effect of donor relationship on outcomes while controlling for donor age, patient age, and patient disease risk factors. We found no difference in outcomes between parent, sibling, or child donors.Conclusions: Increasing donor age was associated with lower relapse risk but higher NRM. These competing effects resulted in no difference in OS based on donor age. Other donor factors including relationship (parent / sibling / child), CMV status, ABO mismatch, donor sex, and HLA match grade were not associated with outcomes. Solomon et al. reported better outcomes with child compared to parent donors, a finding not replicated here, however our analysis controlled for donor age which could have been a proxy for relationship in their study. These data suggest that in peripheral blood haplo-HCT, younger donors may be preferred in patients with high risk of transplant related complications. In contrast, older donors may be preferred in patients where relapse risk is high. Data on HLA-DR and DP match is being analyzed and will be presented at the ASH 2021 meeting. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Microbial-Derived Metabolites Induce Epithelial Recovery Via the Sting Pathway in Mice and Men and Protect from Graft-Versus-Host Disease

Background: Graft-versus-host disease (GVHD) is a dreaded complication after allogeneic stem-cell transplantation (ASCT). Previously, we and others showed that activation of type I interferon (IFN-I) inducing pathways such as RIG-I/MAVS or cGAS-STING can promote the integrity of the intestinal barrier and limit GVHD (Fischer et al., Sci Transl Med, 2017). However, the signals that drive these protective IFN-I responses are poorly understood.Commensal microbiota can (i) have distant effects on immune responses through modulation of IFN-I signaling (Steed et al, Science, 2017; Swimm et al, Blood, 2018) and (ii) predict mortality in ASCT patients (Peled et al., N Engl J Med, 2020). We hypothesized that microbiota-derived products such as microbial metabolites engage IFN-I signaling in immune and non-immune cells poising them for induction of protective responses. We established a prospective, multi-centric clinical study in patients newly diagnosed with acute leukemia and performed longitudinal stool sampling to track changes in microbial community composition and metabolites expression levels. Submitted as a separate abstract to ASH 2021 (Orberg & Meedt et al.), we show that patients with high metabolite expression going into ASCT are less likely to develop GVHD. In this study, we translate our clinical observations to mouse models of acute GVHD and human and mouse intestinal organoids to uncover the molecular mechanisms via which metabolites protect the intestinal barrier during ASCT.Methods: Stool samples from ASCT patients were obtained in Munich and at Regensburg in accordance to IRB-approved study protocols. Patients were sampled at initial diagnosis (Dx), prior to conditioning and weekly after ASCT up to day 28. We analyzed samples by 16S rRNA sequencing and mass spectrometry to obtain a complete picture of microbiome composition and function. Next, we tested metabolites which we detected in patients (desaminotyrosine [DAT], indole-3-carboxaldehyde [ICA]) as treatment in preclinical models in ASCT and GVHD mouse models. Outcomes were assessed by a novel organoid recovery assay in addition to established read-outs. To obtain a mechanistic understanding of the signaling pathways involved, we stimulated WT or IFN-I-signaling-impaired mouse (incl. STING -/-, MAVS -/-, IFNαR -/-) as well as human intestinal crypt-derived organoids with metabolites.Results: Here, we present a 64-year-old female patient diagnosed with AML who received a 9/10 HLA-matched ASCT. At day 7, i.v. antibiotics were started due to fever and Enterococcus bacteraemia. At day 15, the patient developed skin and GI GVHD (Glucksberg III). At the timepoint initial diagnosis (Dx), i.e. the timepoint when we diagnosed AML but before therapy was initiated, we detected rich alpha diversity (Panel 1a) in the patient's stool. Flavonifractor plautii, a producer of the metabolite DAT, was detectable (Panel 1b). We observed high-level expression of metabolites including short-chain fatty acids, DAT, ICA and secondary bile acids (Panel 1c). Following ASCT, and especially at the early time-points day 0 and day 7, alpha diversity and metabolite expression declined drastically. We confirmed this trend in our multi-centric cohort of ASCT patients by comparing levels of DAT and ICA sampled at admission to the transplantation ward (Conditioning) versus at clinical diagnosis of GVHD: in patients with GVHD, metabolite levels were drastically reduced (Panel 2). Next, we prophylactically administered metabolites in a major mismatch mouse ASCT model. Metabolite-treated mice showed significantly showed better outcomes in our organoid recovery assay, which measures the ability of intestinal stem cells to recover after allogeneic injury (Panel 3). This effect that was abrogated in STING -/- recipients. Metabolite stimulation of mouse small intestinal organoids promoted organoid numbers and size, and required intact STING signaling (Panel 4a). Human colon organoids also responded to DAT and ICA, however the metabolite effect was lost when co-administered with the STING-inhibitor H151.Conclusions: We identify that microbial-derived metabolites detected in patients can engage the STING pathway in humans and mice to confer resistance from immune damage. Thus, prophylactic administration of metabolite cocktails or bacterial consortia that can produce these metabolites may reduce occurrence of GVHD in ASCT patients. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Addressing Mismatch between TVET Programs and Skill Needs in the Finance and Banking Sector – A Case Study from Qatar

