played a larger role in super resolution microscopy by lowering the technical barrier to entry. This is possible as Deep Learning algorithms can learn the non-linear function of transforming diffraction-limited images obtained on traditional fluorescent confocal microscopes into their super-resolved counterparts. Generally, these algorithms have required complex encoder-decoder architectures and more recent efforts include complex generative adversarial networks. While useful, these architectures hinder interpretability, demand higher energy consumption, and require high volumes of memory on devices. Herein, we investigate the performance of a simple three-layer convolutional neural network and compare it with a well cited encoder-decoder architecture.

processes. Currently, however, it is primarily produced from petrochemicals via an unsustainable process with high greenhouse gas intensity. Alternatively, malic acid can be microbially produced from renewable biomass sugars via anaerobic fermentation by Escherichia coli. In this carbon-negative process, dissolved inorganic carbon (DIC) serves as an essential co-substrate but there are several limitations associated with its conventional delivery. Typically, bicarbonate salts are used, but their generation is both wasteful and energy intensive. Alternatively, direct delivery of CO2 gas would be advantageous for promoting a circular carbon economy and mitigating climate change. To this end, we systematically developed a series of novel membrane bioreactors that deliver CO2 gas using polymeric hollow fiber membranes (HFMs) in a bubbleless and on-demand manner. A CO2 transfer rate model was developed and utilized to predict dissolution rates of total DIC using HFMs, whereas relevant process conditions were investigated to optimize fermentation performance. At a CO2 delivery pressure (CO2DP) of 2.5 PSIG and HFM specific surface area of 7.4 m-1, a maximum malate titer of 36.4 g/L was achieved at a yield of 0.96 g-malate/g-glucose, both comparable to the performance of a conventional batch fermentation. Meanwhile, by performing a total carbon balance on the system, it was further determined that a ~50% reduction in carbon waste due to off-gassing was possible for the membrane bioreactor compared to a conventional batch process.

at the cell type and single-cell level. Epigenetic heterogeneity is particularly prevalent in breast cancer. Our lab focuses on Triple Negative Breast Cancer (TNBC), the most aggressive breast cancer subtype with a survivability that is worsened among African Americans when compounded by obesity. The molecular relationship between obesity and TNBC is an active area of research with epigenetic repression being identified as a promising biomarker. While several molecular techniques are used to study the functional impact of epigenetic repression, there are limited genetic engineering techniques that allow for live-cell tracking of epigenetic repression with single-cell resolution. To study how the adipocyte-secretome impacts TNBC epigenetic states, we engineered cyan fluorescent protein (CFP) expressing BT549 cells regulated by a doxycycline inducible tet-3XVP16 activator and monitored CFP repression in cells treated with adipocyte-conditioned media (ACM). ACM treatment significantly decreased (p<0.0001) CFP expression in lipogenic cells compared to control groups. The transcriptome of ACM-treated versus control-treated BT549 cells showed the upregulation of genes including oncogenic and lipogenesis pathways, and the downregulation of genes including tumor suppressor genes (FC>=2). This ACM-associated transcriptomic profile aligns with phenotypes of enhanced proliferation, survival, invasion, and lipid synthesis. These data provide insights into how metabolic (i.e lipogenesis) and epigenetic fluctuations influence transgene behavior. Furthermore, these results provide a proof-of-concept for the use of a fluorescent reporter to track TNBC epigenetic states at the single-cell level.

the primary treatment to improve symptoms. We believe that satellite cells (SCs) mediate recovery during SET. SCs are quiescent skeletal muscle stem cells that repair muscle when activated by stimuli (exercise). We hypothesize that activated SCs secrete angiogenic factors inducing collateral vessels due to their adjacency to skeletal muscle capillaries. 40 percent of PAD patients are affected by hyperglycemia, and this impairs muscle cells. We look to investigate SCs' role in angiogenesis, and the effects of hyperglycemia on this niche. 
Methods: SCs angiogenic potential was investigated through histology, cell migration, and ELISA. Mouse aortic smooth muscle (MASM) and endothelial cells (MAEC) migration towards the stimuli: SC conditioned media (CM), DMEM (positive control), Serum free DMEM (control), Denatured CM, was measure in a transwell system. We measured VEGF secretion via ELISA for these same stimuli. Balb/C mice were treated with SCs in alginate, empty alginate, and saline following hind limb ischemia and blood vessel growth was measured with histology. 
Results & Conclusion Migration increased with CM (38.09 ± 27.21, n=8) (*p<0.05) compared to control groups (12.52 ± 12.74, n=8) for MASMs with similar trends for MAECs (CM, 119.75 ± 70, control, 29.7 ± 11.75, n=2). Satellite cell treated mice showed an increase of capillary number (136.36 ± 70.7) (*p<0.05) compared to saline (55.76 ± 28.82). ELISA demonstrated that excess glucose in cell culture increases SC VEGF production in conditioned media (High Glucose CM, 741 pg/mL ± 48.75, CM, 56.15 pg/mL ± 22.15).

