Biology subjects

Farach-Carson, M. C.

Publications and source records attributed to Farach-Carson, M. C..

15 recordsLinked to original sources

bioRxiv2026

Nonlinear rheology of laminin-111-modified hyaluronan hydrogels

We experimentally assess the nonlinear rheology of composite biopolymer hydrogels composed of thiolated hyaluronic acid, poly(ethylene glycol) diacrylate (PEGDA), and laminin-111 in varied concentrations. We focus in particular on the influence of laminin on the mechanics of the assembled hydrogels, reporting nonlinear rheological measurements for gels under applied shear and compressive load. We find that increasing the concentration of laminin in the synthesized gels reduces the linear shear modulus and gives rise to a mild strain softening regime at intermediate strains prior to the onset of strain stiffening. In the stiffening regime, we find that all gels exhibit stress-controlled mechanics with K {propto}{sigma} a, with an apparent stiffening exponent of a {approx} 1, in agreement with observations of a variety of other reconstituted biopolymer gels. We discuss the possible implications of this nonlinear mechanical behavior on mechanotransduction and organoid development in biomimetic extracellular matrices.

biophysics↗

bioRxiv2026

Assessing Bioactivity and Biointegration of Engineered Salivary Tissue Constructs in a Preclinical Unilateral Fractionated Irradiated Rat Model

Human salivary stem/progenitor cell (hS/PC)-loaded hyaluronic acid (HA)-based hydrogels, termed 3D-salivary tissue constructs (3D-ST), hold great promise for restoring salivary gland function post-radiation injury. Here, we developed a next-generation 3D-ST using heparin-modified HA and bioactive peptide-modified hydrogels. This new formulation enables controlled pre-loading and localized presentation of heparin-binding growth factors prior to surgical implantation, providing opportunities to enhance in vivo hS/PC bioactivity. To model clinically relevant radiation injury, we established an athymic rat model subjected to computed tomography (CT)-guided fractionated radiation, resulting in hallmark features of radiation-induced salivary dysfunction. Over 60-days post-irradiation, glands exhibited progressive loss of acini, increased fibrosis, and disruption of endothelial, neuronal, and myoepithelial compartments. Within this injured environment, a surgical pocket was created to precisely implant 3D-STs to assess graft performance. Fluorescent labeling of the 3D-STs enabled longitudinal tracking post-implantation. Over 14 days, implanted 3D-STs remained structurally stable within irradiated glands, and hS/PCs remained viable without evidence of local inflammatory responses. Compared to non-injured glands, the irradiated microenvironment suppressed hS/PC proliferation and phenotype, indicating alterations in the irradiated local tissue negatively impact hS/PC bioactivity. In addition, host neurovascular migration into the 3D-ST was majorly restricted in irradiated glands, providing new opportunities to enhance biointegration. Overall, this work establishes a reproducible preclinical framework for assessing hydrogel biocompatibility and stability, cell bioactivity, and host-graft biointegration prior to scale up into preclinical large animal models. This study has successfully established a tractable approach for improving 3D-ST formulations to enhance hS/PC expansion, differentiation, and biointegration following implantation into radiation-injured beds.

bioengineering↗

bioRxiv2026

Optimizing Primary Human Salivary Stem/Progenitor Cells for Tissue Engineering Applications