Links between Qatar’s labor market and post-secondary education are not fully effective and often result in neglected or duplicated development of human capital. Therefore, most development efforts occur in isolation involving outmoded programs with many complacent faculty unaware of new technologies and developments relevant to labor market sectors. Analyses of secondary data from government departments and international studies were combined with a survey on “Improving and enriching the Human Capital of the State of Qatar through Identification and Development of 21st Century Skills”. This explored perception of both employers and TVET program leaders toward the skills needed for economic and social development in a changing world by meeting human capital needs through 21st century skills. A total of 85 managers and professionals completed the survey, together with 35 TVET program leaders from one university and five government TVET institutions (the survey was adapted to fit the context of TVET institutions surveyed). Thirty-two of the industry managers and professionals were from Hydrocarbon and Energy, 26 from Built Environment and 27 from Banking Finance sectors. Subsequently, 32 semi-structured interviews were conducted. Descriptive statistics using T-test and effect size for comparison, showed a major mismatch between perceptions of TVET program leaders and business finance sector’ managers and professionals in many aspects of 21st century skills requirements. These were mainly in social skills and some specific technology skills. Significantly, the study indicated weak links between employers and TVET institutions. To address these issues, minimizing the skills’ mismatch can be achieved by placing greater emphasis on reforming the curricula of Qatar’s TVET institutions, to facilitate faster transitions into the workplace.

Sexual (in)equality? A meta‐analysis of sex differences in thermal acclimation capacity across ectotherms

Abstract Climate change is putting the fate of ectothermic animals at stake because their body temperature closely tracks environmental temperatures. The ability to adjust thermal limits and preference through acclimation (i.e. acclimation capacity) may compensate for temperature changes. However, although necessary for forecasting the future of ectotherms in a changing climate, knowledge on the factors modulating these plastic responses is fragmentary. For instance, the influence of an animal's sex in driving acclimation capacity has been underappreciated. Here, we present the first systematic review and meta‐analysis on sex differences in thermal acclimation capacity. Using 239 effect sizes from 37 studies and 44 species, we revealed that males and females did not differ significantly in their overall capacity to acclimate their thermal limits and preference. However, in some instances, females expressed significantly greater plastic responses than males. In wild animals, females had a greater heat tolerance plasticity than males. In addition, females had a greater cold tolerance plasticity in terrestrial habitats, but the strength and direction of this sexual dimorphism was associated with the duration of acclimation. We also found a negative correlation between body mass and plasticity. Finally, we demonstrated that the capacity for each sex to adjust their thermal tolerance and preference was remarkably limited. It is important to acknowledge that the above effects were weak and heterogeneous. Hence, in the species we investigated, minor differences in acclimation capacity may not translate into major ecological mismatch between sexes with climate change. Our systematic review also revealed that over 75% of the studies we identified either did not report or confounded the sex of the animals. This under‐reporting may cause to overlook ecologically relevant sex differences in plasticity in ectothermic taxa. We stress the need for further research on sex‐based responses to temperatures. Our synthesis provides additional evidence that the capacity for ectotherms to acclimate to temperatures is limited, and likely insufficient to compensate for the impacts of climate change. A free Plain Language Summary can be found within the Supporting Information of this article.

