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Acta Biomaterialia Apr 2024Cardiac pacing with temporary epicardial pacing wires (TEPW) is used to treat rhythm disturbances after cardiac surgery. Occasionally, TEPW cannot be mechanically...
Cardiac pacing with temporary epicardial pacing wires (TEPW) is used to treat rhythm disturbances after cardiac surgery. Occasionally, TEPW cannot be mechanically extracted and remain in the thorax, where they may rarely cause serious complications like migration and infection. We aim to develop bioresorbable TEPW that will dissolve over time even if postoperative removal is unsuccessful. In the present study, we demonstrate a completely bioresorbable design using molybdenum (Mo) as electric conductor and the resorbable polymers poly(D, L-lactic-co-glycolic acid) (PLGA) and polycaprolactone (PCL) for electrically insulating double-coating. We compared the pacing properties of these Mo TEPW demonstrators to conventional steel TEPW in Langendorff-perfused rat hearts and observed similar functionality. In vitro, static immersion tests in simulated body fluid for up to 28 days elucidated the degradation behaviour of uncoated Mo strands and the influence of polymer coating thereon. Degradation was considerably reduced in double-coated Mo TEPW compared to the uncoated and the PLGA-coated condition. Furthermore, we confirmed good biocompatibility of Mo degradation products in the form of low cytotoxicity in cell cultures of human cardiomyocytes and cardiac fibroblasts. STATEMENT OF SIGNIFICANCE: Temporary pacing wires are routinely implanted on the heart surface to treat rhythm disturbances in the days following cardiac surgery. Subsequently, these wires are to be removed. When removal attempts are unsuccessful, wires are cut at skin level and the remainders are left inside the chest. Retained fragments may migrate within the body or become a centre of infection. These complications may be prevented using resorbable pacing wires. We manufactured completely resorbable temporary pacing wires using molybdenum as electrical conductor and assessed their function, degradation and biological compatibility. Our study represents an important step in the development of a safer approach to the treatment of rhythm disturbances after cardiac surgery.
Topics: Humans; Animals; Rats; Cardiac Pacing, Artificial; Pacemaker, Artificial; Molybdenum; Absorbable Implants; Pericardium
PubMed: 38432350
DOI: 10.1016/j.actbio.2024.02.039 -
Journal of Translational Medicine Mar 2024Cellular stress associated with static-cold storage (SCS) and warm reperfusion of donor lungs can contribute to ischemia-reperfusion (IR) injury during transplantation....
BACKGROUND
Cellular stress associated with static-cold storage (SCS) and warm reperfusion of donor lungs can contribute to ischemia-reperfusion (IR) injury during transplantation. Adding cytoprotective agents to the preservation solution may be conducive to reducing graft deterioration and improving post-transplant outcomes.
METHODS
SCS and warm reperfusion were simulated in human lung epithelial cells (BEAS-2B) by exposing cells to low potassium dextran glucose solution at 4 °C for different periods and then switching back to serum-containing culture medium at 37 °C. Transcriptomic analysis was used to explore potential cytoprotective agents. Based on its results, cell viability, caspase activity, cell morphology, mitochondrial function, and inflammatory gene expression were examined under simulated IR conditions with or without thyroid hormones (THs).
RESULTS
After 18 h SCS followed by 2 h warm reperfusion, genes related to inflammation and cell death were upregulated, and genes related to protein synthesis and metabolism were downregulated in BEAS-2B cells, which closely mirrored gene profiles found in thyroid glands of mice with congenital hypothyroidism. The addition of THs (T3 or T4) to the preservation solution increases cell viability, inhibits activation of caspase 3, 8 and 9, preserves cell morphology, enhances mitochondrial membrane potential, reduces mitochondrial superoxide production, and suppresses inflammatory gene expression.
CONCLUSION
Adding THs to lung preservation solutions may protect lung cells during SCS by promoting mitochondrial function, reducing apoptosis, and inhibiting pro-inflammatory pathways. Further in vivo testing is warranted to determine the potential clinical application of adding THs as therapeutics in lung preservation solutions.
