Biology subjects

Ward, D.

Publications and source records attributed to Ward, D..

At least 19 recordsLinked to original sources

bioRxiv2026

How long is the brain perfusable after global ischemia? A systematic review

BackgroundGlobal cerebral ischemia initiates a cascade of pathophysiological changes that progressively impair subsequent perfusion of brain tissue. Some authors have proposed that adequate cerebral perfusion becomes impossible after approximately 10-30 minutes of global ischemia. However, the extant evidence base for this threshold and its variation across studies have not yet been systematically examined. ObjectiveTo synthesize the literature on post-ischemic cerebral perfusion success as a function of ischemia duration. MethodsWe searched PubMed (February 11, 2026) for studies of global cerebral ischemia in animal or human models that reported quantitative or categorical measures of perfusion quality. Eligible studies included those assessing perfusion via restoration of blood flow, via external perfusion of non-blood solutions, and/or via tracer injection following reperfusion. Studies of focal ischemia were excluded. Data extracted included species, ischemia duration, temperature during ischemia, ischemia model, perfusate type, and perfusion quality assessment method. The perfusion quality outcome was operationalized as either the average percentage of brain tissue perfused or the percentage of brains in a group that were adequately perfused. Study quality was assessed using a custom domain-specific checklist. The review protocol was preregistered on the Open Science Framework (https://osf.io/2qm3w). ResultsWe included 60 studies with 192 study arms reporting on the perfusion of the brains of rabbits, rats, pigs, dogs, cats, and humans. Studies differed in the model of ischemia, the perfusate, the perfusion parameters, the quality assessment methods, and other factors. Longer ischemia was associated with lower perfusion quality, but the relationship appeared to be a gradual decline rather than a sharp threshold. Additionally, the variability across studies was large, and some studies have found that at least partial perfusion is possible after longer periods. Within-study dose-response curves were more consistent than the pooled cross-study pattern. ConclusionsHow long the brain remains perfusable after circulatory arrest has not yet been definitively established. On average, perfusion quality clearly declines rapidly as the duration of global cerebral ischemia increases. However, some studies, often using interventions such as hypothermia or vasopressors, have reported at least partial perfusion of the brain even after 30 or 60 minutes of ischemia. Moreover, at least partial perfusion has been reported in human brain banking studies after postmortem intervals of several hours or days in some donors. Limitations of this review include substantial heterogeneity in study methods and outcome measures, which precluded formal meta-analysis. Future research may benefit from more thorough and precise measures of perfusion quality.

neuroscience↗

bioRxiv2026

immunoPETE: A DNA-based integrated B-cell and T-cell receptor profiling platform

The vast diversity of B and T cell receptors generated through the recombination of Variable (V), Diversity (D), and Joining (J) gene segments plays a critical role in adaptive immunity. Profiling immune repertoires at the DNA level provides a robust and stable approach to capture the clonal composition of these receptors. immunoPETE is an assay designed to target recombined human T-cell Receptor Beta (TRB), T-cell Receptor Delta (TRD), and Immunoglobulin Heavy (IGH) chain genes directly from genomic DNA. Simultaneous profiling of B and T cell receptor chains in a single reaction provides internally normalized clone counts and facilitates the study of B-T cell interactions. Full-length amplicon consensus sequences representative of original template DNA molecules are accurately reconstructed using Unique Molecular Identifiers (UMIs). An in-house pipeline compiles VDJ rearrangements from the Complementarity-Determining Region 3 (CDR3) of TRB, TRD and IGH chains into comprehensive readouts at cell-level resolution. In this study, we describe the immunoPETE end-to-end workflow, followed by a comprehensive benchmarking of its performance in adaptive immune profiling. Where applicable, we used both natural and contrived samples and characterized the assays accuracy, linearity, and reproducibility across several metrics: retrieving CDR3 sequences, determining B and T cell ratios, total cell count, yield, fraction of functional rearrangements, clonal diversity, composition of dominant clones, pairwise similarity, and V/J gene usage frequencies. Furthermore, we assessed its quantitative limits concerning the total number of lymphocytes and the detection of rare clones. As an example of its applications, we show that adding immune biomarkers extracted from immunoPETE data to clinical factors improves prediction of progression-free survival in a cohort of non-muscle invasive bladder cancer (NMIBC) patients. Finally, we discuss the broad applications of immunoPETE in the study of aging, cancers, infections, and autoimmune disorders with reference to select published studies.