supporting immune function. Breast cancer cells invade the lymphatics through lymph vessels in the breast tissue, subsequently entering lymph nodes and traveling to distant metastatic sites. O-linked glycosylation, the attachment of sugars to serine or threonine residues on membrane-anchored macromolecular backbones, plays a pivotal role in mediating cell-cell and cell-extracellular matrix interactions. In cancer, aberrant O-linked glycosylation is associated with an increased production of extracellular vesicles (EVs), promoting metastasis. Using nanoparticle tracking analysis, extracellular vesicles were characterized between triple negative breast cancer (TNBC) breast cancer cell lines, MDA-MB-231 and MDA-MB-468 to confirm concentration and expected size ranges. We then exposed lymphatic endothelial cells (LECs) to EVs from the TNBC cell lines. Both of the TNBC cell lines exhibit elevated expression of the known aberrant glycosylation signatures in cancer, specifically T/Tn antigens and sialylation. Alterations in LEC marker podoplanin and O-linked glycosylation levels were also assessed to examine the role of EVs in mediating tumor-lymphatic interactions in the metastatic cascade. Collectively, these findings underscore the role of aberrant glycosylation patterns in TNBC and LECs. We are currently aiming to understand the role of glycosylation and EVs in tumor-lymphatic crosstalk in breast metastasis to other niches, such as the brain and bone. Overall, this research will be provide insights on mechanisms driving cancer dissemination via lymphatics and to identify potential therapeutic targets in breast metastasis.

protein engineering. By providing additional stabilization through synthetic polymer scaffolds, such as dendrimer colloidal , mechanical, thermal, and viscoelastic properties can be fine-tuned. Herein, smaller generations of poly(amidoamine) (PAMAM) dendrimers serve as low molecular weight gelators interacting with bovine serum albumin (BSA) via electrostatic interactions and subsequently form stable physically crosslinked hydrogel networks upon thermal annealing. Interestingly, adding a reinforcing network through chemical crosslinking with glyoxal via Schiffbase reactions resulted in weakened mechanical and viscoelastic behaviors. Opportunely, potentially new optical properties have been observed, highlighting the versatility of these hydrogels serving broader applications within theranostic nanomedicine. It was concluded that maintaining the viscoelastic properties of empty hydrogels was dependent on entrapped therapeutic payload concentrations. Higher concentrations of doxorubicin resulted in weaker viscoelastic behaviors. Highly porous microstructures were examined to assess the potential loading of small molecules and nanoparticles. BSA-PAMAM hydrogels were degraded within 24 hours utilizing proteinase K during enzymatic degradation experiments, which support future in vivo model studies. Water-soluble tetrazolium 1 (WST) cell viability assay of human embryonic kidney (HEK) 293 cells was determined to be at or above 80% for both physically and chemically crosslinked hydrogel networks. BSA-PAMAM hydrogels highlight the advantages of combining natural and synthetic polymers to produce reinforced soft matter materials exhibiting favorable biocompatibility, high protein content, and accessible modifiability that can be broadly applied across biomedical research applications.

