Injury Model Research Articles

Effectiveness of Insolubilized Poly(vinyl alcohol)-Based Electrospun Fiber-Loaded Methylprednisolone by Enzyme-Catalyzed Cross-Linking in a Rat Spinal Cord Injury Model.

Spinal cord injury (SCI) has been implicated in neural loss and, consequently, motor/sensory impairment. Here, we propose an improved formation for fibrous mat fabrication from the derivatives of poly(vinyl alcohol) (PVA) and gelatin (Gela) through horseradish peroxidase-mediated cross-linking, providing a sustained release of methylprednisolone (MP) for SCI repair. After 28 days, the animals were evaluated in terms of remyelination and apoptosis and underwent behavioral tests. The mechanical properties, hydrophobicity, and degradation rate of PVAPh/GelaPh fibrous mats were significantly improved as compared with those of PVAPh samples. This could provide the desired structure for a sustained MP release. The seeded cells could adhere and proliferate to the composite fibers, which indicates the cytocompatibility of the resultant PVAPh/GelaPh fibrous mat. The results showed significant reductions in the number of apoptotic neurons and a substantial improvement in remyelination in the SCI+ PVAPh/GelaPh + MP group. The behavioral tests confirmed improvement in locomotor hindlimb function following treatment. The MP-loaded PVAPh/GelaPh mat developed through the long-term release of MP and the biocompatible fabricated mat could inhibit axonal demyelination, attenuate apoptosis, and improve the functional outcome, which verified the potential of PVAPh/GelaPh + MP nanocomposites as a bioactive scaffold for SCI regeneration.

Effects of cannabidiol on AMPKα2 /HIF-1α/BNIP3/NIX signaling pathway in skeletal muscle injury

Cannabidiol: (CBD) is a non-psychoactive natural active ingredient from cannabis plant, which has many pharmacological effects, including neuroprotection, antiemetic, anti-inflammatory and anti-skeletal muscle injury. However, the mechanism of its effect on skeletal muscle injury still needs further research. In order to seek a scientifically effective way to combat skeletal muscle injury during exercise, we used healthy SD rats to establish an exercise-induced skeletal muscle injury model by treadmill training, and systematically investigated the effects and mechanisms of CBD, a natural compound in the traditional Chinese medicine Cannabis sativa L., on combating skeletal muscle injury during exercise. CBD effectively improved the fracture of skeletal muscle tissue and reduced the degree of inflammatory cell infiltration. Biochemical indexes such as CK, T, Cor, LDH, SOD, MDA, and GSH-Px in serum of rats returned to normal. Combining transcriptome and network analysis results, CBD may play a protective role in exercise-induced skeletal muscle injury through HIF-1 signaling pathway. The experimental results implied that CBD could down-regulate the expression of IL-6, NF-κB, TNF-α, Keap1, AMPKα2, HIF-1α, BNIP3 and NIX, and raised the protein expression of IL-10, Nrf2 and HO-1. These results indicate that the protective effect of CBD on exercise-induced skeletal muscle injury may be related to the inhibition of oxidative stress and inflammation, thus inhibiting skeletal muscle injury through AMPKα2/HIF-1α/BNIP3/NIX signal pathways.

Magnetic resonance imaging tracing of superparamagnetic iron oxide nanoparticle-labeled mesenchymal stromal cells for repairing spinal cord injury.

Mesenchymal stromal cell transplantation Is an effective and promising approach for treating various systemic and diffuse diseases. However, the biological characteristics of transplanted mesenchymal stromal cells in humans remain unclear, including cell viability, distribution, migration, and fate. Conventional cell tracing methods cannot be used in the clinic. The use of superparamagnetic iron oxide nanoparticles as contrast agents allows for the observation of transplanted cells using magnetic resonance imaging. In 2016, the National Medical Products Administration of China approved a new superparamagnetic iron oxide nanoparticle, Ruicun, for use as a contrast agent in clinical trials.In the present study, an acute hemi-transection spinal cord injury model was established in beagle dogs. The injury was then treated by transplantation of Ruicun-labeled mesenchymal stromal cells. The results indicated that Ruicun-labeled mesenchymal stromal cells repaired damaged spinal cord fibers and partially restored neurological function in animals with acute spinal cord injury. T2*-weighted imaging revealed low signal areas on both sides of the injured spinal cord. The results of quantitative susceptibility mapping with ultrashort echo time sequences indicated that Ruicun-labeled mesenchymal stromal cells persisted stably within the injured spinal cord for over 4 weeks. These findings suggest that magnetic resonance imaging has the potential to effectively track the migration of Ruicun-labeled mesenchymal stromal cells and assess their ability to repair spinal cord injury.

Testing Neuroprotective Strategies in Prolonged Field Care Model of Traumatic Brain Injury and Hemorrhagic Shock.

