Search Results (126)

Somatic mutation patterns observed in cancer genomes are widely used to generate hypotheses about functional relationships among cancer genes and signaling pathways. However, mutation co-occurrence and mutual exclusivity are assessed at multiple levels, including cohorts, bulk specimens, lesions, regions, clones, and individual cells, although each observational level supports a different scope of inference. In this structured narrative review, we clarify these inferential boundaries and distinguish marginal from conditional association, as well as negative association from complete mutual exclusivity. A hypothetical numerical example of Simpson’s reversal illustrates how marginal and conditional associations can differ and why negative association with non-zero overlap should be distinguished from complete mutual exclusivity. We then synthesize evidence from bulk, multi-region, phylogenetic, and single-cell analyses to examine spatial and clonal localization, interclonal cooperation, single-cell error and detection power, and genetic versus non-genetic resistance. We also provide a decision guide for method selection and a staged framework for functional validation. Overall, statistical association, physical localization, and functional interaction are related but distinct inferential targets that require different data, assumptions, and forms of validation. Full article

Chronic hepatitis C virus (HCV) infection is an established risk factor for B-cell lymphoproliferative disorders and represents a paradigmatic model of infection-driven lymphomagenesis. Although direct-acting antivirals have markedly reduced the burden of HCV-related disease, HCV-associated lymphomas continue to occur. Moreover, HCV screening remains incomplete in some geographical areas and healthcare settings, leaving a substantial proportion of infected individuals unaware of their status. This narrative review integrates current evidence on the mechanisms linking chronic HCV infection to mixed cryoglobulinemia and overt B-cell non-Hodgkin lymphoma. HCV lymphotropism and persistent antigenic stimulation could initially promote the selection and expansion of autoreactive B-cell clones, while mixed cryoglobulinemia represents the most informative pre-lymphomatous risk condition. Cytokine-mediated survival signals, particularly those involving B-cell activating factor, reinforce clonal persistence and cooperate with host genetic susceptibility, impaired apoptotic control, and activation-induced cytidine deaminase-mediated genomic instability. The progressive acquisition of somatic driver mutations, copy-number alterations, and epigenetic and transcriptomic changes may enable selected clones to escape functional anergy and become increasingly independent of the original viral stimulus that, in turn, represents an initial trigger of the lymphoproliferative process. Recurrent abnormalities converge on NF-κB, NOTCH, chromatin-regulatory, apoptotic, and cell-cycle pathways, although HCV-associated lymphomas remain molecularly heterogeneous. Emerging microRNA profiles further contribute to the molecular characterization of the transition from chronic infection and cryoglobulinemia to lymphoma. Despite the availability of highly effective antiviral therapies, HCV-associated lymphomagenesis remains clinically relevant and continues to provide an especially informative model for understanding how chronic viral infection can drive human cancer development. Full article

Philadelphia-negative (Ph-negative) myeloproliferative neoplasms (MPNs) include polycythemia vera (PV), essential thrombocythemia (ET), and primary myelofibrosis (PMF), which are clonal hematopoietic disorders caused by somatic gene mutations in the JAK2, CALR, or MPL genes. Mutations activate the JAK–STAT pathway and disrupt NF-κB signaling, leading to a chronic inflammatory state caused by pro-inflammatory cytokines and reactive oxygen species (ROS). This altered microenvironment causes serious clinical features of the disease, such as bone marrow fibrosis, splenomegaly, vascular niche remodeling, and a greater probability of thrombosis or secondary leukemic transformation. Concurrently, MPNs cause both severe immune dysregulation and tumor evasion, as evidenced by progressive lymphopenia, T and B cell exhaustion, Natural Killer cell maturation arrest, and the accumulation of myeloid-derived suppressor cells. Although FDA-approved JAK1/JAK2 inhibitors ruxolitinib, fedratinib pacritinib and momelotinib effectively reduce splenomegaly and symptom burden and have demonstrated survival benefits in clinical trials, their ability to eliminate malignant clones or induce durable disease modification remains limited, and disease progression continues to occur in most patients. Finally, this review assesses the complex immunological dysfunction and chronic inflammatory dysregulation that characterize Ph-negative MPNs, as well as emerging therapeutic strategies, emphasizing the importance of fully understanding these intricate microenvironmental mechanisms for the identification and development of novel precision treatment targets. Full article