hydrogen peroxide. Cholesterol oxidase is an industrially importantenzyme used in the clinical determination of serum cholesterol levels. CO enzyme is used in the biological processes involving conversion of different steroidal and nonsteroidal compounds. Moreover, CO is used as an insecticide in transgenic crop-pest management. CO enzyme biosensors are applied in cholesterol level detection in various samples and in investigating cell membrane interactions with cholesterol. Due to its vast range of applications, the industrial importance and demand of CO have gained increased interest. Current work describes the isolation of a potential cholesterol oxidase producing streptomycete from Egyptian soil. The isolated strain produced cholesterol oxidase in submerged culture using a medium containing glucose, yeast extract, malt extract, and CaCO3 with the addition of cholesterol as an inducer. The isolated strain was identified as Streptomyces rochei NAM-19 based on 16S rRNA sequencing and phylogeny. Optimization of cholesterol oxidase production has been carried out using response surface methodology. The Plackett-Burman design method was used to evaluate the significant components of the production medium followed by Box-Behnken experimental design to locate the true optimal concentrations, which are significantly affecting enzyme production. Results showed that the predicted enzyme response could be closely correlated with the experimentally obtained production. Furthermore, the applied optimization strategy increased volumetric enzyme production by 2.55 times (65.1 U/mL) the initial production obtained before medium optimization (25.5 U/mL).

offer a viable substitute for conventional packed bed chromatography. To efficiently separate proteins, this study functionalized DEAE anion-exchange ligands on cellulose acetate nanofibers, which were hydrolyzed to regenerate cellulose and then used to create an ion-exchange medium. Standard commercially available regenerated cellulose membranes, microfiber mediums, and bleached cotton balls were used as comparisons for the nanofiber felts. When it came to bovine serum albumin (BSA), the DEAE-functionalized nanofiber felt showed an exceptional static binding capacity of 40.0 mg/g, greatly increasing potential yield and purity in the manufacture of therapeutic proteins. Additionally, it demonstrated permeability for Tris buffer (pH 8.0) that was more than five times greater. By adding more felt layers, it effectively reduced system dispersion and maximized throughput without lowering system pressure. In the nanofiber felt, dynamic adsorption of BSA at 10% breakthrough was likewise more effective (26.9 mg/g) than the commercial alternative (20.9 mg/g), suggesting its potential to enhance performance in important bioseparations related to health.

aqueous humor outflow from the eye, causing an increase in intraocular pressure that gradually damages the optic nerve. The trabecular meshwork (TM) is the main tissue implicated in outflow dysfunction, and increased TM stiffness has been associated with high intraocular pressure. Outflow through the TM is non-uniform, meaning there are areas of high flow (HF) and low flow (LF) throughout the tissue. Previous work in human donor eyes has shown that low-flow (LF) regions of the TM are stiffer than high-flow (HF) regions; however, differences in stiffness between HF and LF regions in animal models are unknown. Here we investigate differences in stiffness between HF and LF regions of 4-month-old wild-type C57BL/6 mice. Atomic force microscopy (AFM) force mapping was used to measure Young’s modulus on cryosections from HF and LF regions of the TM. Consistent with results in humans, LF regions were stiffer than HF regions (Young’s modulus = 23.8 ± 14.0 kPa in LF regions vs. 13.9 ± 4.59 kPa in HF regions). However, results showed significant variability among mice; in the two females, LF regions were stiffer, while in the male, the HF region was stiffer (p < 0.001). These data provide a baseline for studying stiffness differences between HF and LF regions in a mouse model of glaucoma, and more animals will be tested to investigate sex-specific differences and identify sources of variability.

microorganisms including Candida (C.) albicans, human immunodeficiency virus (HIV), and severe acute respiratory syndrome coronavirus 2 (SARS‑CoV‑2). FDG Abs were found in HIV-naïve humans, and rhesus macaques (RMs) that were HIV vaccinated or infected with Simian-HIV (SHIV), but the ontogeny of these Abs was not fully defined. Previous studies have postulated that intestinal microbiota (IM) antigens may prime B cells that can be recruited in response to HIV vaccination or infection. Thus, the purpose of this study is to identify glycosylated IM antigens that may prime the induction of FDG B cell lineages.
Methods. We tested FDG Abs from HIV-naïve human (DH1005), SHIV-infected (DH851.3) and HIV vaccinated (DH717.1) RMs to IM and C. albicans lysates via western blot. We will perform immunoprecipitation (IP) with FDG Abs and IM-WCLs and C. albicans lysates using a commercially-available kit followed by mass spectrometry analysis.
Results. Only DH1005 bound IM antigens, whereas DH1005, DH717.1, and DH851.3 bound antigens in both IM and C. albicans lysates. FDG Abs bound antigens of varied molecular weights as observed via western blot, but we will perform IP followed by mass spectrometry to define these antigens.
Conclusions. These data raised the hypothesis that glycosylated antigens in IM and C. albicans lysates bound by FDG Abs may influence the development of these Abs. Moreover, the results of our studies may provide insights for future vaccine regimens against glycosylated pathogens.