immunology↗

bioRxiv2026

Vicarious trauma primes innate immunity and reconfigures human brain networks

Vicarious trauma--witnessing serious harm to others--produces systemic neuroimmune effects in animal models. However, its brain, autonomic, and immune consequences in humans remain unclear. We scanned 88 participants with fMRI during a naturalistic movie-viewing paradigm depicting either industrial cruelty to farm animals (Vicarious Trauma, VT) or positive human-animal interactions (Vicarious Community, VC) with concurrent autonomic physiology and gene expression in circulating monocytes. VT, compared to VC, evoked sustained sympathetic arousal lasting 40 minutes post-viewing and primed innate immune cells to elicit elevated pro-inflammatory (upregulating IL1B and CXCL8) and reduced anti-inflammatory (TGFB1) responses. VT reduced integration in a frontoparietal control system and increased coupling between a salience/action-mode system with paralimbic, basal ganglia, and sensorimotor regions. Disrupted negative coupling between frontoparietal/default-mode and hippocampal systems predicted VT-related CXCL8 increases. These findings shed light on how the brain translates vicarious trauma into the priming of innate immunity.

neuroscience↗

bioRxiv2025

Cyclic-di-GMP controls Type III effector export and symptom development in Pseudomonas syringae infections via the export ATPase HrcN

Pseudomonas syringae is a destructive bacterial pathogen that infects a wide variety of plants. Following apoplastic entry, P. syringae uses its type 3 injectisome (T3I) to secrete host-specific effectors into the cytoplasm, enabling tissue wetting and immune suppression and leading to bacterial proliferation, chlorosis and necrosis. P. syringae strains encode dozens of highly specialised effectors, whose composition defines strain specificity and host range. Effective plant infection depends on the tight temporal and hierarchical control of effector delivery through the T3I. Effector secretion is driven by HrcN, an ATPase complex that interacts with the base of the T3I and is essential for plant infection. HrcN binds specifically to the bacterial signalling molecule cyclic-di-GMP, although the impact of binding on T3I function and P. syringae virulence is currently unknown. To address this, we examined the influence of mutating the predicted cyclic-di-GMP-hrcN binding site on plant infection and effector secretion. Despite maintaining effective bacterial proliferation in Arabidopsis thaliana leaves, two hrcN mutants showed severely compromised disease symptoms, a phenotype linked to reduced translocation of a specific subset of T3I-effectors, with HopAA1-2 particularly important for symptom development. We propose that cyclic-di-GMP binding may represent a novel regulatory mechanism for effector secretion during bacterial infections.

microbiology↗

bioRxiv2025

Differences in brain activity during sentence repetition in people who stutter: a combined analysis of four fMRI studies

Our understanding of the neural correlates of developmental stuttering benefits from the use of functional MRI (fMRI) during speech production. Despite two decades of research, however, we have reached little consensus. In the current study, we analysed pooled fMRI data from four different studies that used the same sentence reading task and methodological approach. The combined sample included 56 adolescents and adults who stutter and 53 demographically matched typically fluent controls. A sparse-sampling design was used in each study, in which participants spoke during the silent period between measurements of brain activity. Sentence reading evoked activity in both groups across frontal and temporal regions bilaterally. At statistical thresholds corrected for family-wise error, there were no significant group differences. An uncorrected threshold was applied to explore group differences in areas previously identified in earlier fMRI studies on stuttering. People who stutter (PWS) showed greater activity compared with controls in right frontal pole, right anterior insula extending to frontal operculum, left planum temporale, and midbrain, at the level of red nucleus. In contrast, PWS showed lower activity in left superior frontal sulcus, subgenual medial prefrontal cortex, right anterior temporal lobe, and portions of inferior parietal lobe bilaterally including the angular gyrus on the left. Despite pooling data across multiple studies to achieve a relatively large sample, group differences in regions involved in speech-motor control only emerged at an uncorrected voxel-wise threshold. Some of these findings align with previous fMRI studies, such as increased activity in the right anterior insular cortex.