The genomic landscape of myelodysplastic syndromes/neoplasms (MDS), a heterogeneous group of myeloid malignancies defined by bone marrow cell dysplasia with ineffective hematopoiesis, includes somatic and, less frequently, germline mutations in hematopoietic stem and progenitor cells, along with chromosomal abnormalities. The latest World Health Organization 2022 classification of myeloid neoplasms, as well as stratification in lower-risk (LR) and higher-risk (HR) MDS using either the Revised International Prognostic Scoring System (IPSS-R) or the Molecular International Prognostic Scoring System (IPSS-M), guide prognostic assessment and risk-adjusted therapy. We report the case of an 81-year-old patient diagnosed with LR-MDS according to IPSS-R that exhibited direct progression to acute myeloid leukemia. The retrospective analysis of paired DNA samples from MDS and leukemic phases, obtained four months apart, using both targeted next-generation sequencing and single nucleotide polymorphism array, indicated swift alterations in the genomic profile, being suggested that the leukemic clone emerged from the clone harboring homozygous TET2 and heterozygous SRSF2 variants that acquired RUNX1, BCOR, BCORL1 likely pathogenic mutations and trisomy 13. By employing IPSS-M for prognostic evaluation at the MDS phase, the patient would have been assigned to the HR-MDS category with a possible benefit from hypomethylating agent therapy. Risk stratification is of pivotal importance in a patient-centered approach to MDS treatment being significantly improved by incorporating the molecular genetic findings. Full article

Somatic cell nuclear transfer (SCNT) in pigs has been a widely used technique for producing gene-edited pigs for biomedical research, yet its wide-spread application through serial cloning remains markedly limited. Unlike in mice, where the serial cloning can be sustained across numerous generations, in pigs it is usually limited to only a few rounds. Specifically, porcine serial cloning has not been reported beyond three consecutive generations in live-born offspring, with blastocyst development rates declining from approximately 4.4% in G1 to 1–5% in G2–G3, and live-birth cloning efficiency (offspring/recipient) dropping sharply with each successive round. Compelling evidence suggests that cumulative epigenetic instability, incomplete embryo genome activation, DNA methylation reprogramming, persistent donor-cell memory, and imprinting disruption collectively erode transcriptional integrity across generations. Although several manipulations, including epigenetic modifiers, transiently improved the early development, they failed to sustain the reprogramming across several generations. Here, we synthesize comparative advances in serial cloning across species and propose that species-specific differences in chromatin plasticity and cytoplasmic reprogramming capacity define a porcine “reprogramming ceiling”. Deciphering and overcoming this barrier will be critical for advancing sustainable livestock engineering, xenotransplantation and translational medicine biotechnology. Full article

Clonal hematopoiesis of indeterminate potential (CHIP) is an age-related condition affecting over 10–20% of individuals older than 70 years, characterized by the expansion of hematopoietic stem cell clones carrying somatic mutations in leukemia-associated driver genes in the absence of overt hematologic disease. Initially recognized as a precursor to hematologic malignancies, CHIP has since been implicated in diverse non-malignant disorders, notably increasing the risk of cardiovascular events by 40%. Recent epidemiological and experimental evidence suggests a potential disease-modifying influence of CHIP in neurodegenerative diseases, particularly Alzheimer’s disease (AD), although findings remain heterogeneous and sometimes contradictory. This review synthesizes recent evidence linking CHIP to AD risk, neuropathology, and disease progression. In this study, we summarize population-based cohort studies reporting a 36 to 54% reduction in the odds of clinical AD among CHIP carriers, alongside emerging data indicating that DNMT3A and TET2 mutations may exert divergent effects on neurodegeneration. Mechanistic insights from experimental models are examined, highlighting the ability of mutated myeloid cells to infiltrate the central nervous system and modulate neuroinflammation and amyloid clearance. We discuss conflicting findings and analyze how CHIP-driven vascular disease and stroke confound neuroprotective signals. We propose that CHIP may differentially influence AD and vascular contributions to cognitive impairment and dementia, shaping mixed dementia phenotypes. Methodological challenges, including survivor bias, competing risks, variable mutation detection thresholds, and incomplete Apolipoprotein E stratification, are discussed. Ultimately, our review clarifies that CHIP is not a simple protective factor, but a complex systemic modulator that reshapes the neurodegenerative and vascular drivers of cognitive decline, necessitating cross-disciplinary neuro-hematology collaboration to establish its role as a novel risk stratificator for improving diagnostic precision and personalizing clinical outcomes in Alzheimer’s disease. Full article

