Genaro Vázquez-Victorio

Vascular remodeling responds to local wall shear stress, but transport efficiency belongs to the network as a whole. We connect these scales by minimizing dissipation under a fixed maintenance budget, with vessel cost proportional to mass raised to an exponent b . At an optimum, dissipated power is proportional to vessel cost. This yields the shear set-point | τ | ∝ r ^{b −1} l ^{( b −1)/2} for radius r and length l , with uniform shear recovered at b = 1. The associated local adaptation rule decreases the global functional, although the topology it reaches retains a memory of development. Simulations with 400 and 700 sinks show that staged growth improves transport mainly by allowing pruned connections to regrow. Fluctuating demand sustains loops without increasing the maintenance budget. The local condition fixes allocation on a given support, whereas growth and variable demand shape the topology that survives.

During liver fibrosis, hepatocytes respond to progressive increases in tissue stiffness by undergoing cytoskeletal reorganisation. This has important consequences such as immune cell infiltration, mainly triggered by the activation of YAP and TAZ proteins. However, other proteins that respond to mechanical modifications remain poorly characterised; in particular, the myocardin‐related transcription factors A and B (MRTFs). Here, we referred to these four proteins as mechanical responsive transcriptional cofactors (MRTcoF). We analysed the regulation of MRTcoF proteins in fibrotic livers induced by carbon tetrachloride (CCl ₄ ) and observed that MRTcoF were differentially regulated during liver fibrosis, predominantly in hepatocytes. Transcriptome analysis suggested that fatty acid and glucose metabolisms, cell proliferation and signalling pathways were regulated by transcriptional enhanced associate domain (TEAD)‐ and serum response factor (SRF)‐transcriptional programmes in CCl ₄ ‐treated livers. Then, we evaluated the protein expression levels of MRTcoF in primary hepatocytes (PH) cultured on tissue culture petri dishes (TCPD). After culture, we observed upregulation of MRTcoF, and downregulation of TAZ after protein increase. Interestingly, despite the increase in MRTFB protein levels, no accumulation of MRTFB was detected in the nucleus. Culture on soft polyacrylamide hydrogels (PAA HGs) attenuated the activation of MRTcoF, having specific implications in the transcriptional regulation of target genes. In contrast, pharmacological inhibition of MRTcoF was not sufficient to halt transcriptional regulation. In summary, a progressive increase of stiffness activates MRTcoF transcriptional complexes in hepatocytes, suggesting a key role of mechano‐transduction during liver fibrosis.

The integration of microfluidic technologies in organoid generation has enhanced the development of precise and functional 3D cell culture models, overcoming the limitations of traditional cultures, especially for brain organoids. The application of controlled morphogen gradients in microfluidic platforms has enabled faithful tissue patterning, cellular differentiation, and spatial organization of brain organoids. Here, we report the design, fabrication, and validation of a custom‐made double‐gradient microfluidic device for organoid culture. The device consists of a central channel for organoid growth, flanked by two parallel lateral channels that serve as reservoirs. These components were assembled using a porous membrane as an interface, bonded using APTES surface treatment and corona discharge activation, followed by thermal curing to ensure stability. Human embryonic stem cells were seeded and labeled with fluorescent dyes, where asymmetric fluorescence intensity confirmed efficient double‐gradient formation. Subsequently, midbrain organoids were loaded into the device under the same conditions, demonstrating gradient penetration into tissue‐level structures. A polarized differentiation protocol was successfully established within the platform, leading to the generation of differentiated dorsal and ventral neurons with asymmetric spatial distribution. These results establish a robust and reproducible pump‐free protocol for a double‐gradient microfluidic device, providing a versatile tool to establish polarized organoids for studies in patterning or the incorporation of more functional organoids in damaged tissues.

Conventional cancer therapies often harm healthy tissues due to their poor specificity, resulting in significant side effects that diminish patients’ quality of life. Parasporins, a group of non-hemolytic parasporal proteins produced by Bacillus thuringiensis, are known for their selective cytotoxicity toward cancer cells. Typically, these proteins require activation through physical or biochemical treatments that fragment them into multiple peptides of varying sizes, which are then tested as mixtures, without purification, against cancer cell lines. In this study, a purification strategy that isolates the protein without prior activation and evaluates the resulting cytotoxic mechanism is proposed. The purification consists of four steps: (1) crystal solubilization with Laemmli buffer, (2) size-based separation via SDS-PAGE, (3) electroelution of the target protein from the gel, and (4) dialysis to remove the elution buffer. From the B. thuringiensis AX isolate, four proteins ranging from ~20 to 60 kDa were recovered, but only AX-2 displayed cytotoxic activity toward MCF-7 breast cancer cells, while remaining non-hemolytic and non-toxic to normal cells (erythrocytes, PBMCs, and MRC-5 fibroblasts). Thus, AX-2 qualifies as a parasporin. AX-2 induces apoptosis in MCF-7 breast cancer cells without generating oxidative stress, and the observed cell death appears to initiate at the plasma membrane rather than through intracellular pathways.