Prolonged Field Care (PFC) is a military adaptation of Tactical Combat Casualty Care providing extended pre-hospital management during delayed extrication. Effects of addition of Valproic Acid (VPA) to Fresh Frozen Plasma (FFP) in a PFC model of hemorrhagic shock and traumatic brain injury (TBI) are not known. We hypothesized that VPA is associated with decreased neurological impairment, and its protective changes are detected at the transcriptomic level. Swine underwent TBI and 40%-blood volume hemorrhage. After 2-hours of shock, they were randomized to: 1)normal saline (NS), 2)NS+250 ml FFP (NS+FFP), or 3)NS+FFP+150 mg/kgVPA(NS+FFP+VPA). At 72-hours, they were transfused packed red blood cells before being euthanized. Intraoperative variables and neurological outcomes were compared. Brain lesion size was measured, and gene expression profiles were analyzed using RNA-sequencing. Pathway and network analyses were performed on differentially expressed genes. Real-time PCR was performed to validate key genes. NS+FFP and NS+FFP+VPA required significantly less crystalloid resuscitation(974 ml-NS+FFP; 1461ml-NS+FFP+VPA vs. 4540 ml-NS, p<0.001), had smaller brain lesion size(2477mm3-NS+FFP; 3018.0mm3-NS+FFP+VPA vs. 4517.0mm3-NS, p<0.01), and less functional neurologic impairment compared to NS. Per pathway analysis of differentially expressed genes, VPA was associated with enrichment of numerous metabolic changes in injured brains, which were not observed with FFP. Network analysis showed enrichment of various gene networks. MT-ATP8 gene was downregulated in VPA-treated animals. The addition of FFP to the resuscitation protocol resulted in a significant reduction in crystalloid requirements. Both, the FFP and FFP+VPA groups showed improved neurological recovery compared to NS alone and had distinctive transcriptomic profiles in injured brains at 72-hours. MT-ATP8, involved in worsening ischemia following brain injury, was down-regulated in VPA-treated animals.

The Gelatin-Chitosan-Tetraethyl Orthosilicate Calcium Hydroxide Composite as a Potential Dental Pulp Medicament (Study on Expression of COX-2, PGP 9.5, TNF-α and Neutrophils number)

Introduction Calcium hydroxide (Ca(OH)2) is the material of choice for pulp therapy. However, Ca(OH)2 has drawbacks such as toxicity, poor sealing, and tunnel defect formation. Alternative materials have been developed to provide more biocompatible materials with better dentin formation ability. The objective of this study was to evaluate the effect of composites containing gelatin (G), chitosan (CH), tetraethyl orthosilicate (TEOS), and Ca(OH)2, namely G-CH-TEOS-Ca(OH)2 (Extended data) on inflammation of the dental pulp (expression of COX-2, PGP 9.5, TNF-α, and neutrophil number). Materials and methods A total of 16 Wistar rat models of acute pulp injury were prepared and divided into two groups, treatment and control, 8 with each. In the treatment group, we applied a pulp-capping material using G-CH-TEOS-Ca(OH)2 and Ca(OH)2. On the 1st and 3rd days, rats were sacrificed. Tissue samples from 4 rats in each group were processed for histological preparation. COX-2, PGP 9.5, and TNF-α were observed using immunohistochemical (IHC) staining, and neutrophil numbers were observed using hematoxylin-eosin staining. Image analysis of COX-2, PGP 9.5, and TNF-α expression was performed using ImageJ software. Results The results showed a decrease in COX-2 expression, but not significantly while PGP 9.5 and TNF-α expression were significantly higher than those in the control group. Neutrophil numbers were lower in the treatment group than in the control group, but the difference was not statistically significant. Conclusion The G-CH-TEOS-Ca(OH)2 composite material may have potential as an exposed pulp medicament by reducing inflammation (COX-2 expression and number of neutrophils) and increasing the regeneration factor (TNF-α expression) and nerve (PGP 9.5 expression).

Microglia ameliorate delirium-like phenotypes in a murine model of acute ventilator-induced lung injury