Expectations and Perceptions of the Internship Program: A Case Study of Tourism Retail and Marketing Students in Macao

ABSTRACT Previous studies show that minimizing the gap between students’ expectations and perceptions of internships is essential to attract students to enter the same industry upon graduation, but research on the concerns in retail tourism internships is lacking. The current study employed mixed methods to address this gap. Qualitative in-depth interviews with 26 senior year Tourism Retail and Marketing Management students in Macao were conducted to examine their expectations and perceptions of internships, identify mismatches between these and explore consequences of perceived mismatches. Then quantitative surveys were carried out to collect opinions from a larger sample. Results show that the majority of respondents perceived gaps between their expectations and perceptions of their internship. Broken promises, negative company atmosphere and lack of supervision were the major mismatches. The majority of the interns either intended to remain in the retail industry or were undecided about their career intentions. Based on the findings, practical suggestions are given to major stakeholders to improve retail tourism internship programs.

Rapid and Efficient Gene Editing for Direct Transplantation of Naive Murine Cas9+ T Cells.

Gene editing of primary T cells is a difficult task. However, it is important for research and especially for clinical T-cell transfers. CRISPR/Cas9 is the most powerful gene-editing technique. It has to be applied to cells by either retroviral transduction or electroporation of ribonucleoprotein complexes. Only the latter is possible with resting T cells. Here, we make use of Cas9 transgenic mice and demonstrate nucleofection of pre-stimulated and, importantly, of naive CD3+ T cells with guideRNA only. This proved to be rapid and efficient with no need of further selection. In the mixture of Cas9+CD3+ T cells, CD4+ and CD8+ conventional as well as regulatory T cells were targeted concurrently. IL-7 supported survival and naivety in vitro, but T cells were also transplantable immediately after nucleofection and elicited their function like unprocessed T cells. Accordingly, metabolic reprogramming reached normal levels within days. In a major mismatch model of GvHD, not only ablation of NFATc1 and/or NFATc2, but also of the NFAT-target gene IRF4 in naïve primary murine Cas9+CD3+ T cells by gRNA-only nucleofection ameliorated GvHD. However, pre-activated murine T cells could not achieve long-term protection from GvHD upon single NFATc1 or NFATc2 knockout. This emphasizes the necessity of gene-editing and transferring unstimulated human T cells during allogenic hematopoietic stem cell transplantation.

Benchmarking of the English Language Component of the Mature Students’ Entrance Examinations in Ghana against the WASSCE English Language Component

This paper was purposed on benchmarking the English language component of the Mature Students’ Entrance Examinations (MSEE) (administered in Ghana by universities to select undergraduate candidates) to the English language component of the West African Senior School Certificate Examination (WASSCE) which constitutes the mainstream Ghanaian university undergraduate entrance examination. The qualitative design (specifically, multiple case study design) was employed for this study. Using the multi-stage sampling technique, six Ghanaian universities (from whom eighteen English language component of the MSEE past questions were elicited) and three sets of the WASSCE English language component past questions administered in 2016, 2017 and 2018) were used for the study. O’Leary’s (2014) eight steps of conducting document analysis were used to analyse the data. The study revealed that there are major mismatches between the two sets of examinations in the areas of the test types, the basic language skills tested and the competences tested. The study advanced, to policy makers, suggestions such as testing Speaking and Listening for the improvement of the two sets of examinations. Received: 2 May 2021 / Accepted: 15 June 2021 / Published: 8 July 2021

Post-Transfusion Overall Survival for HLA- Alloimmunized Platelet Refractorythrombocytopenic Leukemia Patients