This work aimed to establish a translationally viable, xeno-free, serum-free platform and protocol for the isolation and expansion of human salivary stem/progenitor cells (hS/PCs) suitable for regulatory qualification and future FDA-approved first-in-human autologous regenerative therapy trials for the treatment of hyposalivation disorders. Parotid gland specimens from non-cancerous regions/tissues were collected from consented surgical patients. Primary hS/PCs were isolated from tissue specimens, cultured in animal-component-free conditions, expanded to produce millions of cells, then enriched for CD44+ stem/progenitor cells by magnetic cell sorting. Normal epithelial purity was assessed using cytokeratins 5/14. Anti-CD133/PROM1 (cancer marker) and anti- fibroblast (clone TE-7) antibodies were used to demonstrate a lack of contaminating cells. Phenotype validation was performed by flow cytometry and immunocytochemistry on both CD44+ sorted and unsorted populations. Senescence-associated beta-galactosidase (SA-{beta}-gal) assays were performed across serial passages (P1-P6). Pluripotency was demonstrated by culture under conditions supporting lineage-specific differentiation. Primary hS/PCs demonstrated consistent expansion and epithelial morphology under serum-free conditions. CD44 expression remained high (>95%) throughout expansion, with negligible detection of CD133 or fibroblast markers, confirming epithelial purity and absence of tumorigenic or stromal contamination. Immunocytochemistry corroborated these expression profiles. SA-{beta}-gal staining revealed only a minor, passage-dependent increase (5-16%) in senescent cells from multiple donors, indicating retention of proliferative potential. Our defined, animal-free culture system supports stable expansion of pure low passage hS/PCs under conditions compatible with good manufacturing practice (GMP).

cell biology↗

bioRxiv2025

Antibody-Protein L Functionalized Microparticles for Detection of Surface Markers in Heterogeneous Colorectal Lesions

Visualization of colorectal cancer (CRC) lesions is complicated by their location in the colon and tumor morphology. Reliance on a single surface biomarker for direct identification risks false negatives due to temporal changes and/or tumor heterogeneity. We developed a multiplexed system of complementary biomarker targets in an effort to capture a broader range of lesions with diverse temporal and/or phenotypic expression. We identified Mucin-1 (MUC1) and epithelial cell adhesion molecule (EPCAM) as useful targeting pairs by examining multiple colon tumor subtypes in a standard tissue array, and by surveying multiple CRC cell lines, both as 2D cultures and as 3D tumoroids, for the presence of the CRC surface biomarkers. We demonstrated the utility of a "universal" surface functionalization approach using Antibody-Protein L functionalized microparticles (APL-MPs) that enabled the simultaneous incorporation of antibodies recognizing MUC1 and EPCAM. Using CRC cell heterogeneous tumoroids expressing both MUC1 and EPCAM (HET-tumoroids) and orthotopic animal cancer models designed to express both surface antigens, we demonstrated that: 1) APL-MPs identified MUC1- and EPCAM-positive tumoroids in proportion to antigen expression; 2) APL-MPs detected CRC surface antigens on the luminal colon surface in vivo, and 3) concurrent targeting of multiple surface antigens enhanced the sensitivity of detection of heterogeneous CRC lesions. This approach opens the door for the use of antibody-protein L dual-targeting MPs in a variety of applications to detect heterogeneous cancer lesions. SignificanceMulti-antigen targeting of MUC1 and EPCAM with Antibody-Protein L microparticles enhances the detection sensitivity of heterogeneous colorectal cancer lesions, offering a promising strategy for the accurate visualization of tumors in complex environments.

cancer biology↗

bioRxiv2025

Exploring Spike-Dependent and ACE2-Independent Viral Entry into Salivary Epithelial Cells in the Absence of ACE2

Salivary gland infection by SARS-CoV-2 requires viral entry via routes and mechanisms that remain unresolved. This study examined the expression of the angiotensin- converting enzyme 2 (ACE2) receptor in salivary tissues and basal cell-derived human salivary progenitor cells (hS/PCs), an unstudied potential entry point for SARS-CoV-2. Multiple detection modalities, including immunocytochemistry, western blotting, flow cytometry and RT- PCR, demonstrated a consistent lack of ACE2 protein and transcript in both tissue specimens and primary salivary epithelial cells. Antigen retrieval at pH 9 was determined to be optimal for immunodetection protocols, yet ACE2 remained undetectable. Small intestine tissue served as a positive control, confirming the validity of the methods and reagents we used. Considering there can be other receptors for SARS-CoV-2, flow cytometric analyses demonstrated that recombinant SARS-CoV-2 spike protein failed to bind to salivary epithelial cells, in contrast to HEK293 cells engineered to overexpress ACE2, which showed robust spike binding. These findings strongly support our conclusion that salivary cells do not serve as major targets for SARS-CoV-2 infection via ACE2 or spike protein, whether through direct exposure to viral particles in ductal fluids or via access to basal cells across the basement membrane.