Topics: Humans; Mice; Animals; Organ Preservation; Lung; Reperfusion Injury; Reperfusion; Epithelial Cells; Thyroid Hormones
PubMed: 38429788
DOI: 10.1186/s12967-024-05024-x -
Bone May 2024Human mesenchymal stem cells (hMSCs) sense and respond to biomechanical and biophysical stimuli, yet the involved signaling pathways are not fully identified. The...
INTRODUCTION
Human mesenchymal stem cells (hMSCs) sense and respond to biomechanical and biophysical stimuli, yet the involved signaling pathways are not fully identified. The clinical application of biophysical stimulation including pulsed electromagnetic field (PEMF) has gained momentum in musculoskeletal disorders and bone tissue engineering.
METHODOLOGY
We herein aim to explore the role of PEMF stimulation in bone regeneration by developing trabecular bone-like tissues, and then, culturing them under bone-like mechanical stimulation in an automated perfusion bioreactor combined with a custom-made PEMF stimulator. After selecting the optimal cell seeding and culture conditions for inspecting the effects of PEMF on hMSCs, transcriptomic studies were performed on cells cultured under direct perfusion with and without PEMF stimulation.
RESULTS
We were able to identify a set of signaling pathways and upstream regulators associated with PEMF stimulation and to distinguish those linked to bone regeneration. Our findings suggest that PEMF induces the immune potential of hMSCs by activating and inhibiting various immune-related pathways, such as macrophage classical activation and MSP-RON signaling in macrophages, respectively, while promoting angiogenesis and osteogenesis, which mimics the dynamic interplay of biological processes during bone healing.
CONCLUSIONS
Overall, the adopted bioreactor-based investigation platform can be used to investigate the impact of PEMF stimulation on bone regeneration.
Topics: Humans; Electromagnetic Fields; Transcriptome; Bone and Bones; Bone Regeneration; Bioreactors
PubMed: 38428556
DOI: 10.1016/j.bone.2024.117065 -
ACS Biomaterials Science & Engineering Mar 2024Encapsulating multiple growth factors within a scaffold enhances the regenerative capacity of engineered bone grafts through their localization and controls the...
Encapsulating multiple growth factors within a scaffold enhances the regenerative capacity of engineered bone grafts through their localization and controls the spatiotemporal release profile. In this study, we bioprinted hybrid bone grafts with an inherent built-in controlled growth factor delivery system, which would contribute to vascularized bone formation using a single stem cell source, human adipose-derived stem/stromal cells (ASCs) in vitro. The strategy was to provide precise control over the ASC-derived osteogenesis and angiogenesis at certain regions of the graft through the activity of spatially positioned microencapsulated BMP-2 and VEGF within the osteogenic and angiogenic bioink during bioprinting. The 3D-bioprinted vascularized bone grafts were cultured in a perfusion bioreactor. Results proved localized expression of osteopontin and CD31 by the ASCs, which was made possible through the localized delivery activity of the built-in delivery system. In conclusion, this approach provided a methodology for generating off-the-shelf constructs for vascularized bone regeneration and has the potential to enable single-step, in situ bioprinting procedures for creating vascularized bone implants when applied to bone defects.
Topics: Humans; Bioprinting; Tissue Engineering; Bone and Bones; Intercellular Signaling Peptides and Proteins; Stromal Cells
PubMed: 38416687
DOI: 10.1021/acsbiomaterials.3c01222 -
Applied Microbiology and Biotechnology Feb 2024Cell culture-based production of vector-based vaccines and virotherapeutics is of increasing interest. The vectors used not only retain their ability to infect cells but...