BACKGROUND: Platelet alloimmunization and refractoriness is an established complication of platelet (PLT) transfusion, known to be treatment-limiting with potential life-threatening complications for hemato-oncologic patients undergoing chemotherapy (Hess et al.). Previous studies have explored the association of PLT refractoriness with lymphocytotoxic antibody positivity, more frequent platelet transfusion and platelet age (Slichter et al, Blood, 2005). They have also developed cross matching or HLA matching for donor selection as the basis of managing refractory patients. However, the impact of patient characteristics on the clinical outcomes of refractoriness is not fully understood.Objective: To analyze overall survival (OS) of PLT refractory leukemia patients transfused with HLA-matched platelets in association with the demographic and clinical characteristics of these patients.METHODS: We analyzed the demographics, clinical characteristics and platelet refractoriness (defined as two consecutive <24 hr post-transfusion corrected platelet count increments (CCI) of < 5 K/uL), ABO matching (fully compatible, minor mismatch or major mismatch), and HLA match grade (A, B, C). The threshold for transfusion was <7 K/uL and only patients with recorded platelet counts pre- and post-transfusion were included. The majority of post-transfusion counts were performed < 24 hrs. Transfusion efficacy was defined as having: i) difference in platelet counts > 5 pre- and post-transfusion; ii) ABO match or compatible; iii) HLA match grade A or B1.We included two measurements for overall survival (OS): from the date of the first transfusion and also from the last transfusion to the date of last follow-up or date of death. A PLT transfusion record for each measure was obtained using a majority rules algorithm. Group differences in OS were assessed using the log-rank test. For differences in PLT counts, the median value for each patient was used. The association between mean PLT count difference, age, and weight with OS was evaluated using univariate Cox proportional regression models.RESULTS: We identified 33 leukemia patients who received a total of 145 HLA-matched platelet transfusions from 2010-2017. A majority [60%] of the patients had a diagnosis of acute myeloid leukemia or myelodysplastic syndrome and received ≤2 HLA-matched platelet transfusions. Complex cytogenetics was identified in 7 of 23 (30%) patients, while the majority (97%) showed broad reactivity against HLA Class I and Class II antigens. Less than half (45%) of the platelets transfused were HLA match grade B and a third of the platelets (35%) were ABO compatible.The median OS was 16.7 months and the 1-year OS rate was 64%. A significant association between post-transfusion OS with age [p=0.016] and complex cytogenetics [p=0.013] was observed. However, no significant association between OS and gender, weight, ethnicity, and the number of pregnancies was noted. Although the median OS was longer for patients with ABO match/compatible transfusions (vs. incompatible), HLA match grades B2/B3/C platelets (vs. A/B1), and >2 platelets transfusions (vs. ≤ 2), none of these differences were statistically significant (Table 1).CONCLUSIONS: In summary, our findings reveal a significant association of OS was noted with age and complex cytogenetics in platelet refractory hemato-oncologic patients who received HLA-matched platelet transfusions. This association prompts further investigation into how patient profiles may be used in HLA-alloimmunized patients with platelet refractoriness. However, this may need to be studied in a larger cohort of patients in the future. [Display omitted] DisclosuresDiNardo:Celgene: Honoraria, Research Funding; Agios: Honoraria, Research Funding; AbbVie: Honoraria, Research Funding; Novartis: Honoraria, Research Funding; Daiichi-Sankyo: Honoraria, Research Funding. Cortes:ImmunoGen: Consultancy, Research Funding; ARIAD: Consultancy, Research Funding; Novartis Pharmaceuticals Corporation: Consultancy, Research Funding; BMS: Consultancy, Research Funding; Teva: Research Funding; Sun Pharma: Research Funding; Pfizer: Consultancy, Research Funding. Kantarjian:Delta-Fly Pharma: Research Funding; Amgen: Research Funding; Novartis: Research Funding; Bristol-Meyers Squibb: Research Funding; Pfizer: Research Funding; ARIAD: Research Funding.

A Novel Role for AMP-Activated Protein Kinase (AMPK) in Controlling T Cell Sensitivity to IL-6