cell biology↗

bioRxiv2025

Targeted Molecular MRI of Colorectal Cancer by Antibody Functionalized Hyperpolarized Silicon Particles

The development of non-invasive, non-ionizing sensitive molecular targeting approaches to detect colorectal cancer (CRC) lesions is warranted to improve high risk patient outcomes. Hyperpolarized silicon nanoparticles and microparticles are potentially well-suited to act as targeted molecular imaging agents because of their overall biocompatibility and long-lasting enhanced magnetic resonance imaging (MRI) signals. In this study, dynamic nuclear polarization was performed on silicon particles functionalized with an antibody to Mucin-1 (MUC1), a surface mucin glycoprotein aberrantly expressed in CRC. Antibody conjugation to the particle surface did not affect 29Si hyperpolarization characteristics. Similarly, conjugation and the dynamic nuclear polarization process did not adversely affect the affinity of the targeting antibody. In vivo MRI scans performed 10-15 minutes after luminal administration of targeted hyperpolarized particles into human MUC1-expressing orthotopic CRC mouse models showed that particles actively targeted tumor sites. These results were supported by chemical and biological controls and blocking experiments as well as correlative immunohistochemical analysis. These surface-functionalized silicon particles are under development as a platform technology that will allow non-invasive molecular targeting of CRC using hyperpolarized MRI. Single Sentence SummaryTargeted molecular MR imaging of colorectal cancer by hyperpolarized silicon particles functionalized with mucin 1 antibody.

cancer biology↗

bioRxiv2024

Computational Modeling and Analysis of the TGF-β-induced ERK and SMAD Pathways

TGF-{beta}, an important cytokine that plays a key role in many diseases regulates a wide array of cellular and physiologic processes via several TGF-{beta}-driven signaling cascades, including the SMAD and non-SMAD-driven pathways. However, the detailed mechanisms by which TGF-{beta} induces such diverse responses remain poorly understood. In particular, compared to the SMAD-dependent pathway, SMAD-independent pathways such as the ERK/MAPK pathway, which is critical in cancer progression, are less characterized. Here, we develop an integrated mechanistic model of the TGF-{beta}-triggered ERK activation pathway and its crosstalk with the SMAD pathway, an analysis of which demonstrates how SMAD dynamics can be significantly modulated and regulated by the ERK pathway. In particular, SMAD-mediated transcription can be altered and delayed due to expedited phosphorylation of the linker of SMAD by TGF-{beta}-activated ERK; and enhanced ERK activity, but attenuated SMAD activity, can be achieved simultaneously by fast turnover of TGF-{beta} receptors via lipid-rafts. Also, in silico mutations of the TGF-{beta} pathways reveal that the dynamic characteristics of both SMAD and ERK signaling may change significantly during cancer development. Specifically, normal cells may exhibit enhanced and sustained SMAD signaling with transient ERK activation, whereas cancerous cells may produce elevated and prolonged ERK signaling with enervated SMAD activation. These distinctive differences between normal and cancerous signaling behavior provide clues concerning, and potential explanations for, the seemingly contradictory roles played by TGF-{beta} during cancer progression. We demonstrate how crosstalk among various branch pathways of TGF-{beta} can influence overall cellular behavior. Based on model analysis, we hypothesize that aberrant molecular alterations drive changes in the intensity and duration of SMAD and ERK signaling during cancer progression and ultimately lead to an imbalance between the SMAD and ERK pathways in favor of tumor promotion. Thus, to treat cancer patients with a genetic signature of oncogenic Ras effectively may require at least a combination therapy to restore both the expression of TGF-{beta} receptors and the GTPase activity of Ras.