Cell culture-based production of vector-based vaccines and virotherapeutics is of increasing interest. The vectors used not only retain their ability to infect cells but also induce robust immune responses. Using two recombinant vesicular stomatitis virus (rVSV)-based constructs, we performed a proof-of-concept study regarding an integrated closed single-use perfusion system that allows continuous virus harvesting and clarification. Using suspension BHK-21 cells and a fusogenic oncolytic hybrid of vesicular stomatitis virus and Newcastle disease virus (rVSV-NDV), a modified alternating tangential flow device (mATF) or tangential flow depth filtration (TFDF) systems were used for cell retention. As the hollow fibers of the former are characterized by a large internal lumen (0.75 mm; pore size 0.65 μm), membrane blocking by the multi-nucleated syncytia formed during infection could be prevented. However, virus particles were completely retained. In contrast, the TFDF filter unit (lumen 3.15 mm, pore size 2-5 μm) allowed not only to achieve high viable cell concentrations (VCC, 16.4-20.6×10 cells/mL) but also continuous vector harvesting and clarification. Compared to an optimized batch process, 11-fold higher infectious virus titers were obtained in the clarified permeate (maximum 7.5×10 TCID/mL). Using HEK293-SF cells and a rVSV vector expressing a green fluorescent protein, perfusion cultivations resulted in a maximum VCC of 11.3×10 cells/mL and infectious virus titers up to 7.1×10 TCID/mL in the permeate. Not only continuous harvesting but also clarification was possible. Although the cell-specific virus yield decreased relative to a batch process established as a control, an increased space-time yield was obtained. KEY POINTS: • Viral vector production using a TFDF perfusion system resulted in a 460% increase in space-time yield • Use of a TFDF system allowed continuous virus harvesting and clarification • TFDF perfusion system has great potential towards the establishment of an intensified vector production.
Topics: Humans; Animals; HEK293 Cells; Vesicular Stomatitis; Vesicular stomatitis Indiana virus; Vesiculovirus; Cell Culture Techniques; Genetic Vectors
PubMed: 38413399
DOI: 10.1007/s00253-024-13078-6 -
Chemico-biological Interactions Mar 2024Acute kidney injury (AKI) is a disease characterised by acute onset, high mortality, and poor prognosis, and is mainly caused by ischemia-reperfusion (I/R). Human...
Acute kidney injury (AKI) is a disease characterised by acute onset, high mortality, and poor prognosis, and is mainly caused by ischemia-reperfusion (I/R). Human urine-derived stem cells (USCs) exhibit antioxidant, anti-inflammatory, and anti-apoptotic cytoprotective effects. Previously, we found that exosomes from USCs had the ability to inhibit apoptosis and protect kidneys from I/R injury. This study aimed to investigate the role of USC-derived exosomes (USC-Exos) in reducing pyroptosis and alleviating I/R-AKI. Models of HK-2 cells hypoxia-reoxygenation (H/R) and I/R kidney injury was established in Sprague Dawley rats to simulate AKI in vitro and in vivo. USC-Exos were isolated using ultracentrifugation and identified via electron microscopy and western blotting. USC-Exos were co-cultured with HK-2 cells and injected into rats via the tail vein. The expression of pyroptosis-related molecules (GSDMD, caspase-1, and NLRP-3) was verified using PCR and western blotting. Changes in renal function were reflected in the serum creatinine, urea, and cystatin C levels. The degree of renal injury was determined using haematoxylin and eosin and immunohistochemical staining. The levels of IL-1β and IL-18 were detected using enzyme-linked immunosorbent assay (ELISA) to verify the role of USC-Exos in pyroptosis. Differentially expressed circRNAs in I/R rat kidneys were screened by transcriptome sequencing, and a dual-luciferase experiment was used to verify the interaction between upstream and downstream molecules. Ischemia-reperfusion resulted in significantly impaired renal function and expression of pyroptosis molecules, and significantly increased concentrations of inflammatory factors. These effects were reversed by injecting USC-Exos. Circ DENND4C was the most significantly decreased circRNA in I/R rat renal tissue, and knock-down of circ DENND4C can aggravate AKI in vivo and in vitro. DAVID(http://david.abcc.ncifcrf.gov) website showed that miR 138-5p/FOXO3a is a potential downstream target of circ DENND4C. Knock-down of circ DENND4C in HK-2 cells resulted in increased expression of miR 138-5p and increased miR 138-5p can reverse the regulation of FOXO3a. Dual-luciferase assay verified the reverse interaction between circ DENND4C, miR 138-5p, and FOXO3a. Exosomes promote cell proliferation and inhibit the activation of NLR family pyrin domain containing 3 through the circ DENND4C/miR 138-5p/FOXO3a pathway, thereby reducing pyroptosis and AKI. Circ DENND4C may be a potential therapeutic target for AKI.