Allogeneic hematopoietic stem cell transplantation (alloHSCT) is a curative treatment for high-risk leukemia and many non-malignant hematologic disorders. However, the use of alloHSCT as a therapy remains limited by acute graft-versus-host disease (GVHD), where activated donor T cells attack and destroy host tissues in the skin, gastrointestinal tract, and liver. We have previously shown that alloreactive T cells increase phosphorylation of the intracellular energy sensor AMP-activated protein kinase (AMPK) as early as day 3 post-transplant (levels 8-fold higher in alloreactive T cells compared to naive or syngeneic controls (p=0.0003)) and donor T cells lacking AMPK (AMPK KO) cause significantly less GVHD in both major- and minor-histocompatibility mismatch models of GVHD. At the same time, AMPK KO T cells equivalently clear leukemia cells and extend recipient survival in leukemia models with high tumor burden. Transplantation of cells lacking AMPK lowers the number of donor T cells recovered on day 7 post-transplant (3.15 +/- 0.49x106 versus1.87 +/- 0.53x106, p=0.0006, wildtype (wt) versus AMPKKO respectively) but interestingly, increases both the percentage and total number of regulatory T cells (Treg) (0.85 +/- 0.32x104 vs. 1.69 +/- 0.34x104, wt vs. KO, p=0.004).To understand these findings mechanistically, we sought to study the role of AMPK in donor Treg expansion and stabilization post-alloHSCT. Canonically, interleukin-2 (IL-2) signaling, through phosphorylation of Stat5, promotes Treg generation and stability, while inflammatory signals (e.g. IL-6 signaling through Stat3) are known to decrease Treg stability post-transplant. Studies testing IL-2 sensitivity in KO cells, or the degree of Stat5 phosphorylation post-transplant, were unrevealing. However, AMPK KO Tregs significantly decrease their sensitivity to IL-6, with a 50% reduction in phosphorylated-Stat3 levels following intermediate levels of cytokine stimulation (Figure 1A). To demonstrate the functional consequences of this difference in cytokine sensitivity, we exposed in vitro generated Treg to increasing concentrations of IL-6. As expected, the percentage of wt FoxP3+ cells decreased sharply with increasing IL-6 concentrations (decreasing from 50% to 14%), while AMPK KO Tregs were minimally affected over this dose range (47% versus 42%; Figure 1B, p<0.02).To determine if increased Treg stability was the predominant reason for improvements in GVHD seen with AMPK KO cells, CD25+ cells were removed from the donor inoculum of AMPK KO or wt mice by magnetic selection (to eliminate >90% of Treg), and then either CD25-replete or CD25-depleted sets of cells were transplanted into B6xDBA2 F1 recipients in a major MHC mismatch model of GVHD. Despite a decrease in AMPK KO Treg numbers on day 7 to levels commiserate with wt donors (Figure 2A), elimination of AMPK KO Treg had only a modest effect on GVHD severity (Figure 2B), implicating a substantial role for AMPK in the non-Treg compartment.To explore the role of AMPK in non-Tregs further, we investigated the sensitivity of unmanipulated wt and AMPK KO T cells to IL-6 stimulation. Similar to FoxP3+ cells, both naïve CD4 and CD8 T cells lacking AMPK experienced lower levels of Stat3 phosphorylation in response to graded concentrations of IL-6 (Figure 3A). Furthermore, if IL-6 was added to a 4-day culture with anti-CD3 stimulation, the number of wt T cells recovered after 96 hours increased >2.2-fold. In contrast, AMPK KO cells experienced minimal changes in T cell number over a wide range of IL-6 concentrations (Figure 3B).In conclusion, we define in these studies a novel role for AMPK in controlling donor T cell sensitivity to IL-6, both at the level of Stat3 phosphorylation, and in terms of functional consequences. Not only does this striking result elegantly explain our previous observation of increased Treg percentages, and decreased donor T cell numbers, following transplantation of AMPK KO cells, it continues to make the case that inhibition of AMPK represents an attractive clinical target for the amelioration of GVHD. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

Investigation of risk factors associated with erythrocyte engraftment after ABO-incompatible hematopoietic stem cell transplantation.

ABO-incompatible hematopoietic stem cell transplantations (HSCTs) are widely practiced; however, the delay in erythrocyte engraftment can be problematic. While erythrocyte engraftment is usually indicated by an increase in reticulocyte levels without the need for erythrocyte transfusions, the disappearance of recipient-derived anti-A/B isoagglutinin and detection of donor-derived A/B antigens can also be used as other parameters. We conducted a retrospective analysis of 68 ABO-incompatible HSCTs, focusing on major and bidirectional mismatch. We analyzed known clinical risk factors associated with delayed erythrocyte engraftment using the three parameters (disappearance of anti-A/B isoagglutinin in recipient, detection of donor-derived A/B antigen, and reticulocyte levels >1%). Although the three parameters were well correlated, the results showed heterogeneity when analyzing the associated risk factors for delayed erythrocyte engraftment. In the analysis of all cases, the requirement for an HLA-matched platelet transfusion was a common risk factor. Furthermore, erythrocyte engraftment was slower in adults than in children. In adults, cytomegalovirus antigenemia was a risk factor for two parameters; however, in children, underlying disease was a common risk factor for all parameters. There is a complex relationship between erythrocyte engraftment and various factors related to HSCTs. Our results suggest that greater accuracy is possible by using analysis methods other than the measurement of reticulocyte levels.