Cutaneous T-cell lymphoma (CTCL) comprises a heterogeneous group of extranodal non-Hodgkin lymphomas. With the publication of the fifth edition of the World Health Organization Classification of Hematolymphoid Tumors, the diagnostic framework for CTCL has shifted from primarily morphologic phenotypes toward an emphasis on molecular drivers. Current research suggests that malignant clones may arise from somatic mutations at the hematopoietic stem cell stage and may follow a continuous hematogenous dissemination model with bidirectional trafficking between the skin and systemic circulation. At the molecular level, genomic instability, often associated with somatic copy-number variations, may promote activation of the janus kinase-signal transducer and activator of transcription (JAK/STAT) signaling pathway through gene-dosage effects. In parallel, chromatin remodeling linked to EZH2 overexpression and reduced special SATB1 expression may support a Th2-polarized program. This phenotype may contribute to epidermal barrier impairment via cytokines such as Interleukins-4 (IL-4) and IL-13, potentially creating conditions permissive for Staphylococcus aureus colonization. Microbial superantigens and exotoxins may further contribute to tumor progression and therapeutic resistance by reinforcing JAK/STAT signaling, particularly STAT3, and reducing CD8+ T-cell–mediated immune surveillance. In the dermis, reprogramming of cancer-associated fibroblasts and polarization of macrophages toward an M2 phenotype may collectively contribute to an immunosuppressive niche. Emerging biomarkers, including CD74, and acquired resistance mechanisms after anti-C-C chemokine receptor 4 therapy further extend the translational relevance of recent pathologic findings. Overall, CTCL evolution appears to be a systemic process shaped by interactions between tumor-intrinsic genetic alterations and the skin microenvironment. Full article

Embryonic stem cells represent a valuable germplasm resource with significant implications for breed conservation, development, and utilization. However, the scarcity of genetic resources in endangered species poses a fundamental constraint on obtaining gametes for embryonic stem cell derivation. Therefore, generating embryonic stem cells from somatic cell nuclear transfer blastocysts offers an optimal alternative for conservation cloning. In this study, we established ApèiJiaza somatic cell nuclear transfer ESCs (APNT-ESCs) from cloned embryos, using ApèiJiaza cattle ear fibroblasts as nuclear donors. APNT-ESCs could be passaged for over 30 generations in vitro, exhibiting high expression of key pluripotency markers, genomic stability, and the ability to form embryoid bodies and differentiate into cell types of all three germ layers. This research established an effective biotechnological framework for the genetic conservation of other endangered species lacking accessible gametes. Full article

Background/Objectives: Clonal evolution is mainly defined based on the appearance or expansion of clones harboring specific somatic mutations and/or cytogenetic abnormalities, whereas few studies have investigated immunophenotypic heterogeneity assessed by flow cytometry and its relationship with disease progression. In this study, flow cytometry immunophenotyping of acute myeloid leukemia (AML) was carried out to identify phenotypic subclones based on antigen expression and to investigate clonal sweep. Methods: A total of 24 patients diagnosed with AML followed at the Hematology and Transplant Center of Salerno were included. Bone marrow or peripheral blood specimens were subjected to flow cytometry immunophenotyping and leukemic cell characterization. Phenotypic profiles were also compared to molecular alterations detected by next-generation sequencing. Results: We found that flow cytometry-defined clonal heterogeneity was more complex than molecular heterogeneity at diagnosis and disease relapse. Flow cytometry enabled the identification of small phenotypic subclones that were not detected by molecular profiling and that, in several cases, expanded over time, consistent with a phenotypic clonal sweep. The presence of small clones was associated with shorter progression-free survival and overall survival. Conclusions: Flow cytometric clonal heterogeneity, especially the presence of small clones (defined by antigen expression from 2 to 30%), may serve as an additional prognostic factor in AML. Immunophenotyping integrated with molecular data may improve risk stratification, enhance measurable residual disease assessment, and contribute to a more personalized disease monitoring strategy. Full article