Topics: Animals; Humans; Rats; Acute Kidney Injury; Apoptosis; Exosomes; Ischemia; Kidney; Luciferases; MicroRNAs; Pyroptosis; Rats, Sprague-Dawley; Reperfusion; Reperfusion Injury; Stem Cells
PubMed: 38412628
DOI: 10.1016/j.cbi.2024.110922 -
Research Square Feb 2024Studies in adults have linked stress-related activation of the immune system to the manifestation of psychiatric conditions. Using a translational design, this study...
Studies in adults have linked stress-related activation of the immune system to the manifestation of psychiatric conditions. Using a translational design, this study aimed to examine the impact of social stress on immune activity in adolescents and on neuronal activity in a preclinical mouse model. Participants were 31 adolescents (ages 12-19), including 25 with mood and anxiety symptoms. Whole-blood samples were collected before and after the Trier Social Stress Test (TSST), a stress-inducing public speaking task, then cultured for 6 hours in the presence and absence of the inflammatory endotoxin lipopolysaccharide (LPS). Effects of TSST and LPS on 41 immune biomarkers were examined using repeated-measures analysis of variance. Separately, juvenile (8-week-old) male mice were non-stressed or exposed to reminder social defeat then intraperitoneally injected with saline or LPS (n = 6/group). Brains were perfused and collected for immunohistochemistry and confocal microscopy at 0, 1, 6, and 24 hours post-injection. Activity was determined by the density of cFos-positive neurons in the paraventricular hypothalamus, paraventricular thalamus, and basolateral amygdala, regions known to show sustained activation to immunological challenge. Analyses in the adolescent study indicated a strong effect of LPS but no effects of TSST or TSST×LPS interaction on immune biomarkers. Similarly, reminder social defeat did not induce sustained neuronal activity changes comparable to LPS immunological challenge in juvenile mice. Our convergent findings across species suggest that the acute immune response to stress documented in adults is not present in youth. Thus, aging and chronicity effects may play an important role in the inflammatory response to acute psychosocial stress.
PubMed: 38405791
DOI: 10.21203/rs.3.rs-3845793/v1 -
Journal of Clinical Medicine Feb 2024Mixed martial arts (MMA) fighters use their arms and hands for striking with the fists, grappling, and defensive techniques, which puts a high load on the forearms and...
Mixed martial arts (MMA) fighters use their arms and hands for striking with the fists, grappling, and defensive techniques, which puts a high load on the forearms and hand muscles. New methods are needed to decrease the risk of injury and increase the effectiveness of regeneration. This study aimed to assess the effectiveness of cryo-compression (CC) therapy of different times (3 and 6 min) on forearm muscles in MMA fighters by investigating muscle pain, stiffness, tension, elasticity strength, and perfusion. Twenty professional male MMA fighters aged 26.5 ± 4.5 years, with training experience of 10.3 ± 5.0 years, were enrolled on an experimental within-group study design. The participants underwent CC therapy at a temperature of 3 °C and compression of 75 mmHg for 3 min and, in the second session, for 6 min. The investigated parameters were in the following order: (1) perfusion in non-reference units (PU), (2) muscle tone (T-[Hz]), (3) stiffness (S-[N/m]), (4) elasticity (E-[arb]), (5) pressure pain threshold (PPT-[N/cm]), and (6) maximum isometric force (Fmax [kgf]) at two time points: (1) at rest-2 min before CC therapy (pre) and (2) 2 min after CC therapy (post). There were significant differences between 3 and 6 min of CC therapy for PU and T. Meanwhile, F, E, PPT, and S were significantly different when comparing pre- to post-conditions. These results provide evidence that CC therapy is a stimulus that significantly affects parameters characterizing muscle biomechanical properties, pain threshold, strength, and tissue perfusion.
PubMed: 38398489
DOI: 10.3390/jcm13041177 -
PloS One 2024In this study we used a spatial transcriptomics approach to identify genes specifically associated with either high or low outflow regions in the trabecular meshwork...