neuroscience↗

bioRxiv2025

Schistosome exposure and diet induced effects on candidate immune gene expression in an African snail vector

Schistosoma mansoni is a parasitic helminth that is vectored through freshwater snails. While the anti-schistosome defense of the South American snail, Biomphalaria glabrata, is well studied, little is known about the immune response of the African snail, Biomphalaria sudanica. We measured expression of five candidate immune genes in B. sudanica 8, 24, and 72 hours post-exposure to S. mansoni using reverse transcription quantitative PCR. Expression patterns of resistant snails were compared to susceptible snails and those sham exposed. We also assessed how diet (lettuce vs. pellet) affected expression of three genes, given prior findings that pellet-fed snails were more susceptible to S. mansoni. Results indicated that resistant snails constitutively expressed higher levels of superoxide dismutase 1 (SOD1) than susceptible snails, consistent with expression patterns of resistant B. glabrata. Parasite-induced expression occurred at 8 hours in SOD1, biomphalysin, thioester protein 1 (TEP1), and granulin (GRN); however, for biomphalysin and TEP1, induced expression was only detected for susceptible snails. At 24 hours, biomphalysin expression increased in exposed resistant snails, and at 72 hours, all exposed snails decreased biomphalysin expression compared to controls. Parasite-induced expression of SOD1, biomphalysin, TEP1, and GRN supports the hypothesis that these genes play a role in B. sudanica anti-schistosome defense, however increased expression does not necessarily yield clearance of S. mansoni. SOD1 expression was higher in lettuce-fed snails at 8 and 24 hours, consistent with their greater resistance. Together, these results demonstrate the conserved and unique aspects of the B. sudanica anti-schistosome response.

microbiology↗

bioRxiv2025

Microbiome functional gene pathways predict cognitive performance in older adults with Alzheimers disease

Disturbances in the gut microbiome is increasing correlated with neurodegenerative disorders, including Alzheimers Disease. The microbiome may in fact influence disease pathology in AD by triggering or potentiating systemic and neuroinflammation, thereby driving disease pathology along the "microbiota-gut-brain-axis". Currently, drivers of cognitive decline and symptomatic progression in AD remain unknown and understudied. Changes in gut microbiome composition may offer clues to potential systemic physiologic and neuropathologic changes that contribute to cognitive decline. Here, we recruited a cohort of 260 older adults (age 60+) living in the community and followed them over time, tracking objective measures of cognition, clinical information, and gut microbiomes. Subjects were classified as healthy controls or as having mild cognitive impairment based on cognitive performance. Those with a diagnosis of Alzheimers Diseases with confirmed using serum biomarkers. Using metagenomic sequencing, we found that relative species abundances correlated well with cognition status (MCI or AD). Furthermore, gene pathways analyses suggest certain microbial metabolic pathways to either be correlated with cognitive decline or maintaining cognitive function. Specifically, genes involved in the urea cycle or production of methionine and cysteine predicted worse cognitive performance. Our study suggests that gut microbiome composition may predict AD cognitive performance.