systems biology↗

bioRxiv2023

Development and Characterization of a low intensity vibrational system for microgravity studies

The advent of extended-duration human spaceflight demands a better comprehension of the physiological impacts of microgravity. One primary concern is the adverse impact on the musculoskeletal system, including muscle atrophy and bone density reduction. Ground-based microgravity simulations have provided insights, with vibrational bioreactors emerging as potential mitigators of these negative effects. Despite the potential they have, the adaptation of vibrational bioreactors for space remains unfulfilled, resulting in a significant gap in microgravity research. This paper introduces the first automated low-intensity vibrational (LIV) bioreactor designed specifically for the International Space Station (ISS) environment. Our research covers the bioreactors design and characterization, the selection of an optimal linear guide for consistent 1-axis acceleration, a thorough analysis of its thermal and diffusion dynamics, and the pioneering use of BioMed Clear resin for enhanced scaffold design. This advancement sets the stage for more authentic space-based biological studies, vital for ensuring the safety of future space explorations.

bioengineering↗

bioRxiv2023

Increased deformations of aging bone marrow during physical activity is dispensable for cell mechanoresponse in bone analogs

Aged individuals and astronauts experience bone loss despite rigorous physical activity. Bone mechanoresponse is in-part regulated by mesenchymal stem cells (MSCs) that respond to mechanical stimuli. Direct delivery of low intensity vibration (LIV) recovers MSC proliferation in senescence and simulated microgravity models, indicating that age-related reductions in mechanical signal delivery within bone marrow may contribute to declining bone mechanoresponse. To answer this question, we developed a 3D bone marrow analog that controls trabecular geometry, marrow mechanics and external stimuli. Validated finite element (FE) models were developed to quantify strain environment within hydrogels during LIV. Bone marrow analogs with gyroid-based trabeculae of bone volume fractions (BV/TV) corresponding to adult (25%) and aged (13%) mice were printed using polylactic acid (PLA). MSCs encapsulated in migration-permissive hydrogels within printed trabeculae showed robust cell populations on both PLA surface and hydrogel within a week. Following 14 days of LIV treatment (1g, 100 Hz, 1 hour/day), type-I collagen and F-actin were quantified for the cells in the hydrogel fraction. While LIV increased all measured outcomes, FE models predicted higher von Mises strains for the 13% BV/TV groups (0.2%) when compared to the 25% BV/TV group (0.1%). Despite increased strains, collagen-I and F-actin measures remained lower in the 13% BV/TV groups when compared to 25% BV/TV counterparts, indicating that cell response to LIV does not depend on hydrogel strains and that bone volume fraction (i.e. available bone surface) directly affects cell behavior in the hydrogel phase independent of the external stimuli. Overall, bone marrow analogs offer a robust and repeatable platform to study bone mechanobiology.

bioengineering↗

bioRxiv2022

Adipose triglyceride lipase is regulated by CAMKK2-AMPK signaling and drives advanced prostate cancer

Lipid metabolism plays a central role in prostate cancer. To date, the major focus on prostate cancer lipid metabolism has centered on de novo lipogenesis and lipid uptake with little consideration for how cancer cells access these lipids once they are created or taken up and stored. Patient-derived phosphoproteomics identified adipose triglyceride lipase (ATGL), a previously suspected tumor suppressor, as a CAMKK2-AMPK signaling target that, conversely, promotes castration-resistant prostate cancer (CRPC) progression. Phosphorylation of ATGL increased its lipase activity, cancer cell proliferation, migration, and invasion. Shotgun lipidomics and mass spectrometry imaging demonstrated ATGLs profound regulation of lipid metabolism in vitro and in vivo, remodeling membrane composition. Inhibition of ATGL induced metabolic plasticity, causing a glycolytic shift that could be exploited therapeutically by co-targeting both metabolic pathways. Together, these data nominate ATGL and intracellular lipolysis as potential therapeutic targets for the treatment of CRPC and provide insights for future combination therapies.

