Biology subjects

Velazquez, J.

Publications and source records attributed to Velazquez, J..

7 recordsLinked to original sources

bioRxiv2025

Differentiating the Roles of Metabolic Similarity and Ionic Coupling in Determining the Beta Hub Cell Phenotype

Pancreatic beta cells regulate circulating glucose levels by releasing insulin. Beta cells transduce elevated blood glucose into electrical activity through an intrinsic cascade of metabolic and electrophysiologic responses. These responses are synchronized across the electrically connected network of beta cells, such that insulin release is also relatively synchronous. Despite their coordinated behavior, individual beta cells exhibit significant functional heterogeneity. This heterogeneity is thought to provide cells with specific functional phenotypes the ability to control the activity of the broader network. Hub cells, identified by their synchronized [Ca2+] activity, are one such functional subpopulation, and are believed to orchestrate the second, oscillatory phase of insulin release. However, it remains unclear whether the cell-autonomous characteristics of hub cells, such as their metabolic activity and electrophysiologic properties, are more important than their network characteristics (i.e. gap junctional coupling) for their ability to influence broader network activity. In this study, we investigate the roles of intrinsic metabolic and electrophysiologic properties and ionic coupling in determining the beta hub cell phenotype. Using a computational islet model of 1,000 beta cells, our analysis revealed that both intrinsic metabolic properties and structural coupling via gap junctions are crucial for determining the hub cell phenotype. After investigating the intrinsic coupling conductance of neighboring cells and the number of structural (direct electrical) links as independent contributors to the hub cells local electrical coupling, we find that the number of cells to which a beta cell is directly coupled may be a key determinant of its propensity to serve as a hub cell in the model. As predicted for this subpopulation, we also demonstrate that decoupling hub cells impairs the functional connectivity of the entire network. Our findings indicate the importance of both autonomous cellular dynamics and non-autonomous structural coupling for the hub cell phenotype. These insights help build a fundamental understanding of hub cells, which in turn may contribute to identifying potential approaches to preserve or improve beta cell function and thereby manage the progression of diabetes.

bioengineering↗

bioRxiv2023

Genetically engineering endothelial niche in human kidney organoids enables multilineage maturation, vascularization, and de novo cell types

Vascularization plays a critical role in organ maturation and cell type development. Drug discovery, organ mimicry, and ultimately transplantation in a clinical setting thereby hinges on achieving robust vascularization of in vitro engineered organs. Here, focusing on human kidney organoids, we overcome this hurdle by combining an inducible ETS translocation variant 2 (ETV2) human induced pluripotent stem cell (iPSC) line, which directs endothelial fate, with a non-transgenic iPSC line in suspension organoid culture. The resulting human kidney organoids show extensive vascularization by endothelial cells with an identity most closely related to endogenous kidney endothelia. Vascularized organoids also show increased maturation of nephron structures including more mature podocytes with improved marker expression, foot process interdigitation, an associated fenestrated endothelium, and the presence of renin+ cells. The creation of an engineered vascular niche capable of improving kidney organoid maturation and cell type complexity is a significant step forward in the path to clinical translation. Furthermore, this approach is orthogonal to native tissue differentiation paths, hence readily adaptable to other organoid systems and thus has the potential for a broad impact on basic and translational organoid studies. Translational StatementDeveloping therapies for patients with kidney diseases relies on a morphologically and physiologically representative in vitro model. Human kidney organoids are an attractive model to recapitulate kidney physiology, however, they are limited by the absence of a vascular network and mature cell populations. In this work, we have generated a genetically inducible endothelial niche that, when combined with an established kidney organoid protocol, induces the maturation of a robust endothelial cell network, induces a more mature podocyte population, and induces the emergence a functional renin population. This advance significantly increases the clinical relevance of human kidney organoids for etiological studies of kidney disease and future regenerative medicine strategies. Graphical AbstractGenetically engineered endothelial niche induces mature cell populations in human kidney organoids O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=100 SRC="FIGDIR/small/542848v1_ufig1.gif" ALT="Figure 1"> View larger version (35K): org.highwire.dtl.DTLVardef@1b129d4org.highwire.dtl.DTLVardef@313ae3org.highwire.dtl.DTLVardef@111c91org.highwire.dtl.DTLVardef@16a8d45_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioengineering↗

bioRxiv2022

Platform-Agnostic CellNet (PACNet) enables cross-study meta-analysis of cell fate engineering protocols