microbiology↗

bioRxiv2025

Progesterone regulation of cervix ripening in preterm and term birth

The cervix functions both as gatekeeper barrier to maintain pregnancy and virtually vanish for birth at term in mammals. The period of remodeling well-before term is characterized by an inflammatory process associated with reduced cell nuclei density and cross-linked collagen, as well as increased density of resident macrophages in cervix stroma. Contemporarily, progesterone (P4) is at or near peak concentrations in maternal circulation. The functional or actual loss of response to P4 is thought to drive the process that enhances uterine contractile activity for labor and parturition at term. The objective of the present study was to determine if actual or functional loss of P4 regulated cytomorphological characteristics associated with prepartum cervix ripening at term and with preterm birth. On day 16 of pregnancy. Ovaries were removed to eliminate the main source of P4 production and a silastic capsule implanted (with vehicle or P4, Ovx or Ovx+P4, respectively). Controls received a vehicle-filled capsule, while a P4 capsule was implanted into an addition group of Intact mice to ensure sustained concentrations throughout pregnancy (Intact+P4). Pups were born in controls at term (days 19-20 postbreeding), but deliveries were preterm in Ovx mice within 24h (day 17). In the Ovx+P4 group, births were delayed to term and post-term in most Intact+P4 mice with adverse pregnancy outcomes commonplace. Characteristics of cell nuclei and degradation of cross-linked collagen were advanced with preterm birth in Ovx mice compared to controls that gave birth by at term. Treatment of Ovx mice with P4 blocked preterm birth, but parturition was complicated by dystocia. In addition, P4 given to ovary-intact mice sustained peak pregnancy concentrations, but had minimal effects on cytoarchitecture of the prepartum cervix stroma except term birth was forestalled with dystocia and fetal morbidity. Density of resident macrophages in the cervix stroma in term Ovx+P4 mice was reduced along with area of macrophage stain versus postpartum controls. Thus, analyses of cervix cellular cytoarchitecture provided useful biomarkers of local inflammation to assessment the ripening process for preterm and term parturition. Collectively, findings suggest a functional loss of prepartum cervix responses to progesterone are part of a final common mechanism for parturition across mammals. SummaryLoss of response to progesterone withdrawal is associated with cervix ripening while some cytoarchitectural characteristics of remodeling are regulated to block preterm birth and dystocia at term

physiology↗

bioRxiv2025

N,N-dimethylacetamide blocks inflammation-induced preterm birth and remediates maternal systemic immune responses

The common excipient, N,N-dimethylacetamide (DMA), prevents imminent endotoxin-induced preterm birth in mice. The present study hypothesized that DMA forestalls preterm birth to term (defined as day 18.5 or later) by attenuating bacterial endotoxin lipopolysaccharide (LPS)-induced maternal systemic inflammatory responses and cervix remodeling. Accordingly, LPS (i.p.) on day 15 postbreeding stimulated preterm delivery within 24 h while mice treated with DMA 2 h preceding and 9 h following LPS administration remained pregnant, comparable to saline and DMA controls, to deliver viable pups at term. Irrespective of LPS or DMA+LPS treatment, maternal plasma pro- and anti-inflammatory cytokines on day 15.5 (12 h post-LPS) increased 10-fold compared to baseline concentrations in controls. On day 16 of pregnancy, plasma concentrations of G-CSF and TNF were reduced in the prepartum LPS+DMA group compared to those in postpartum mice given LPS. By day 18 of pregnancy, all cytokines returned to baseline - equivalent to low systemic levels throughout the study in saline and DMA controls that gave birth at term. In addition, maternal plasma progesterone declined within 12 h in prepartum LPS-treated mice to postpartum concentrations on day 16. Although a similar transient decrease occurred by 12 h in DMA+LPS mice, plasma progesterone returned to baseline concentrations in controls. Contemporaneously, the progression of prepartum cervix remodeling leading to preterm delivery was acutely forestalled by DMA without impeding birth at term. These findings support the hypothesis that DMA not only prevents inflammation-driven preterm birth, but rescues pregnancy for birth to occur at term. The results raise the possibility that maternal signals can forecast risk of preterm birth while selective suppression of systemic inflammation can mitigate adverse pregnancy outcomes. SummaryDMA rescues pregnancy from inflammation-induced preterm birth through a progesterone-independent mechanism that involves multiple proinflammatory cytokines in maternal circulation. Supported by NIH Grant 1R16GM145586 and the Ines Mandl Research Foundation

pharmacology and toxicology↗

bioRxiv2024

Exploring mechanisms of change in a Southern Ocean fishery with a co-produced network model