including melanoma and breast cancer. Melanoma, a type of skin cancer, and triple-negative breast cancer (TBNC), an aggressive form of breast cancer, are both detected at an early stage. However, TBNC tends to be more aggressive and has a higher likelihood of rapid growth and metastasis. Immunotherapy has transformed treatments for both cancers but has yet to reach full efficacy. A potential barrier to the efficacy of approved immunotherapies is the glycocalyx barrier around the cell, which interferes with the ability of T cells to effectively attack cancer cells. This study focuses on identifying a way to break down the glycocalyx barrier, allowing T cells to access cancer cells. We hypothesize that treating cancer cells with chondroitinase, hyaluronidase, heparinase or neuraminidase will reduce impairments caused by the glycocalyx layer, allowing for better T cell-mediated cancer cell killing. We utilized confocal imaging to measure the glycocalyx degradation after enzyme introduction. This demonstrated a significant reduction of the glycocalyx layer around cancer cells, implying that enzymes effectively break down the barrier. This research can improve immunotherapies for melanoma and breast cancer by showing a more effective method for T cells to reach cancer cells. Reducing the glycocalyx can improve T cell-mediated destruction of cancer cells, resulting in long-term survival for patients with breast cancer and melanoma.

therapies, immunotherapy is gaining significant attention as a promising option for the development of therapeutic treatments for ovarian cancer research. Nanoparticles are used to transport cytokines that induce a proinflammatory immune response into tumor micro-environments. In this study, highly soluble liposomal based nanoparticles deliver Interleukin-12 (IL-12) to activate immune cells while reducing toxicity and increase biocompatibility. It is suggested that polymeric layer by layer methods and highly effective IL-12 conjugation techniques are linked to prolonged cytokine retention which improves stability and increases localization for an increased activation of immune cells. This study focuses on comparing different molecular conjugation techniques of Nickel-His and Cobalt chemistries by observing nanoparticle’s size, polymeric layer by layer loading efficiency and protein retention to test their stability in media. By rigorously evaluating and refining the covalent chemistries of nanoparticles, the efficacy and safety of targeting ovarian cancer cells can be significantly enhanced.

biogas production through anaerobic digestion, but BMP tests are complex and time-consuming. To address this challenge, we developed a random forest regression model to predict BMP for a wide range of agricultural biogas feedstocks based on their fresh mass (FM) cellulose, hemicellulose, and lignin contents, offering insights into their influence on biomethane production. The model was trained on data sourced from the comprehensive database developed by Lallement et al. (2023). The model accuracy was evaluated on independent test datasets using key metrics (R², RMSE, and MAE). The model’s learning curve was assessed using a 5-fold cross-validation technique. Shapley Additive Explanations (SHAP) analysis was done to provide insights into the importance of input variables in the model's predictions. The model demonstrated strong performance on the test dataset, with a high R2 value (0.84), low RMSE (284.31 Nm³ CH₄/ton FM), and low MAE (171.66 Nm³ CH₄/ton FM), confirming its reliability and applicability. The learning curve shows no overfitting or underfitting, demonstrating the model's robustness and capacity to generalize. SHAP analysis identified hemicellulose as the most significant factor in the BMP prediction, with lignin and cellulose also playing important roles. The developed model offers a practical and efficient tool for the biogas industry, eliminating the need for extensive laboratory experiments, saving time and resources, and allowing for better decision-making in feedstock selection and process optimization, ultimately enhancing the efficiency and sustainability of biogas production.

polysaccharides, such as cellulose and chitin. Hence, protein engineering is highly required to enhance their catalytic efficiencies. To this effect, we optimized the protein sequence encoding for an LPMO from Bacillus amyloliquefaciens (BaLPMO10A) using the sequence consensus method. Enzyme activity was determined using the chromogenic substrate 2,6-Dimethoxyphenol (2,6-DMP). Compared with the wild type (WT), the variants exhibit up to a 93.7% increase in activity against 2,6-DMP. We also showed that BaLPMO10A can hydrolyze p-nitrophenyl-β-D-cellobioside (PNPC), carboxymethylcellulose (CMC), and phosphoric acid-swollen cellulose (PASC). In addition to this, we investigated the degradation potential of BaLPMO10A against various substrates such as PASC, filter paper (FP), and Avicel, in synergy with the commercial cellulase, and it showed up to 2.7-, 2.0- and 1.9-fold increases in production with the substrates PASC, FP, and Avicel, respectively, compared to cellulase alone. Moreover, we examined the thermostability of BaLPMO10A. The mutants exhibited enhanced thermostability with an apparent melting temperature increase of up to 7.5 C compared to the WT. The engineered BaLPMO10A with higher activity and thermal stability provides a better tool for cellulose depolymerization.