Pulmonary embolism (PE) is biologically heterogeneous. Despite guideline-directed anticoagulation, a subset of patients develops recurrent venous thromboembolism, persistent exertional limitation, residual perfusion defects, and progression to chronic thromboembolic pulmonary disease (CTEPD) or chronic thromboembolic pulmonary hypertension (CTEPH). Conventional risk factors explain much of the index event but incompletely account for thrombus non-resolution and chronic sequelae. Clonal hematopoiesis of indeterminate potential (CHIP)—the age-associated expansion of hematopoietic clones carrying somatic mutations—defines a measurable thrombo-inflammatory endophenotype that is strongly genotype- and clone-size (variant allele frequency; VAF)-dependent. Across human studies, JAK2-CHIP and TET2-CHIP show the most consistent associations with VTE/PE, whereas isolated DNMT3A-CHIP is frequently neutral, and larger clones tend to confer stronger effects. Mechanistically, CHIP can bias myeloid cells toward inflammasome/IL-1β signaling and endothelial activation, increase monocyte tissue factor activity, and promote immunothrombosis with neutrophil extracellular trap (NET) formation. NET-rich thrombi may adopt a dense fibrin–DNA–histone architecture that resists endogenous fibrinolysis, favoring organization and persistence. CTEPH offers a translational window to interrogate this model because thrombotic material and deep phenotyping are accessible. We synthesize genotype- and VAF-resolved clinical and mechanistic evidence using a structured strength-of-evidence framework and propose a pragmatic phenotyping roadmap with testable predictions for prospective post-PE validation. CHIP testing in PE/CTEPH remains investigational and should not currently change standard care. Full article

Pig somatic cell nuclear transfer (SCNT) has valuable applications in agriculture, biomedicine, and life sciences, yet low cloning efficiency remains a major constraint limiting its application. To systematically investigate factors related to the production efficiency of pig cloning, this study conducted a retrospective analysis of 367,701 SCNT embryos transferred into 2019 surrogate sows over five years, focusing on breeds of donor cells, the season of embryo transfers, and the number of embryos transferred per surrogate. Our data demonstrate that the genetic background of donor cells is a critical determinant. SCNT embryos generated by wild-type (WT) Pietrain and Duroc pigs yielded significantly higher cloning efficiencies compared to those from Large White and Yorkshire pigs. This breed-specific influence was also observed with genetically modified (GM) donor cells. Nevertheless, within the GM groups, GM-Duroc and GM-Yorkshire showed superior efficiency compared to GM-Large White and GM-Bama. Furthermore, Summer was identified as the least favorable season for embryo transfer, with significantly lower pregnancy rates, delivery rates, and cloning efficiency compared to the other seasons. Importantly, we established that transferring 100–150 embryos per recipient optimized cloning efficiency, significantly outperforming groups receiving higher embryo numbers without compromising pregnancy rates, delivery rates, or average litter sizes. Our findings provide valuable guidance for optimizing large-scale SCNT protocols in swine. Full article

Somatic cell nuclear transfer (SCNT) or cloning technology is widely used in agriculture and biomedicine. However, the application of this technology is limited by the low developmental competence of cloned embryos or fetuses, which frequently exhibit abnormal development of trophoblast cells or placentas. The purpose of this study was to investigate the possible causes of the erroneous placental development of SCNT-derived pig fetuses. The placental transcriptomic and lipidomic profiles were compared between 30-day-old SCNT- and artificial insemination (AI)-produced pig fetuses. Differentially expressed lipid metabolites between two groups of placentas were selected to test their effects on porcine trophoblast cell activity. The results showed that SCNT placentas exhibit impaired lipid metabolism and function. The level of a metabolite, lysophosphatidylcholine (LPC), in the glycerophospholipid metabolism pathway was substantially increased in SCNT placentas, compared with AI placentas. The elevation in LPC content may lead to impaired placental development in cloned pig fetuses, as LPC inhibited the proliferation and migration of porcine trophoblast cells. This study discovers a main cause of erroneous development of cloned pig fetuses, which will be beneficial for understanding the regulation of SCNT embryo development, as well as developing new methods to improve the efficiency of pig cloning. Full article