A key challenge in planning long-term fisheries sustainability is overcoming uncertainties in predicted changes in target populations and catch rates in response to climate change and fishery trends. We combine transdisciplinary knowledge co-production and qualitative network modelling to advance system understanding and elucidate likely responses of a Patagonian toothfish fishery to future change. We co-developed a model of the Kerguelen Plateau biophysical-socioeconomic system with knowledge holders from industry and science; the first whole-of-system qualitative network model of intermediate complexity for this region. We present new approaches for dealing with uncertainty in network structure, and for investigating how perturbations propagate through a network. Using these tools, we found multiple potential pathways of decline for toothfish population and catch, but also some possible mechanisms of increase dependent on magnitude of certain effects. Our results highlight critical information gaps, including the potential role of scavenging benthos in fishery-ecosystem interactions and likely changes in the prey-field in response to warmer water, that require filling to improve predictions for toothfish populations and catch and forward planning for the fishery.

ecology↗

bioRxiv2024

No Evidence of Altered Language Laterality in People Who Stutter across Different Brain Imaging Studies of Speech and Language

A long-standing neurobiological explanation of stuttering is the incomplete cerebral dominance theory, which refers to competition between two hemispheres for "dominance" over handedness and speech, causing altered language lateralisation. Renewed interest in these ideas came from brain imaging findings in people who stutter (PWS) of increased activity in the right hemisphere during speech production or of shifts in activity from right to left when fluency increased. Here, we revisited this theory using functional MRI data from children and adults who stutter, and typically fluent speakers (119 participants in total) during four different speech and language tasks: overt sentence reading, overt picture description, covert sentence reading and covert auditory naming. Laterality indices (LIs) were calculated for the frontal and temporal lobes using the LI toolbox running in Statistical Parametric Mapping. We also repeated the analyses with more specific language regions, namely the pars opercularis (Brodmann Area 44) and pars triangularis (Brodmann Area 45). Laterality indices in PWS and typically fluent speakers (TFS) did not differ and Bayesian analyses provided moderate to anecdotal levels of support for the null hypothesis (i.e., no differences in laterality in PWS compared with TFS). The proportions of the PWS and TFS who were left lateralised or had atypical rightwards or bilateral lateralisation did not differ. We found no support for the theory that language laterality is reduced or differs in PWS compared with TFS.

neuroscience↗

bioRxiv2024

Microbiota encoded fatty-acid metabolism expands tuft cells to protect tissues homeostasis during Clostridioides difficile infection in the large intestine

Metabolic byproducts of the intestinal microbiota are crucial in maintaining host immune tone and shaping inter-species ecological dynamics. Among these metabolites, succinate is a driver of tuft cell (TC) differentiation and consequent type 2 immunity-dependent protection against invading parasites in the small intestine. Succinate is also a growth enhancer of the nosocomial pathogen Clostridioides difficile in the large intestine. To date, no research has shown the role of succinate in modulating TC dynamics in the large intestine, or the relevance of this immune pathway to C. difficile pathophysiology. Here we reveal the existence of a three-way circuit between commensal microbes, C. difficile and host epithelial cells which centers around succinate. Through selective microbiota depletion experiments we demonstrate higher levels of type 2 cytokines leading to expansion of TCs in the colon. We then demonstrate the causal role of the microbiome in modulating colonic TC abundance and subsequent type 2 cytokine induction using rational supplementation experiments with fecal transplants and microbial consortia of succinate-producing bacteria. We show that administration of a succinate-deficient Bacteroides thetaiotaomicron knockout ({Delta}frd) significantly reduces the enhanced type 2 immunity in mono-colonized mice. Finally, we demonstrate that mice prophylactically administered with the consortium of succinate-producing bacteria show reduced C. difficile-induced morbidity and mortality compared to mice administered with heat-killed bacteria or the vehicle. This effect is reduced in a partial tuft cell knockout mouse, Pou2f3+/-, and nullified in the tuft cell knockout mouse, Pou2f3-/-, confirming that the observed protection occurs via the TC pathway. Succinate is an intermediary metabolite of the production of short-chain fatty acids, and its concentration often increases during dysbiosis. The first barrier to enteric pathogens alike is the intestinal epithelial barrier, and host maintenance and strengthening of barrier integrity is vital to homeostasis. Considering our data, we propose that activation of TC by the microbiota-produced succinate in the colon is a mechanism evolved by the host to counterbalance microbiome-derived cues that facilitate invasion by intestinal pathogens.