important cellular functions. Evolved to be histone-PTM specific, histone binding domains (HBD) from chromatin reader-effector proteins are gaining more attention as modules to alter and probe the epigenome beyond antibodies. Chromatin marks change during solid tumor progression, making them an important biomarker in solid cancer studies. A repressive chromatin mark known as H3K27me3 has been associated with triple negative breast cancer, the most aggressive form of breast cancer that has shown resistance to chemotherapy. The Haynes lab intends to engineer histone-binding probes (HBPs) that identify H3K27me3 within chromatin. However, it is time-consuming to test various HBP designs since each probe needs to be constructed in total before being tested. Here, we have integrated coiled-coil (CC) proteins into the probe design to allow the reader and probe domains to be engineered separately before being incorporated into cells. We fused HBPs to coiled coil proteins to increase design flexibility while demonstrating high affinity and specificity for the H3K27me3 chromatin mark. We show dimerization of CC proteins results in nuclear localization of CFP in human cells.

including the ability to reliably segregate during cell divisions due to having a functional centromere. However, HACs have persisting limitations, including random multimerization of input sequences required to form a functional centromere. In response to this issue, Gambogi et al. reported use of a precursor yeast artificial chromosome (YAC) sufficient in size to form HACs without multimerization, effectively creating single-copy HACs. However, HACs still have complications including AT richness, which makes sequencing and PCR difficult, and limited gene expression. As such, we are shifting toward using a large YAC with stuffer DNA to facilitate sequencing and eventual HAC optimization. For iterative HAC debugging, we would retrofit Saccharomyces cerevisiae carrying this YAC using homologous recombination along with selections for and counter-selections against the URA3 auxotrophic marker. The native URA3 is currently knocked out, but a transposon disrupts it. This causes it to recombine with any URA3-based elements incorporated into the YAC, effectively precluding YAC modification. I hypothesize that the YAC can be effectively optimized by deleting the native URA3 to prevent unwanted recombination events. When combined with URA3 selection and counter-selection, this would allow for iterative engineering with fewer limitations. We first successfully repaired the native URA3, then attempted to delete it from the yeast genome. After successful deletion of URA3 and reincorporation into the YAC along with HAC formation and optimization components, future directions include creating HACs and verifying their stability.

immune-related disorder treatment1. A critical challenge with this approach is the selective delivery of RNAs to organs where target cells reside2. For instance, macrophages are prominent cellular targets of anti-tumor immunotherapies due to their broad therapeutic effector functions3,4. However, selective RNA delivery to lymphoid organs where macrophages reside poses a significant barrier to macrophage-reprogramming gene therapy development. Existing FDA-approved lipid nanoparticle (LNP) formulations primarily target the liver when administered systemically, limiting IVT-mRNA's broad applicability as a systemic therapy2. To address these challenges, we conducted a multi-tier library screening of 162 mRNA-nanoparticle formulations based on dendrimer-lipid hybrid materials (dLNPs). From this screening, we aim to identify nanoformulations with the greatest therapeutic potential by assessing macrophage transfection in vitro and nanoparticle biodistribution in vivo. RESULTS: In vitro library screening identified 12 lead formulations (9 novel) with comparable or superior transfection efficiency and safety to clinically relevant formulations. In vivo biodistribution studies revealed 5 formulations with tropism for myeloid cell tissue reservoirs, such as the spleen. Spleen-targeting dLNPs contained internal secondary amines, while liver-targeting dLNPs contained solely tertiary amines. Two formulations showed superior targeting and transfection efficiency compared to clinically relevant SM-102 and SORT-lipid modified DLin-MC3-DMA. CONCLUSIONS: Our findings establish a structure-function relationship between dendrimer internal amine structures and selective in vivo targeting. These insights inform rational nanoformulation design, enhancing the potential for extrahepatic targeted RNA delivery for cancer and immune-related disorder treatment.