Pure red blood cell aplasia: patient management pitfalls in major ABO-incompatible haematopoietic cell transplantation.

In this issue of British Journal of Haematology, Longval and colleagues report on the successes and pitfalls of prediction, diagnosis and management of pure red blood cell aplasia (PRCA) after major ABO-incompatible allogeneic stem cell transplantation.1 Their data confirm prior reports indicating a paucity of cases of PRCA after umbilical cord blood transplantation.2 Among the remaining 587 patients receiving major ABO-incompatible donor grafts, 66 cases of PRCA were identified. In this series, high pre-transplant isohaemagglutinin titre was the only pre-transplant variable associated with subsequent development of PRCA. Despite the various methodologies for performing isohaemagglutinin titre assays,4, 5 this observation highlights the need for pre-transplant quantitation of isohaemagglutinin titre in instances of major ABO mismatch in order to identify patients at highest risk. Furthermore, at some centres pre-conditioning therapeutic plasma exchange is considered for patients with high titres.6 In some patients, residual recipient plasma cells may secrete sufficient quantities of antibody to prevent nascent erythroblast precursors from achieving timely maturation,3 forming a rationale for consideration of plasma cell-directed therapy in patients with persistently high isohaemagglutinin titres. Interestingly, several low titre isohaemagglutinin recipient–donor pairs resulting in post-transplant PRCA were identified, pointing to the need to identify additional predictors of PRCA. This work also exposes a gap in knowledge within the transplant field, namely when can we be sure that red blood cell engraftment has transpired? To date, no universal standard for red blood cell engraftment exists, and across institutions several working definitions exist including red cell transfusion independence, ABO blood type conversion, and absence of reticulocytopenia. Advantages and disadvantages of various case definitions are outlined in Table I. Standard diagnostic assessments for profound anaemia, reticulocytopenia, and absence of red blood cell precursors in the bone marrow are certainly essential when patients display prolonged red cell transfusion dependence,7 but in the absence of alternative causes, the presumptive diagnosis in this setting is PRCA due to ABO incompatibility, and persistence of some level of the corresponding isohaemagglutinin is typical. In this series, over 10% of patients receiving transplants from donors with a major ABO incompatibility developed PRCA, but this immunologic insult did not significantly affect overall survival, which is in line with results of previous studies.8, 9 As such, any therapeutic intervention must be considered in the context of the inherent risk of the intervention itself. Introduction of rituximab,10 donor lymphocyte infusions,11 tapering of immunosuppressive medications, or use of a proteasome inhibitor12 all carry risk–benefit calculations, and to date have demonstrated mixed results in treating post-transplant PRCA. Anecdotal reports of the successful use of the anti-CD38 monoclonal antibody daratumumab for management of post-transplant PRCA13, 14 deserve mention, particularly given that the physiology of prolonged PRCA after transplant is presumably a reflection of residual host plasma cell clones producing the isohaemagglutinin. In the current study, no particular intervention was associated with more rapid resolution of PRCA, although no patients in this series received daratumumab and only a handful received a proteasome inhibitor. Management of patients undergoing major ABO-incompatible allogeneic stem cell transplantation requires close collaboration with the blood bank to identify patients at greatest risk prior to transplantation, and to optimize transfusion strategies during the post-transplant period. This may include pre-transplant minor red blood cell antigen typing to reduce the likelihood of alloimmunization,15 and consideration of pre-transplant plasma exchange for patients with very high isohaemagglutinin titres. It is fair to say that there is no ‘gold standard’ for treatment of active post-transplant PRCA, and this study does not inform us regarding how best to manage these patients. Fortunately, the situation typically resolves over time, but further studies to establish effective treatment strategies are sorely needed given the significant morbidity and symptom burden associated with this not so rare complication. As a final thought, we might speculate two distinct (but not mutually exclusive) mechanisms for what we call ‘PRCA’ in this setting: residual host isohaemagglutinin levels that simply take time to dissipate because of high pre-transplant titres versus persistence of host isohaemagglutinin-producing plasma cells. We have the molecular and blood banking tools to systematically study these post-transplant events, which could better elucidate management approaches most appropriate for individual patients. Variability in RBC products Complicated by suspected transfusion reactions Provider/institutional variability for transfusion thresholds Time to independence is not a universal definition (e.g,, 30 days without transfusion versus 60 days) Inter-observer variability in scoring of agglutination Interference between residual transfused isohaemagglutinins from platelets/plasma No established grading system for mixed field observations Interference from intravenous immune-globulin treatment Results often not reported in patient’s medical record Difficulty in interpretation for non-transfusion medicine personnel Monoclonal antibodies for minor RBC antigens are licensed Low cost and quick turn-around time Only viable if minor antigen discrepancy exists between donor/recipient. If patient has positive Direct Coombs’ test, results could be invalid based on reagent used