Somatic cell nuclear transfer (SCNT) holds promise for animal cloning but remains limited by low efficiency and phenotypic abnormalities, often attributed to incomplete nuclear reprogramming. This study presents an integrative genomic and epigenomic analysis of cloned buffaloes and their respective donors using long-read Oxford Nanopore sequencing. Our results showed a high degree of genomic similarity between clones and donors, with most variations located in non-coding regions and structural variants (SV) distributions highly correlated at the chromosomal level. Gene and protein level overlap of SV-affected loci revealed 70.9–73.3% gene-level and 69.7–72.5% protein-level similarity. Despite this genetic similarity, DNA methylation analysis identified differentially methylated regions (DMRs), particularly in intergenic and promoter regions. Clones exhibited slightly lower CpG methylation than the donors. The DMRs in donor vs. clone comparisons indicated higher hypomethylated regions than hypermethylated regions. Functional enrichment of DMR-associated genes highlighted pathways linked to mitochondrial function, oxidative phosphorylation, and reproductive processes. Although clones showed moderate genome-wide methylation correlation with donors, key differences in methylation suggest incomplete epigenetic reprogramming. Despite these epigenetic differences, all clones were phenotypically normal and healthy into adulthood. This study offers the first comprehensive SV and methylome profile of SCNT-derived buffaloes and emphasizes the role of epigenetic mechanisms in clone development and health, providing valuable insights to enhance cloning efficiency. Full article

The asynchrony of cytoplasmic and nuclear maturation in cumulus–oocyte complexes (COCs) due to prematurely declining concentrations of cyclic adenosine monophosphate (cAMP) has been shown to result in reduced oocyte developmental competence. The objective of this study was to evaluate the effect of pre-IVM treatment with cAMP modulators for different durations on the developmental potential of equine oocytes used for cloned embryo production. Collected COCs were transferred to cryovials filled with transport medium at 20–22 °C. Within the cryovials, the COCs were either untreated (Control) for 18 h or treated with 50 µM forskolin and 100 µM 3-isobutyl-1-methylxanthine for the first 4 h (Pre-IVM 4 h) or the entire 18 h (Pre-IVM 18 h). Oocytes were then transferred to maturation medium and incubated for a further 22–24 h at 38.5 °C in 5% CO₂ in air. Somatic cell nuclear transfer embryos were then produced using the meiotically mature oocytes and donor cells from six different fibroblast cell lines. The rates of maturation and embryo development did not differ significantly between the groups, though blastocyst formation tended to be inferior in the Pre-IVM 4 h group compared with the Control group (p = 0.06). Of 67 blastocysts produced, 23 were transferred to recipient mares on Day 4 or 5 post-ovulation. Regarding the pregnancy outcomes, no significant differences were found between the groups, and four viable foals were born, each derived from a different donor cell line. The findings expand on those from previous evaluations of this biphasic IVM system, and indicate that the cAMP-modulating treatments exert limited effects under the pre-IVM conditions used here. Full article

Background: In the adaptive immune response of the dromedary (Camelus dromedarius, Camdro), the T cell receptor (TR) repertoire of the gamma–delta (γδ) T cells is unusually diversified both by somatic hypermutation in rearranged TR gamma (TRG) and delta (TRD) genes and by the diversity in sequence and length of the third complementarity-determining region (CDR3) of the TRD chain. Methods: The purpose was to investigate, in the absence of 3D structures, the role of Camdro γδ T cells, focusing on the binding interactions at the interface between the V-gamma and V-delta domains, and in complex with the CD1D, a major histocompatibily class I (MH1)-like glycoprotein presenting lipid antigen in association with B2M. A combination of hypermutated TRG dromedary cDNA clones was paired with TRD clones bearing very long, long, or short CDR3s, all isolated from the spleen of a single animal. Results: The 3D models of the Camdro TRG_TRD/CD1D_B2M complexes were inferred using the Homo sapiens 3D structure and the ImMunoGeneTics (IMGT) numbering for V, C, and G domains, and investigated for binding interactions at the interface of the paired V-gamma_V-delta and at the interface with CD1D. Our results suggest that transcripts with long CDR3s may derive from a population of CD1D-restricted γδ T cells. Both the CD1D G-alpha1-like and G-alpha-2 like domain helices were contacted by both the V-gamma and V-delta CDR-IMGT loops. Conclusions: Our findings further emphasize the similarity between the γδ T cells population we analyzed in Camelus dromedarius and the CD1D-restricted γδ NKT cells in Homo sapiens. Full article