BackgroundDelirium affects 50–85% of patients on mechanical ventilation and is associated with increased mortality, prolonged hospitalization, and a three-fold higher risk of dementia. Microglia, the resident immune cells of the brain, exhibit both neuroprotective and neurotoxic functions; however, their effects in mechanical ventilation-induced acute lung injury (VILI) are unknown. We hypothesize that in a model of short-term VILI, microglia play a neuroprotective role to ameliorate delirium-like phenotypes.MethodsMicroglia depletion (n = 18) was accomplished using an orally administered colony stimulating factor 1 receptor inhibitor, while controls received a vehicle diet (n = 18). We then compared extent of neuronal injury in the frontal cortex and hippocampus using cleaved caspase-3 (CC3) and multiple delirium-like behaviors in microglia depleted and non-microglia depleted male mice (C57BL/6 J aged 4–9 months) following VILI. Delirium-like behaviors were evaluated using the Open Field, Elevated Plus Maze, and Y-maze assays. We subsequently evaluated whether repopulation of microglia (n = 14 repopulation, 14 vehicle) restored the phenotypes.ResultsFrontal/hippocampal neuronal CC3 levels were significantly higher in microglia depleted VILI mice compared to vehicle-treated VILI controls (p < 0.01, p < 0.01, respectively). These structural changes were accompanied by worse delirium-like behaviors in microglia depleted VILI mice compared to vehicle controls. Specifically, microglia depleted VILI mice demonstrated: (1) significantly increased time in the periphery of the Open Field (p = 0.01), (2) significantly increased coefficient of variation (p = 0.02), (3) trend towards reduced time in the open arms of the Elevated Plus Maze (p = 0.09), and (4) significantly decreased spontaneous alternations on Y-maze (p < 0.01). There was a significant inverse correlation between frontal CC3 and percent spontaneous alternations (R2 = 0.51, p < 0.01). Microglia repopulation showed a near-complete return to vehicle levels of delirium like-behaviors.ConclusionsThis study demonstrates that microglia depletion exacerbates structural and functional delirium-like phenotypes after VILI, while subsequent repopulation of microglia restores these phenotypes. These findings suggest a neuroprotective role for microglia in ameliorating neuronal and functional delirium-like phenotypes and call for consideration of interventions that leverage endogenous microglia physiology to mitigate delirium.

Tetramethylpyrazine Analogue T-006 Protects Neuronal and Endothelial Cells Against Oxidative Stress via PI3K/AKT/mTOR and Nrf2 Signaling.

T-006, a novel neuroprotective derivative of tetramethylpyrazine (TMP), exhibits multifunctional neuroprotective properties. T-006 has been shown to improve neurological and behavioral functions in animal models of ischemic stroke and neurodegenerative diseases. The present study aims to further elucidate the mechanisms underlying the protective effects of T-006 against oxidative injuries induced by glutamate or hypoxia. Mouse hippocampal HT22 cells were used to evaluate the neuroprotective effects of T-006 against glutamate-induced injuries, while mouse brain endothelial bEnd.3 cells were used to evaluate the cerebrovascular protective effects of T-006 against oxygen-glucose deprivation followed by reperfusion (OGD/R)-induced injuries. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and flow cytometry were used to measure cell viability and oxidative stress. Western blot and immunofluorescence analyses of protein expression were used to study cell signaling pathways. T-006 exhibited significant protective effects in both oxidative injury models. In HT22 cells, T-006 reduced cell death and enhanced antioxidant capacity by upregulating mTOR and nuclear factor erythroid 2-related factor 2/Heme oxygenase-1 (Nrf2/HO-1) signaling. Similarly, in bEnd.3 cells, T-006 reduced oxidative injuries and preserved tight junction integrity through Nrf2/HO-1 upregulation. These effects were inhibited by LY294002, a Phosphoinositide 3-kinase (PI3K) inhibitor. T-006 may exert its neuroprotective and cerebrovascular protective effects via the regulation of PI3K/AKT-mediated pathways, which facilitate downstream mTOR and Nrf2 signaling, leading to improved cell survival and antioxidant defenses.

Evaluation of Blood–Brain Barrier Disruption Using Low- and High-Molecular-Weight Complexes in a Single Brain Sample in a Rat Traumatic Brain Injury Model: Comparison to an Established Magnetic Resonance Imaging Technique

Traumatic brain injury (TBI), a major cause of death and disability among young people, leads to significant public health and economic challenges. Despite its frequency, treatment options remain largely unsuitable. However, examination of the blood–brain barrier (BBB) can assist with understanding the mechanisms and dynamics of brain dysfunction, which affects TBI sufferers secondarily to the injury. Here, we present a rat model of TBI focused on two standard BBB assessment markers, high- and low-molecular-weight complexes, in order to understand BBB disruption. In addition, we tested a new technique to evaluate BBB disruption on a single brain set, comparing the new technique with neuroimaging. A total of 100 Sprague–Dawley rats were separated into the following five groups: naive rats (n = 20 rats), control rats with administration (n = 20 rats), and TBI rats (n = 60 rats). Rats were assessed at different time points after the injury to measure BBB disruption using low- and high-molecular-weight complexes. Neurological severity score was evaluated at baseline and at 24 h following TBI. During the neurological exam after TBI, the rats were scanned with magnetic resonance imaging and euthanized for assessment of the BBB permeability. We found that the two markers displayed different examples of BBB disruption in the same set of brain tissues over the period of a week. Our innovative protocol for assessing BBB permeability using high- and low-molecular-weight complexes markers in a single brain set showed appropriate results. Additionally, we determined the lower limit of sensitivity, therefore demonstrating the accuracy of this method.