modeling the effects of toxins and drugs. Currently, the FDA as an organization has limited experience in evaluating preclinical data based on experimentation in 3D in-vitro models. Our research teams, funded by the Department of Defense (DoD), are currently developing strategies for generating stable and physiologically relevant human micro-engineered organ equivalents (MOEs) for majority of tissue types. Our group is more focused on MOE models for liver cells using Indium-Tin Oxide (ITO) as a scaffold for drug toxicity screening in a Self-Assembled Monolayer (SAM), using 2D and 3D approaches. Preliminary studies on this scaffold will be presented, along with self-assembled monolayer modifications and metabolomic data tracking the changes in the cells and plates over time using Nuclear Magnetic Resonance (NMR) Metabolomics. NMR spectroscopy is a non-invasive and nondestructive technique which gives a detailed insightful observation of intracellular responses and hepatotoxicity study using the passaged medium containing cellular exudates.

chronic pain. While conventional amino-amide and amino-ester local anesthetics are effective, their typical nerve block or infiltration duration is relatively short, around 2-3 hours, due to rapid clearance. In addition, all clinically used local anesthetics act on both sensory and motor neurons, resulting in motor function loss alongside pain relief. To address these challenges, we developed capsaicin-loaded melanin nanoparticles (Cap-MNPs). Capsaicin selectively acts on sensory neurons without affecting motor neurons, thereby achieving sensory-selective nerve blockade. Melanin, known for its exceptional biocompatibility, biodegradability, and abundance in pigmented human tissue, serves as a carrier for capsaicin for sustained release. Melanin's chemical structure enables the encapsulation and sustained release of capsaicin through π-π interactions. The drug loading efficiency of Cap-MNPs was 82.99±1.55%, the drug loading capacity was 67.47±4.24%, and capsaicin was continuously released for more than 360 hours. In rats, a single injection of Cap-MNPs containing 8.04 mg of capsaicin produced a sciatic sensory nerve block lasting for 6 hours without causing any motor block, local toxicity, and capsaicin-related systemic toxicity. Cap-MNPs show promise as clinically useful therapeutics for pain management.

(strain ATCC 27853), and Acinetobacter baumannii (strain ATCC-BAA-1605) have been classified as extremely drug-resistant (XDR) by the World Health Organization (WHO). This study hypothesised that rhizospheric microbiota of plants surviving in metal-laden environments, with extreme environmental stress and limited nutrients, would have developed resilience due to competitive exclusion. This resilience, could in part, be due to the secretion of antimicrobials, which may be potent enough to inhibit the growth of XDR pathogens. In order to investigate this hypothesis, our study evaluates antimicrobial secretions from heavy-metal-contaminated rhizospheres which have been exposed to further unnatural environmental stresses, using novel methodology. Crude extracts of secondary metabolites were subsequently tested against the pathogens using the agar well diffusion test. Results indicate that these secretions inhibit both XDR pathogens to the value of 50 mm where the multi-drug-resistant pathogens are at a concentration of 1.0 McFarland (about 3.0 × 108 CFU/mL). Moreover, pathogens were more responsive to exudates of microbiota from environmentally stressed rhizospheres than microbiota from an organic rhizosphere. The components of the extracted secondary metabolites were tentatively identified using HPLC-MS/UV. Ultimately, the novelty of the secondary metabolites needs to be validated through UPLC-MS and NMR, and the calculated to warrant further investigations on their pivotal contributions to biomedicine and biopharmaceutics. Keywords: Antimicrobial; Antibiotics resistance; Antibiotics susceptibility; Competitive exclusion; Environmental stress; Rhizospheric microorganisms; Pseudomonas aeruginosa; Acinetobacter baumannii; Bacillus siamensis; Bacillus amyloliquefaciens.