TGF-β is a pleiotropic cytokine with both stimulatory and inhibitory effects on immune cells, depending on the microenvironmental context. It targets mast cells (MCs) in different physio-pathological conditions, such as inflammation and cancer. Besides acting as a potent chemoattractant for MCs, TGF-β regulates many other aspects of MCs’ physiology, including the secretion of many regulatory molecules. MCs secrete a variety of mediators, either pre-formed or newly synthesized, upon appropriate stimulation. CCL-2 chemokine and TNF cytokine act as potent chemoattractants for several immune cells and participate in the initiation of inflammatory responses by recruiting them to injured tissues. TGF-β regulates CCL-2 and TNF secretion in different cell types and under distinct cellular contexts. Here, we report that the treatment with TGF-β alone induces the secretion of both pre-formed and newly synthesized CCL-2 in the rat RBL-2H3 mast cells but not in mouse bone marrow-derived mast cells (BMMCs). TGF-β-induced CCL-2 secretion depends on rapid rearrangements of the actin cytoskeleton and, remarkably, on the early secretion of soluble TNF that triggers an autocrine TNF signaling. In conclusion, we found cooperation between TGF-β and TNF signaling pathways to promote the secretion of CCL-2 chemokine by MCs in a cell-context specific manner.

Surface modification of polydimethylsiloxane (PDMS) for organ‐on‐a‐chip (OOC) systems is fundamental for the success of cell physiological assays. Although UV light is commonly used for this purpose, surface chemical modifications are only temporary. To overcome these limitations, an alternative approach is proposed: a physicochemical modification using ozone and the heterofunctional crosslinker sulfo‐SANPAH (SS). This simple one‐step approach is carried out on PDMS microchannels of OOC platforms. A broad physicochemical characterization based on spectroscopy and microscopy techniques confirms successful modification of the PDMS surface. The PDMS surface contact angle decreases after addition of SS (from 101.4° to 79.9°) but decreases as low as 20.8° after extracellular matrix deposition, demonstrating successful transition of the surface from hydrophobic to hydrophilic. For biological characterization, human liver sinusoidal endothelial cells (LSEC) and murine primary hepatocytes are cultured in OOC channels for up to 7 days. LSECs and hepatocytes establish strong cell adhesions under laminar flow and form a well‐established canaliculi network, respectively. This one‐step method of PDMS surface modification for OOC is simpler than others reported in the literature (e.g., based on 3‐aminopropyltriethoxysilane) and ideal for beginners interested in using microfluidic platforms for cell culture.

Purinergic signaling has emerged as an important paracrine–autocrine intercellular system that regulates physiological and pathological processes in practically all organs of the body. Although this system has been thoroughly defined since the nineties, recent research has made substantial advances regarding its role in aspects of liver physiology. However, most studies have mainly targeted the entire organ, 70% of which is made up of parenchymal cells or hepatocytes. Because of its physiological role, the liver is exposed to toxic metabolites, such as xenobiotics, drugs, and fatty acids, as well as to pathogens such as viruses and bacteria. Under injury conditions, all cell types within the liver undergo adaptive changes. In this context, the concentration of extracellular ATP has the potential to increase dramatically. Indeed, this purinergic response has not been studied in sufficient detail in non-parenchymal liver cells. In the present review, we systematize the physiopathological adaptations related to the purinergic system in chronic liver diseases of non-parenchymal liver cells, such as hepatic stellate cells, Kupffer cells, sinusoidal endothelial cells, and cholangiocytes. The role played by non-parenchymal liver cells in these circumstances will undoubtedly be strategic in understanding the regenerative activities that support the viability of this organ under stressful conditions.

This work analyzes the role of the tight junction (TJ) protein ZO-2 on mechanosensation. We found that the lack of ZO-2 reduced apical membrane rigidity measured with atomic force microscopy, inhibited the association of γ-actin and JAM-A to the cell border, and instead facilitated p114RhoGEF and afadin accumulation at the junction, leading to an enhanced mechanical tension at the TJ measured by FRET, with a ZO-1 tension probe, and increased tricellular TJ tension. Simultaneously, adherens junction tension measured with an E-cadherin probe was unaltered. The stability of JAM-A and ZO-2 binding was assessed by a collaborative in silico study. The absence of ZO-2 also impacted the cell response to the substrate, as monolayers plated in 20 kPa hydrogels developed holes not seen in parental cultures and displayed a retarded elongation and formation of cell aggregates. The absence of ZO-2 was sufficient to induce YAP and Snail nuclear accumulation in cells cultured over glass, but when ZO-2 KD cells were plated in nanostructured ridge arrays, they displayed an increased abundance of nuclear Snail and conspicuous internalization of claudin-4. These results indicate that the absence of ZO-2 also impairs the response of cells to substrate stiffness and exacerbates transformation triggered by substrate topography.

In this work, we present an accessible benchtop fabrication technique to obtain a planar array of gold nanowrinkled surface electrodes (ANSE) for the construction of electrochemical cells, specifically to monitor soluble biomarkers of interest in cell culture environments. We present a complete characterization of the array and its response as an electrochemical cell. To validate our sensor, we evaluated the device sensitivity to detect nitric oxide (NO), an important molecule produced by endothelial cells as a response to environmental signals such as mechanics and growth factors. While testing measurements of nitric oxide in aqueous solutions with isotonic salt concentrations, we evidenced the influence of the environmental conditions for such electrochemical measurements, showing that the aqueous medium, usually not accounted for, significantly impacts the outcome. Finally, we present the application of the electrochemical sensor for the detection of nitric oxide released from stimulated endothelial cells as a proof of concept.