cancer biology↗

bioRxiv2022

Gabapentin Disrupts Binding of Perlecan to the α2δ1 Voltage Sensitive Calcium Channel Subunit and Impairs Skeletal Mechanosensation

Our understanding of how osteocytes, the principal mechanosensors within bone, sense and perceive force remains unclear. Previous work identified "tethering elements" (TEs) spanning the pericellular space of osteocytes and transmitting mechanical information into biochemical signals. While we identified the heparan sulfate proteoglycan perlecan (PLN) as a component of these TEs, PLN must attach to the cell surface to induce biochemical responses. As voltage-sensitive calcium channels (VSCCs) are critical for bone mechanotransduction, we hypothesized that PLN binds the extracellular 2{delta}1 subunit of VSCCs to couple the bone matrix to the osteocyte membrane. Here, we showed co-localization of PLN and 2{delta}1 along osteocyte dendritic processes. Additionally, we quantified the molecular interactions between 2{delta}1 and PLN domains and demonstrated for the first time that 2{delta}1 strongly associates with PLN via its domain III. Furthermore, 2{delta}1 is the binding site for the commonly used pain drug, gabapentin (GBP), which is associated with adverse skeletal effects when used chronically. We found that GBP disrupts PLN::2{delta}1 binding in vitro, and GBP treatment in vivo results in impaired bone mechanosensation. Our work identified a novel mechanosensory complex within osteocytes composed of PLN and 2{delta}1, necessary for bone force transmission and sensitive to the drug GBP. This work provides insights into the mechanisms underlying mechanotransduction and will inform future studies to understand the mechanisms responsible for the negative effects of GBP on bone.

molecular biology↗

bioRxiv2021

TWIST1 interacts with adherens junction proteins during neural tube formation and regulates fate transition in cranial neural crest cells

Cell fate determination is a necessary and tightly regulated process for producing different cell types and structures during development. Cranial neural crest cells (CNCCs) are unique to vertebrate embryos and emerge from the neural fold borders into multiple cell lineages that differentiate into bone, cartilage, neurons, and glial cells. We previously reported that Irf6 genetically interacts with Twist1 during CNCC-derived tissue formation. Here, we investigated the mechanistic role of Twist1 and Irf6 at early stages of craniofacial development. Our data indicates that TWIST1 interacts with /{beta}/{gamma}-CATENINS during neural tube closure, and Irf6 is involved in the structural integrity of the neural tube. Twist1 suppresses Irf6 and other epithelial genes in CNCCs during epithelial-to-mesenchymal transition (EMT) process and cell migration. Conversely, a loss of Twist1 leads to a sustained expression of epithelial and cell adhesion markers in migratory CNCCs. Disruption of TWIST1 phosphorylation in vivo leads to epidermal blebbing, edema, neural tube defects, and CNCC-derived structural abnormalities. Altogether, this study describes an uncharacterized function of Twist1 and Irf6 in the neural tube and CNCCs and provides new target genes of Twist1 involved in cytoskeletal remodeling. Furthermore, the association between DNA variations within TWIST1 putative enhancers and human facial morphology is also investigated. SUMMARY STATEMENTThis study uncovers a new function of Twist1 in neural tube development and epithelial-to-mesenchymal transition in cranial neural crest cells. Data further shows that Twist1-interacting Irf6 is involved in regulating neural tube integrity.

developmental biology↗

bioRxiv2021

Functionalized Biomimetic Hydrogels Enhance Salivary Stem/Progenitor Cell Organization

