Biology subjects

Reesink, H. L.

Publications and source records attributed to Reesink, H. L..

7 recordsLinked to original sources

bioRxiv2026

Functional Differentiation of Type II and Type I Collagen Articular Models in Synovial Fluid Film Formation and Recombinant Lubricin Retention

Collagen type II (Col-II) and collagen type I (Col-I) are major components of articular cartilage present at different ratios at its surface. Understanding how each of these components mediates the assembly of molecular films derived from synovial fluid (SF), the native lubricant of synovial joints, is critical to explain the loss of mechanical performance in pathological conditions, guide the design of biomaterial implants meant to be in contact with SF, and develop molecular therapies to restore SF properties. This work demonstrates that Col-II articular surface model assists in scaffolding of full SF-derived films, while Col-I model lacks SF film scaffolding capabilities. However, when Col-II and Col-I are exposed to recombinant lubricin (rLub) alone, the major boundary lubricant in SF, both adsorbed and retained similar amounts. These insights, deduced from quartz crystal microbalance with dissipation, diffuse reflectance circular dichroism, and atomic force microscopy, reveal possible mechanisms underlying the loss of mechanical performance of synovial joints in pathology, where Col-I becomes the major collagenous component of the articular cartilage surface, as well as considerations for designing functional biomaterial implants. Furthermore, this work reinforces the idea of rLub as an intra-articular osteoarthritis therapy with the ability to bind to Col-II and Col-I, irrespectively. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=170 SRC="FIGDIR/small/726594v1_ufig1.gif" ALT="Figure 1"> View larger version (69K): org.highwire.dtl.DTLVardef@138a77borg.highwire.dtl.DTLVardef@7b8512org.highwire.dtl.DTLVardef@15d7060org.highwire.dtl.DTLVardef@17ccf20_HPS_FORMAT_FIGEXP M_FIG C_FIG

biophysics↗

bioRxiv2026

A role for heavy chain-modification in protecting hyaluronan from free radical fragmentation during inflammation

The glycosaminoglycan hyaluronan (HA) is an essential and ubiquitous component of human tissues and biofluids. The only known covalent modification of HA entails the attachment of heavy chains (HC) from the inter-alpha-inhibitor (II) family of proteoglycans, forming stable complexes (HC*HA) that arise during inflammation. In some contexts, HC*HA is thought to contribute to pathology, whereas in others it may form part of a protective pathway. However, its exact roles are not fully understood. Here, we report that HC modifications can protect HA from fragmentation by reactive oxygen species (ROS) produced during the inflammatory cascade. Using solid-state nanopore molecular size analysis, we show that HA is highly resistant to degradation from exogenous ROS in vitro when in the context of HC*HA complexes, while the unmodified HA polymer is fragmented rapidly under the same conditions. Experiments performed with admixtures of HA and unbound antioxidant proteins - including HC-bearing components - demonstrate the necessity of covalent HC attachment to the polysaccharide for the protection. Finally, we find that HA with high-HC content from inflammatory equine synovial fluid has increased resilience to ROS damage compared to low-HC HA from a healthy joint. Collectively, these results demonstrate that covalent HC modification is an effective biological strategy for preserving HA integrity against ROS fragmentation, including in inflammatory conditions.

molecular biology↗

bioRxiv2025

Lectin Microarray-based Glycomics and Machine Learning Identify Shared Osteoarthritis Biomarkers in Humans, Dogs, and Horses

Post-traumatic osteoarthritis (PTOA) is a common sequela to joint injury in both humans and companion animal species such as horses and dogs. Despite the increasing prevalence of osteoarthritis (OA) in humans, investigation of glycosylation changes associated with OA remains in its infancy. Recent advances, such as lectin microarray analysis, now enable detailed glycan profiling in complex biofluids such as synovial fluid. Using lectin microarray technology, this study characterized glycosylation patterns in synovial fluid samples from healthy and OA-affected joints in horses, dogs, and humans. Comparative glycan-binding profiles within and between species revealed conserved and distinct glycomic signatures associated with OA. Machine learning models, including classification algorithms, effectively distinguished OA from healthy joints, identifying key lectins and glycan epitopes crucial to these predictions. The identified lectin markers reflect specific glycosylation pathways and potential inflammatory mechanisms, demonstrating their value in differentiating between healthy and OA phenotypes. Our findings underscore the promise of integrated glycomic profiling and machine learning to enhance our understanding of glycan involvement in the pathogenesis of OA and to facilitate the development of diagnostic and therapeutic strategies applicable to both veterinary and human medicine. In BriefOsteoarthritis affects humans and companion animals; however, its molecular features remain unclear. Using lectin microarrays and machine learning, we identified conserved and species-specific glycan signatures in synovial fluid that differentiate between control and osteoarthritic joints. This One Health approach highlights shared molecular mechanisms of joint degeneration and establishes data-driven glycomic profiling as a framework for understanding osteoarthritis across species. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=162 SRC="FIGDIR/small/682971v1_ufig1.gif" ALT="Figure 1"> View larger version (34K): org.highwire.dtl.DTLVardef@1644beaorg.highwire.dtl.DTLVardef@1b4133dorg.highwire.dtl.DTLVardef@15b286aorg.highwire.dtl.DTLVardef@1ba5d62_HPS_FORMAT_FIGEXP M_FIG C_FIG

molecular biology↗

bioRxiv2025

Delayed lubricin injection improves cartilage repair tissue quality in an in vivo rabbit osteochondral defect model