The optimization of cell fate engineering protocols requires evaluating their fidelity, efficiency, or both. We previously adopted CellNet, a computational tool to quantitatively assess the transcriptional fidelity of engineered cells and tissues as compared to their in vivo counterparts based on bulk RNA-Seq. However, this platform and other similar approaches are sensitive to experimental and analytical aspects of transcriptomics methodologies. This makes it challenging to capitalizing on the expansive, publicly available sets of transcriptomic data that reflect the diversity of cell fate engineering protocols. Here, we present Platform-Agnostic CellNet (PACNet), which extends the functionality of CellNet by enabling the assessment of transcriptional profiles in a platform-agnostic manner, and by enabling the comparison of user-supplied data to panels of engineered cell types from state-of-the-art protocols. To demonstrate the utility of PACNet, we evaluated a range of cell fate engineering protocols for cardiomyocytes and hepatocytes. Through this analysis, we identified the best-performing methods, characterized the extent of intra-protocol and inter-lab variation, and identified common off-target signatures, including a surprising neural and neuroendocrine signature in primary liver-derived organoids. Finally, we made our tool accessible as a user-friendly web application that allows users to upload their own transcriptional profiles and assess their protocols relative to our database of reference engineered samples. Highlights* The development of Platform-Agnostic CellNet (PACNet) that classifies engineered cell populations from transcriptome data regardless of profiling method or transcript abundance estimation method * PACNet enables cross-study comparisons of cell fate engineering protocols * Comparison of cardiomyocyte engineering protocols emphasizes metabolic selection as a key step in achieving a strong cardiomyocyte fate. * PACNet identifies an unexpected off-target neural and neuroendocrine signature in primary liver-derived organoids. eTOC BlurbCahan and colleagues created a computational resource, PACNet, which evaluates the fidelity of cell engineering expression profiles in a platform-agnostic manner to facilitate cross-protocol benchmarking. Examining state-of-the-field cardiomyocyte and hepatocyte derivation protocols, they identified that two techniques in cardiomyocyte engineering best increase cardiac identity and that an off-target neural/neuroendocrine signature in primary liver-derived organoids may reflect a cholangiopathic signature. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=116 SRC="FIGDIR/small/506886v1_ufig1.gif" ALT="Figure 1"> View larger version (32K): org.highwire.dtl.DTLVardef@e4e985org.highwire.dtl.DTLVardef@9f485borg.highwire.dtl.DTLVardef@1475d0borg.highwire.dtl.DTLVardef@2d8c31_HPS_FORMAT_FIGEXP M_FIG C_FIG

developmental biology↗

bioRxiv2020

Effectiveness of fusion peptide-based vaccine TT-P0 on the dynamics of salmon lice (Lepeophtheirus salmonis) infection in Atlantic salmon (Salmo salar L.)

Infection with parasitic copepod salmon louse Lepeophtheirus salmonis, represents one of the most important limitations to sustainable Atlantic salmon (Salmo salar L.) farming today in the North Atlantic region. The parasite exerts negative impact on health, growth and welfare of farmed fish as well as impact on wild salmonid populations. It is therefore central to ensure continuous low level of salmon lice with the least possible handling of the salmon and drug use. To address this, vaccination is a cost-effective and environmentally friendly control approach. In this study, efficacy of a vaccine candidate, containing a peptide derived from ribosomal protein P0, was validated post infestation with L. salmonis, at the lab-scale. The sampling results showed good potential of the vaccine candidate when administered intraperitoneally in the host, in reducing the ectoparasite load, through reduction of adult female lice counts and fecundity and with greater presumptive effect in F1 lice generation. The sampling results correlated well with the differential modulation of pro-inflammatory, Th1, Th2 and T regulatory mediators at the transcript level at different lice stages. Overall, the results supports the effectiveness of the vaccine candidate in controlling salmon lice infestation load. However, further validation is necessary under field conditions.