both atmospheric and aqueous environments. Microelectromechanical systems (MEMS) have emerged as a widely utilized platform for the development of portable and low-power chemical sensors. The chemical sensor employed in this study is a MEMS device that utilizes a resonant cantilever with a polymer coating for the purpose of detecting volatile organic compounds (VOCs) in the gas phase. The cantilever is produced using microfabrication techniques. The inclusion of a thick polymer sensing film on the surface of the microfabricated sensor enhances the sensitivity of the sensor. However, it also introduces a drawback in the form of reduced sensor reaction time, as the analyte must diffuse through the comparatively thick sensing film. The primary objective of this research is to alter the surfaces of the microsensors, which are based on silicon, by including nanoscale features created by 3D printing. The final aim is to develop sensing films that possess both exceptional sensitivity and rapid reaction time. Consequently, the utilization of high-surface area, nano-structured sensing films is a viable approach to enhance reaction times without compromising sensor sensitivity. The present invention demonstrates the capability to generate intricate three-dimensional (3D) patterns with exceptional precision within a photoresist devoid of solvents, utilizing a UV light-sensitive polymer. The study employed a 3D direct-write lithography method to fabricate high-surface-area nano-patterns directly onto the sensor surfaces of silicon-based sensors.

the most common treatment, often results in poor long-term outcomes, with insufficient defect fill and fibrous tissue formation leading to repair failure and progression of osteoarthritis. While strategies such as bioactive factors and scaffolds aim to enhance MFx outcomes, early clot remodeling remains understudied. This study investigated the potential of platelet lysate (PL), an off-the-shelf orthobiologic, to improve volumetric maintenance and cellular activity in cartilage repair both in vitro and in vivo.
Methods: Macro-Scale Fibrin Gel Studies (100 µL): Fibrin gels (25 mg/mL fibrinogen) were formed with thrombin, calcium chloride, and/or platelet lysate (PL; Stem Cell Technologies), gelled for 60 minutes, imaged via SEM, and mechanically tested. For contraction analysis, hMDCs (2M cells/mL; Lonza) were embedded, cultured in basal media with aprotinin (100 KIU/mL) and TGF-β3 (1 ng/mL) for 14 days, and imaged to quantify contraction.
Fibrin Microgel Studies (10 µL): hMDCs (200k/mL; Lonza) were seeded in fibrin gels with or without PL (Stem Cell Technologies), cultured for 3 days with aprotinin (100 KIU/mL) and TGF-β3 (1 ng/mL), then fixed, stained (Phalloidin, Ki-67, SOX9, α-SMA), and imaged by confocal microscopy to assess early cell behavior.
In Vivo Evaluation: In a pig model, three 6 mm trochlear defects were created per knee and treated with MFx, comparing a control and two PL formulations. Tissues were collected at 5 weeks for mechanical testing, micro-CT, and histology.
Results: PL supplementation altered MDC morphology, promoting a larger, rounder shape (Figure 1A, D-E), and increased proliferation, as shown by Ki-67 staining (Figure 1B-C). SOX9 intensity decreased, while α-SMA expression was reduced, indicating lower chondrogenic activity and fibrotic potential, respectively (Figure 1F-G). In vivo, mechanical properties of defects were not improved with PL treatment (Figure 2A-C). Histological analysis revealed no differences in matrix deposition between treatment groups (Figure 3A), and ICRS II scores were comparable across animals (Figure 3B-C).
Discussion: PL supplementation influenced MDC morphology and proliferation while reducing fibrotic potential, as indicated by decreased α-SMA expression. However, the reduction in SOX9 intensity and lack of improvement in in-vivo mechanical properties or histological outcomes suggest further refinement is needed to optimize PL for cartilage repair applications.
References: [1] Erggelet+, J Clin Orthop, 2016. [2] Carluccio+, Cells, 2020
Acknowledgement: Emory Orthopaedics; Regenerative Engineering Medicine (REM).

and function. The implications of fibrotic healing highlight the need for a model to study the mechanisms supporting tendon healing in a fibrosis-free environment. Spiny mice are the only known mammals capable of regenerating large tissue defects without fibrosis, as observed in scar-free healing of skin and muscle. However, the mechanism of tendon homeostasis and healing in this regenerative mouse model is unknown. This study examines tendon homeostasis in spiny mice compared to CD1 mice by analyzing the biomechanical and histological properties of healthy male and female Achilles tendons. Achilles tendons from adult spiny and CD1 mice were subjected to uniaxial tensile testing and histological analysis. Ultimate tensile stress (UTS) and Young’s modulus (E) were statistically compared between species and sexes (p < 0.05). 5-μm thick sagittal sections were stained with hematoxylin and eosin (H&E) to visualize tissue composition. Spiny mice tendons exhibited a 60-70% lower UTS and E than CD1 mice. Female spiny tendons had significantly lower mechanics than the female CD1 tendons. Histological scoring revealed no statistically significant differences in collagen fiber alignment, cellularity, or matrix integrity between spiny mice and CD1 mice (p > 0.05), indicating comparable tendon structure across species. These findings suggest that regenerative abilities in spiny mice may be linked to unique extracellular matrix composition. Identifying key regenerative mechanisms in spiny mice could lead to therapies to mitigate tendon fibrosis.