In this study we used a spatial transcriptomics approach to identify genes specifically associated with either high or low outflow regions in the trabecular meshwork (TM) that could potentially affect aqueous humor outflow in vivo. High and low outflow regions were identified and isolated from organ cultured human anterior segments perfused with fluorescently-labeled 200 nm FluoSpheres. The NanoString GeoMx Digital Spatial Profiler (DSP) platform was then used to identified genes in the paraffin embedded tissue sections from within those regions. These transcriptome analyses revealed that 16 genes were statistically upregulated in high outflow regions and 57 genes were statistically downregulated in high outflow regions when compared to low outflow regions. Gene ontology enrichment analysis indicated that the top three biological categories of these differentially expressed genes were ECM/cell adhesion, signal transduction, and transcription. The ECM/cell adhesion genes that showed the largest differential expression (Log2FC ±1.5) were ADAM15, BGN, LDB3, and CRKL. ADAM15, which is a metalloproteinase that can bind integrins, was upregulated in high outflow regions, while the proteoglycan BGN and two genes associated with integrin signaling (LDB3, and CRKL) were downregulated. Immunolabeling studies supported the differential expression of ADAM15 and showed that it was specifically upregulated in high outflow regions along the inner wall of Schlemm's canal and in the juxtacanalicular (JCT) region of the TM. In addition to these genes, the studies showed that genes for decorin, a small leucine-rich proteoglycan, and the α8 integrin subunit were enriched in high outflow regions. These studies identify several novel genes that could be involved in segmental outflow, thus demonstrating that digital spatial profiling could be a useful approach for understanding segmental flow through the TM. Furthermore, this study suggests that changes in the expression of genes involved in regulating the activity and/or organization of the ECM and integrins in the TM are likely to be key players in segmental outflow.
Topics: Humans; Trabecular Meshwork; Aqueous Humor; Sclera; Proteoglycans; Integrins; Intraocular Pressure; Membrane Proteins; ADAM Proteins
PubMed: 38394161
DOI: 10.1371/journal.pone.0298802 -
Bioengineering (Basel, Switzerland) Jan 2024Distal outflow bleb-forming procedures in ophthalmic surgery expose subconjunctival tissue to inflammatory cytokines present in the aqueous humor, resulting in impaired...
Distal outflow bleb-forming procedures in ophthalmic surgery expose subconjunctival tissue to inflammatory cytokines present in the aqueous humor, resulting in impaired outflow and, consequently, increased intraocular pressure. Clinically, this manifests as an increased risk of surgical failure often necessitating revision. This study (1) introduces a novel high-throughput screening platform for testing potential anti-fibrotic compounds and (2) assesses the clinical viability of modulating the transforming growth factor beta-SMAD2/3 pathway as a key contributor to post-operative outflow reduction, using the signal transduction inhibitor verteporfin. Human Tenon's capsule fibroblasts (HTCFs) were cultured within a 3D collagen matrix in a microfluidic system modelling aqueous humor drainage. The perfusate was augmented with transforming growth factor beta 1 (TGFβ1), and afferent pressure to the tissue-mimetic was continuously monitored to detect treatment-related pressure elevations. Co-treatment with verteporfin was employed to evaluate its capacity to counteract TGFβ1 induced pressure changes. Immunofluorescent studies were conducted on the tissue-mimetic to corroborate the pressure data with cellular changes. Introduction of TGFβ1 induced treatment-related afferent pressure increase in the tissue-mimetic. HTCFs treated with TGFβ1 displayed visibly enlarged cytoskeletons and stress fiber formation, consistent with myofibroblast transformation. Importantly, verteporfin effectively mitigated these changes, reducing both afferent pressure increases and cytoskeletal alterations. In summary, this study models the pathological filtration bleb response to TGFβ1, while demonstrating verteporfin's effectiveness in ameliorating both functional and cellular changes caused by TGFβ1. These demonstrate modulation of the aforementioned pathway as a potential avenue for addressing post-operative changes and reductions in filtration bleb outflow capacity. Furthermore, the establishment of a high-throughput screening platform offers a valuable pre-animal testing tool for investigating potential compounds to facilitate surgical wound healing.
PubMed: 38391628
DOI: 10.3390/bioengineering11020142