immunology↗

bioRxiv2019

C. difficile-associated antibiotics prime the host for infection by a microbiome-independent mechanism.

The most clinically relevant risk factor for Clostridioides difficile-associated disease (CDAD) is recent antibiotic treatment. Though most broad-spectrum antibiotics significantly disrupt the structure of the gut microbiota, only particular ones increase CDAD risk, suggesting additional factors might increase the risk from certain antibiotics. Here we show that commensal-independent effects of antibiotics collectively prime an in vitro germ-free human gut for CDAD. We found a marked loss of mucosal barrier and immune function with CDAD-associated antibiotic pretreatment distinct from pretreatment with an antibiotic unassociated with CDAD, which did not reduce innate immune or mucosal barrier functions. Importantly, pretreatment with CDAD-associated antibiotics sensitized mucosal barriers to C. difficile toxin activity in primary cell-derived enteroid monolayers. These data implicate commensal-independent host changes in the increased risk of CDAD with specific antibiotics. Our findings are contrary to the previously held belief that antibiotics allow for CDAD solely through disruption of the microbiome. We anticipate this work to suggest potential avenues of research for host-directed treatment and preventive therapies for CDAD, and to impact human tissue culturing protocols.

molecular biology↗

bioRxiv2019

Gut-Liver physiomimetics reveal paradoxical modulation of IBD-related inflammation by short-chain fatty acids

Association between the microbiome, IBD and liver diseases are known, yet cause and effect remain elusive. By connecting human microphysiological systems of the gut, liver and circulating Treg/Th17 cells, we modeled progression of ulcerative colitis (UC) ex vivo. We show that microbiome-derived short-chain fatty acids (SCFA) may either improve or worsen disease severity, depending on the activation state of CD4 T cells. Employing multiomics, we found SCFA increased production of ketone bodies, glycolysis and lipogenesis, while markedly reducing innate immune activation of the UC gut. However, during acute T cell-mediated inflammation, SCFA exacerbated CD4+ T cell effector function, partially through metabolic reprograming, leading to gut barrier disruption and hepatic injury. These paradoxical findings underscore the emerging utility of human physiomimetic technology in combination with systems immunology to study causality and the fundamental entanglement of immunity, metabolism and tissue homeostasis.\n\nO_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=87 SRC=\"FIGDIR/small/706812v2_ufig1.gif\" ALT=\"Figure 1\">\nView larger version (31K):\norg.highwire.dtl.DTLVardef@10006b0org.highwire.dtl.DTLVardef@b7344eorg.highwire.dtl.DTLVardef@1cb0c06org.highwire.dtl.DTLVardef@1826e38_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioengineering↗

bioRxiv2018

Iron and Heme Coordinate Erythropoiesis through HRI-Mediated Regulation of Protein Translation and Gene Expression

Iron and heme play central roles in red blood cell production. However, the mechanisms by which iron and heme levels coordinate erythropoiesis remain incompletely understood. HRI is a heme-regulated kinase that controls translation by phosphorylating eIF2. Here, we investigate the global impact of iron, heme and HRI on protein translation in vivo in murine primary erythroblasts using ribosome profiling. By defining the underlying changes in translation during iron and HRI deficiencies, we validate known regulators of this process, including Atf4, and identify novel pathways such as co-regulation of ribosomal protein mRNA translation. Surprisingly, we found that heme and HRI pathways, but not iron-regulated pathways, mediate the major protein translational and transcriptional responses to iron deficiency in erythroblasts in vivo and thereby identify previously unappreciated regulators of erythropoiesis. Our genome-wide study uncovers the major impact of the HRI-mediated integrated stress response for the adaptation to iron deficiency anemia.

cell biology↗