such as microfracture and autologous chondrocyte implantation, face challenges in achieving lasting functional recovery. A key issue is the dedifferentiation of chondrocytes in 2D culture, resulting in altered gene expression and mechanical properties. To address this, encapsulating chondrocytes in hydrogel matrices has emerged as a promising strategy. Heparin-based hydrogels are particularly noteworthy as they can sequester growth factors and modulate their release, thereby enhancing chondrogenic differentiation and matrix production. This study is the first to investigate the impact of heparin sulfation levels on chondrocyte maintenance in vitro. We utilized N-desulfated heparin (Hep-N) and fully desulfated heparin (Hep-) hydrogels as scaffolds to evaluate chondrocyte re-differentiation. Over four weeks, chondrocytes in both heparin groups demonstrated viability and dispersal, with upregulation of chondrogenic genes COL II and ACAN. Notably, matrix production of collagen II, aggrecan, and glycosaminoglycans occurred in both groups, but chondrocytes in Hep-N hydrogels exhibited a significant increase in aggrecan deposition. Furthermore, both hydrogel types accumulated endogenous BMP-2 and TGF-β1 during culture, with Hep- hydrogels showing greater TGF-β1 sequestration. This heparin hydrogel system has the potential to enhance existing autologous chondrocyte transplantation methods by promoting chondrogenic gene expression and matrix production in vivo.

tissue environments for biomedical research. This study focuses on the regeneration of biopolymers dissolved in ionic liquids (ILs) and their subsequent fabrication into materials with directional porosity. Ionic liquids, known for their ability to dissolve and modify natural biopolymers, offer a tunable platform for material processing that preserves biocompatibility while allowing structural control at the micro- and nanoscales of a desired biopolymer system. The regenerated biopolymers, processed using tailored phase separation and templating techniques, exhibit anisotropic porosity optimized for cell migration, proliferation, and differentiation assays. Directional porosity not only enhances nutrient transport and waste removal but also mimics the natural extracellular matrix, promoting more physiologically relevant cell behavior. This work demonstrates the potential of ionic liquid-based biopolymer regeneration in producing structured biomaterials that can be customized for various cell culture applications, offering a versatile tool for tissue engineering and regenerative medicine.

the environment. Though they contain multiple components, these pollutants contain varying concentrations of metal ions, such as cadmium, arsenic, mercury, and uranium. However, a major challenge in sensing and detection platforms is the lack of high-specificity receptors to target heavy metal ions. Therefore, to develop new methods to detect metal ions, we look at aptamers, which are short protein or nucleic acid molecules. Aptamers provide a cheap, highly specific, and sensitive method to bind to target analytes, such as metal ions. However, heavy metal ion-specific aptamers lack effective recognition sites. It is challenging for aptamers to specifically bind and recognize metal ions in the same group due to similar structures of different metal ions and the simple structure and single binding site for heavy metal ions. This leads to poor specificity of the aptamer sequence to the target ion. The goal of this work is to develop theoretical approaches for aptamer screening to target lead ions (Pb2+) to address this challenge. Two G-quadruplex structures of lead-specific aptamers have been crystalized and shared as Protein Data Bank (PDB) files (7D31 and 7D32). Overall, our objective is to observe the conformational changes of the aptamer sequences for Pb2+ to understand the influence of the stability and conformational changes of aptamers with G-quadruplex structures. Our strategy is to (1) mutate 3D aptamers based on the sequences that were unable to be crystalized with a TT or TGT sequence, (2) execute molecular dynamic (MD) simulations to characterize changes of the binding site of aptamers in varying concentrations of salt water, and (3) execute MM/QM simulations to study the interactions of a DNA aptamer with a lead ion. Recent papers have published the sequence of a G-quadruplex structure for a lead-specific aptamer. The TT and TGT sequences are important for the crystallization and stability of the aptamer. This theoretical method provides evidence of the stability and structural changes of aptamer sequences unable to be crystalized without the TT and TGT sequences. These results may play a role in increasing the potential applications of aptamers in new sensing strategies for metal ions.