Complex branched salivary structures remain challenging to replicate within implant ready hydrogels. We showed previously that hyaluronic acid (HA)-based hydrogels enable growth and organization of primary salivary-derived human stem/progenitor cells (hS/PCs) into multicellular spheroids. Here, we systematically functionalized three components of migration-permissive hydrogels to foster salivary tissue morphogenesis. We separately analyzed contributions of an enzymatically degradable crosslinker, a pendant integrin-binding site, and hydrogel porosity to best support high viability, integrin-dependent cell adhesion and migration. Structure size, frequency, and morphology were all affected by hydrogel crosslink density and integration of biofunctional peptides. Viability and proliferation data suggested that integration of integrin binding sites had the greatest effect on hS/PCs behavior. A larger internal matrix space, created by increasing both crosslinker length and PEG content, was needed to form large multicellular hS/PC structures. Peptide-modified hydrogels with more internal space shifted hS/PC organization from spheroidal, surrounded by thick basement membrane, to an asymmetric arrangement with punctate matrix proteins defining a "wrinkled" perimeter. Integrin-binding peptides activated integrin {beta}1, with highest activation observed in hydrogels having both cleavable peptide and integrin ligand. The design parameters we prescribe allowed us to encapsulate hS/PCs in a humanized biomimetic hydrogel matrix able to support morphogenesis needed for salivary restoration.

bioengineering↗

bioRxiv2020

SULF1 suppresses Wnt3A-driven growth of bone metastatic prostate cancer in perlecan-modified 3D cancer-stroma-macrophage triculture models

Bone marrow stroma influences metastatic prostate cancer (PCa) progression, latency, and recurrence. At sites of PCa bone metastasis, cancer-associated fibroblasts and tumor-associated macrophages interact to establish a perlecan-rich desmoplastic stroma. As a heparan sulfate proteoglycan, perlecan (HSPG2) stores and stabilizes growth factors, including heparin-binding Wnt3A, a positive regulator of PCa cell growth. Because PCa cells alone do not induce CAF production of perlecan in the desmoplastic stroma, we sought to discover the sources of perlecan and its growth factor-releasing modifiers SULF1, SULF2, and heparanase in PCa cells and xenografts, bone marrow fibroblasts, and macrophages. SULF1, produced primarily by bone marrow fibroblasts, was the main glycosaminoglycanase present, a finding validated with primary tissue specimens of PCa metastases with desmoplastic bone stroma. Expression of both HSPG2 and SULF1 was concentrated in SMA-rich stroma near PCa tumor nests, where infiltrating pro-tumor TAMs also were present. To decipher SULF1s role in the reactive bone stroma, we created a bone marrow biomimetic hydrogel incorporating perlecan, PCa cells, macrophages, and fibroblastic bone marrow stromal cells. Finding that M2-like macrophages increased levels of SULF1 and HSPG2 produced by fibroblasts, we examined SULF1 function in Wnt3A-mediated PCa tumoroid growth in tricultures. Comparing control or SULF1 knockout fibroblastic cells, we showed that SULF1 reduces Wnt3A-driven growth, cellularity, and cluster number of PCa cells in our 3D model. We conclude that SULF1 can suppress Wnt3A-driven growth signals in the desmoplastic stroma of PCa bone metastases, and SULF1 loss favors PCa progression, even in the presence of pro-tumorigenic TAMs.

cancer biology↗

bioRxiv2017

Inflammatory Breast Cancer: a model for investigating cluster-based dissemination

Metastases claim more than 90% of cancer-related patient deaths and are usually seeded by a subset of circulating tumor cells (CTCs) shed off from the primary tumor. In circulation, CTCs are found both as single cells and as clusters of cells. The clusters of CTCs, although many fewer in number, possess much higher metastatic potential as compared to that of individual CTCs. In this review, we highlight recent insights into molecular mechanisms that can enable the formation of these clusters - (a) hybrid epithelial/mesenchymal (E/M) phenotype of cells that couples their ability to migrate and adhere, and (b) intercellular communication that can spatially coordinate the cluster formation and provide survival signals to cancer cells. Building upon these molecular mechanisms, we also offer a possible mechanistic understanding of why clusters are endowed with a higher metastatic potential. Finally, we discuss the highly aggressive Inflammatory Breast Cancer (IBC) as an example of a carcinoma that can metastasize via clusters and corroborates the proposed molecular mechanisms.

cancer biology↗