Osteochondral lesions (OCL) are common among young patients and often require surgical interventions since cartilage has a poor capacity for self-repair. Bone marrow stimulation (BMS) has been used clinically for decades to treat OCLs, however a persisting challenge with BMS and other cartilage repair strategies is the inferior quality of the resulting fibrocartilaginous repair tissue. Lubrication-based therapies have the potential to improve the quality of cartilage repair tissue as joint lubrication is linked to local cartilage tissue strains and subsequent cellular responses including death and apoptosis. Recently, a full length recombinant human lubricin (rhLubricin) was developed and has been shown to lower friction in cartilage. This study investigated the effect of a single delayed injection of rhLubricin on cartilage repair in an in vivo rabbit OCL model using gross macroscopic evaluation, surface profilometry, histology, and tribology. Moderate improvement in macroscopic scores for cartilage repair were observed. Notably, quantitative analysis of Safranin-O histology showed that rhLubricin treated joints had significantly higher glycosaminoglycan content compared to saline treated joints, and there were no differences in repair integration between groups. Furthermore, rhLubricin treated joints had significantly lower friction coefficients tested across three sliding speeds compared to saline treated joints (rhLubricin: 0.15 {+/-} 0.03 at 0.1 mm/s to 0.12 {+/-} 0.03 at 10 mm/s, Saline: 0.22 {+/-} 0.06 at 0.1 mm/s to 0.19 {+/-} 0.05 at 10 mm/s). Overall, a single delayed injection of rhLubricin improved the quality and lubricating ability of the repair cartilage tissue without inhibiting repair tissue integration.

bioengineering↗

bioRxiv2024

Leucine zipper-based SAIM imaging identifies therapeutic agents to disrupt the cancer cell glycocalyx for enhanced immunotherapy

The abnormally thick glycocalyx of cancer cells can physically limit immune-cell engagement and reduce the efficacy of cellular immunotherapies, but strategies for identifying pharmacological agents that relieve this barrier remain limited. Here, we establish glycocalyx thickness as a pharmacologically actionable physical phenotype. Disruption of mucin-type O-glycosylation reduced glycocalyx thickness and increased NK- and CAR-NK-mediated killing, demonstrating that pharmacological glycocalyx remodeling can enhance innate and CAR-directed cytotoxicity. To screen this phenotype directly, we developed a scanning angle interference microscopy (SAIM) platform using complementary leucine zipper pairs as a stable extracellular plasma-membrane reference. A focused screen of glycosylation and metabolic inhibitors resolved compound- and cell-type-specific nanoscale changes in glycocalyx thickness and identified several thinning agents. Castanospermine, an inhibitor of early N-glycan processing, was among the strongest hits and reduced mucin-probe binding in an independent mucin-rich gastric cancer model. Castanospermine-treated gastric cancer cells retained MUC17 antibody recognition and showed increased physical avidity for MUC17 CAR-T cells, while castanospermine enhanced MUC17 CAR-T-mediated tumor control in real-time co-culture assays. These findings establish direct glycocalyx-thickness measurement as a physical-phenotype-guided screening strategy and identify castanospermine as a candidate modulator for combination with MUC17 CAR-T cells.

bioengineering↗

bioRxiv2024

Recombinant manufacturing of multispecies biolubricants

Lubricin, a lubricating glycoprotein abundant in synovial fluid, forms a low-friction brush polymer interface in tissues exposed to sliding motion including joints, tendon sheaths, and the surface of the eye. Despite its therapeutic potential in diseases such as osteoarthritis and dry eye disease, there are few sources available. Through rational design, we developed a series of recombinant lubricin analogs that utilize the species-specific tissue-binding domains at the N- and C-termini to increase biocompatibility while replacing the central mucin domain with an engineered variant that retains the lubricating properties of native lubricin. In this study, we demonstrate the tissue binding capacity of our engineered lubricin product and its retention in the joint space of rats. Next, we present a new bioprocess chain that utilizes a human-derived cell line to produce O-glycosylation consistent with that of native lubricin and a purification strategy that capitalizes on the positively charged, hydrophobic N- and C-terminal domains. The bioprocess chain is demonstrated at 10 L scale in industry-standard equipment utilizing commonly available ion exchange, hydrophobic interaction and size exclusion chromatography resins. Finally, we confirmed the purity and lubricating properties of the recombinant biolubricant. The biomolecular engineering and bioprocessing strategies presented here are an effective means of lubricin production and could have broad applications to the study of mucins in general.

bioengineering↗

bioRxiv2022

Mucins form a nanoscale material barrier against immune cell attack

The cancer cell glycocalyx serves as a major line of defense against immune surveillance. However, how specific physical properties of the glycocalyx contribute to immune evasion and how these properties are regulated are not well understood. Here, we uncover how the surface density, glycosylation, and crosslinking of cancer-associated mucins contribute to the nanoscale material thickness of the glycocalyx, and further analyze the effect of the glycocalyx thickness on resistance to effector cell attack. Natural Killer (NK) cell-mediated cytotoxicity exhibits a near perfect inverse correlation with the glycocalyx thickness of target cells regardless of the specific glycan structures present. NK cells expressing a chimeric antigen receptor (CAR) have an enhanced ability to breach the glycocalyx and kill target cells. Equipping the NK cell surface with a mucin-digesting enzyme also improves killing with a performance enhancement that rivals or exceeds CARs in some cases. Together, our results provide new considerations for improving cancer immunotherapies.

biophysics↗