immunology↗

bioRxiv2023

Improved science communication and student gains from an undergraduate biomedical research experience

Summer research experiences expose undergraduate students to biomedical research in a laboratory or clinical setting, but often do not incorporate formal learning on scientific communication. Proper written and oral communication of science is essential to succeed in biomedical fields. This study examined whether participation in a science communication series (SCS) would increase gains in research and written communication abilities among students participated in the University of Pittsburgh School of Medicine Department of Pediatrics Summer Research Internship Program. Surveys were administered at the beginning and end of the program to evaluate their summer undergraduate research experience (SURE). Positive personal and professional gains in research and communication skills were identified through participation in both SURE and SCS. Participation in the SCS also significantly improved the quality and presentation of research abstracts. Focused learning in science communication during SUREs would improve undergraduate students personal and professional abilities in biomedical research and medicine.

scientific communication and education↗

bioRxiv2023

DNA storage in thermoresponsive microcapsules for repeated random multiplexed data access

Owing to its longevity and extremely high information density, DNA has emerged as an attractive medium for archival data storage. Scalable parallel random access of information is a desirable property of any storage system. For DNA-based storage systems, however, this yet has to be robustly established. Here we develop thermoconfined PCR, a novel method that enables multiplexed, repeated random access of compartmentalized DNA files. Our strategy is based on stable localization of biotin-functionalized oligonucleotides inside microcapsules with temperature-dependent membrane permeability. At low temperatures, microcapsules are permeable to enzymes, primers, and amplified products, while at high temperatures membrane collapse prevents molecular crosstalk during amplification. We demonstrate that our platform outperforms non-compartmentalized DNA storage with respect to repeated random access and reducing amplification bias during multiplex PCR. Using fluorescent sorting, we additionally demonstrate sample pooling and data retrieval by barcoding of microcapsules. Our thermoresponsive microcapsule technology offers a scalable, sequence-agnostic approach for repeated random access of archival DNA files.

synthetic biology↗

bioRxiv2022

Evolved bacterial resistance to the chemotherapy gemcitabine modulates its efficacy

Drug metabolism by the microbiome can influence anti-cancer treatment success. We previously suggested that chemotherapies with antimicrobial activity can select for adaptations in bacterial drug metabolism that can inadvertently influence the hosts chemoresistance. We demonstrated that evolved resistance against fluoropyrimidine chemotherapy lowered its efficacy in worms feeding on drug-evolved bacteria (Rosener et al., 2020). Here we examine a model system that captures local interactions that can occur in the tumor microenvironment. Gammaproteobacteria colonizing pancreatic tumors can degrade the nucleoside-analog chemotherapy gemcitabine and, in doing so, can increase the tumors chemoresistance. Using a genetic screen in Escherichia coli, we mapped all loss-of-function mutations conferring gemcitabine resistance. Surprisingly, we found that one third of resistance mutations increase or decrease bacterial drug breakdown and therefore can either lower or raise the gemcitabine load in the local environment. Experiments in three E. coli strains revealed that evolved adaptation converged to inactivation of the nucleoside permease NupC, an adaptation that increased the drug burden on co-cultured cancer cells. The two studies provide complementary insights on the potential impact of microbiome adaptation to chemotherapy by showing that bacteria-drug interactions transpire locally and systemically and can influence chemoresistance in the host.