Under-deployment of extra-large drug-eluting stent: an adapted provisional technique for selected patients with distal lesions in large left main.

In provisional technique, main vessel (MV) drug-eluting stent (DES) diameter is usually selected according to distal MV to reduce carina shift. Proximal optimization technique (POT) is used to expand the DES in the proximal MV. Occasionally, the size discrepancy between left main (LM) and left anterior descending artery (LAD) may be huge and this may cause stent malapposition and poor vessel wall coverage in large-sized LM. Recently, some manufactures designed extra-large DES to treat large vessels. We developed an "adapted" provisional strategy based on under-deployment of extra-large DES in case of major size mismatch between LM and proximal LAD. Bench tests were realized in appropriately designed LM bifurcation model using an extra-large DES (Onyx XL, Medtronic, Santa Rosa, CA, USA). This technique was adopted when such "rare" anatomy was found in our clinical practice. At bench test, Onyx XL 4.5 mm stent reaches 3.8 mm at 5-6 atmospheres, with favorable stent deformation achieved after POT, kissing balloon and re-POT. This technique was performed in 10 patients undergoing unprotected LM stenting with large LM and major mismatch toward LAD. Angiographic success was achieved in all cases and optical coherence tomography assessment was performed in 5 patients revealing optimal stent result. After a follow-up of 557 days (range: 90-1369 days), clinical course was uneventful in all treated patients. Under-deployment of extra-large DES is a technical option that can be considered to optimize the provisional stenting technique in selected patients with major diameter mismatch between large-sized LM and LAD.

Berberine Prolongs Mouse Heart Allograft Survival by Activating T Cell Apoptosis via the Mitochondrial Pathway.

Berberine, which is a traditional Chinese medicine can inhibit tumorigenesis by inducing tumor cell apoptosis. However, the immunoregulatory of effects berberine on T cells remains poorly understood. Here, we first examined whether berberine can prolong allograft survival by regulating the recruitment and function of T cells. Using a major histocompatibility complex complete mismatch mouse heterotopic cardiac transplantation model, we found that the administration of moderate doses (5 mg/kg) of berberine significantly prolonged heart allograft survival to 19 days and elicited no obvious berberine-related toxicity. Compared to that with normal saline treatment, berberine treatment decreased alloreactive T cells in recipient splenocytes and lymph node cells. It also inhibited the activation, proliferation, and function of alloreactive T cells. Most importantly, berberine treatment protected myocardial cells by decreasing CD4+ and CD8+ T cell infiltration and by inhibiting T cell function in allografts. In vivo and in vitro assays revealed that berberine treatment eliminated alloreactive T lymphocytes via the mitochondrial apoptosis pathway, which was validated by transcriptome sequencing. Taken together, we demonstrated that berberine prolongs allograft survival by inducing apoptosis of alloreactive T cells. Thus, our study provides more evidence supporting the potential use of berberine in translational medicine.