systems biology↗

bioRxiv2022

Functional traits trade-offs define plant population stability worldwide

O_LIEcological theory posits that temporal stability patterns in plant populations are associated with differences in species ecological strategies. However, empirical evidence is lacking about which traits, or trade-offs, underlie species stability, specially across different ecosystems. C_LIO_LITo address this, we compiled a global collection of long-term permanent vegetation records (>7000 plots from 78 datasets) from a wide range of habitats and combined this with existing trait databases. We tested whether the observed inter-annual variability in species abundance (coefficient of variation) was related to multiple individual traits and multivariate axes of trait variations (PCoA axes). C_LIO_LIWe found that species with greater leaf dry matter content and seed mass were consistently more stable over time (lower variability in species abundance) although other leaf traits played a significant role as well, albeit weaker. Using multivariate axes did not improve predictions by specific traits. C_LIO_LIOur results confirm existing theory, providing compelling empirical evidence on the importance of specific traits, which point at ecological trade-offs in different resource use and dispersal strategies, on the stability of plant populations worldwide. C_LI

ecology↗

bioRxiv2022

Doctoring Direct-to-Consumer Genetic Tests with DNA Spike-Ins

Direct-to-consumer (DTC) genetic testing companies have provided personal genotyping services to millions of customers. Customers mail saliva samples to DTC service providers to have their genotypes analyzed and receive back their raw genetic data. Both consumers and the DTC companies use the results to perform ancestry analyses, relative matching, trait prediction, and estimate predisposition to disease, often relying on genetic databases composed of the data from millions of other DTC-genotyped individuals. While the digital integrity risks to this type of data have been explored, we considered whether data integrity issues could manifest upstream of data generation through physical manipulation of DNA samples themselves, for example by adding synthetic DNA to a saliva sample ("spiked samples") prior to sample processing by a DTC company. Here, we investigated the feasibility of this scenario within the standard DTC genetic testing pipeline. Starting with the purchase of off-the-shelf DTC genetic testing kits, we found that synthetic DNA can be used to precisely manipulate the results of saliva samples genotyped by a popular DTC genetic testing service and that this method can be used to modify arbitrary single nucleotide polymorphisms (SNPs) in multiplex to create customized doctored genetic profiles. This capability has implications for the use of DTC-generated results and the outcomes of their downstream analyses.

genetics↗

bioRxiv2021

Electrical signature of heterogeneous human mesenchymal stem cells

Human mesenchymal stem cells (hMSCs) have gained traction in transplantation therapy due to their immunomodulatory, paracrine, immune-evasive, and multipotent differentiation potential. Given the heterogeneous nature of hMSCs, therapeutic treatments and robust in vivo and in vitro experiments require additional biomarkers to ensure reproducibility when using these stem cells. In this work, we utilized dielectrophoresis (DEP), a label-free electrokinetic phenomenon, to investigate and quantify the heterogeneity of hMSCs derived from the bone marrow (BM) and adipose tissue (AD). Through computer simulation, we identified that the transient slope of the DEP force spectra can be used as a metric of heterogeneity. The electrical properties of BM-hMSCs were compared to homogeneous mouse fibroblasts (NIH-3T3), human fibroblasts (WS1), and human embryonic kidney cells (HEK-293). BM-hMSCs DEP profile was most different from HEK-293 cells. We compared the DEP profiles of BM-hMSCs and AD-hMSCs and found they have similar membrane capacitances, differing cytoplasm conductivity, and transient slopes. Inducing both populations to differentiate into adipocyte and osteocyte cells revealed they behave differently in response to differentiation-inducing cytokines. Histology and RT-qPCR analyses of the differentiation-related genes revealed differences in heterogeneity between BM-hMSCs and AD-hMSCs. The differentiation profiles correlate well with the DEP profiles developed and indicate that these BM-hMSCs have higher differentiation potential than AD-hMSCs. Our results demonstrate using DEP, membrane capacitance, cytoplasm conductivity, and transient slope can uniquely characterize the inherent heterogeneity of hMSCs to guide robust and reproducible stem cell transplantation therapies.

bioengineering↗