Impaired pericyte-Müller glia interaction via PDGFRβ suppression aggravates photoreceptor loss in a rodent model of light-induced retinal injury.

To investigate the involvement of pericyte-Müller glia interaction in retinal damage repair and assess the influence of suppressing the platelet-derived growth factor receptor β (PDGFRβ) signaling pathway in retinal pericytes on photoreceptor loss and Müller glial response. Sprague-Dawley rats were exposed to intense light to induce retinal injury. Neutralizing antibody against PDGFRβ were deployed to block the signaling pathway in retinal pericytes through intravitreal injection. Retinal histology and Müller glial reaction were assessed following light injury. In vitro, normal and PDGFRβ-blocked retinal pericytes were cocultured with Müller cell line (rMC-1) to examine morphological and protein expression changes upon supplementation with light-injured supernatants of homogenized retinas (SHRs). PDGFRβ blockage 24h prior to intense light exposure resulted in a significant exacerbation of photoreceptor loss. The upregulation of GFAP and p-STAT3, observed after intense light exposure, was significantly inhibited in the PDGFRβ blockage group. Further upregulation of cytokines monocyte chemoattractant protein 1 (MCP-1) and interleukin-1β (IL-1β) was also observed following PDGFRβ inhibition. In the in vitro coculture system, the addition of light-injured SHRs induced pericyte deformation and upregulation of proliferating cell nuclear antigen (PCNA) expression, while Müller cells exhibited neuron-like morphology and expressed Nestin. However, PDGFRβ blockage in retinal pericytes abolished these cellular responses to light-induced damage, consistent with the in vivo PDGFRβ blockage findings. Pericyte-Müller glia interaction plays a potential role in the endogenous repair process of retinal injury. Impairment of this interaction exacerbates photoreceptor degeneration in light-induced retinal injury.

Evaluation of Blood-Brain Barrier Breakdown in a Mouse Model of Mild Traumatic Brain Injury

Evaluation of Blood-Brain Barrier Breakdown in a Mouse Model of Mild Traumatic Brain Injury

Directly monitoring the dynamic in vivo metabolisms of hyperpolarized 13C-oligopeptides.

Peptides play essential roles in biological phenomena, and, thus, there is a growing interest in detecting in vivo dynamics of peptide metabolisms. Dissolution-dynamic nuclear polarization (d-DNP) is a state-of-the-art technology that can markedly enhance the sensitivity of nuclear magnetic resonance (NMR), providing metabolic and physiological information in vivo. However, the hyperpolarized state exponentially decays back to the thermal equilibrium, depending on the spin-lattice relaxation time (T1). Because of the limitation in T1, peptide-based DNP NMR molecular probes applicable in vivo have been limited to amino acids or dipeptides. Here, we report the direct detection of in vivo metabolic conversions of hyperpolarized 13C-oligopeptides. Structure-based T1 relaxation analysis suggests that the C-terminal [1-13C]Gly-d2 residue affords sufficient T1 for biological uses, even in relatively large oligopeptides, and allowed us to develop 13C-β-casomorphin-5 and 13C-glutathione. It was found that the metabolic response and perfusion of the hyperpolarized 13C-glutathione in the mouse kidney were significantly altered in a model of cisplatin-induced acute kidney injury.

Acute kidney disease in mice is associated with early cardiovascular dysfunction

Acute kidney injury (AKI) and chronic kidney disease (CKD) are major health concerns due to their increasing incidence and high mortality. They are interconnected syndromes; AKI without recovery evolves into acute kidney disease (AKD), which can indicate an AKI-to-CKD transition. Both AKI and CKD are associated with a risk of long-term cardiovascular complications, but whether vascular and cardiac dysfunctions can occur as early as the AKD period has not been studied extensively. In a mouse model of kidney injury (KI) with non-recovery, we performed vasoreactivity and echocardiography analyses on days 15 (D15) and 45 (D45) after KI. We determined the concentrations of two major gut-derived protein-bound uremic toxins known to induce cardiovascular toxicity—indoxyl sulfate (IS) and para-cresyl sulfate (PCS)—and the levels of inflammation and contraction markers on D7, D15, and D45. Mice with KI showed acute tubular and interstitial kidney lesions on D7 and D15 and chronic glomerulosclerosis on D45. They showed significant impairment of aorta relaxation and systolic-diastolic heart function, both on D15 and D45. Such dysfunction was associated with downregulation of the expression of two contractile proteins, αSMA and SERCA2a, with a more pronounced effect on D15 than on D45. KI was also followed by a rapid increase in IS and PCS serum concentrations and the expression induction of pro-inflammatory cytokines and endothelial adhesion molecules in serum and cardiovascular tissues. Therefore, these results highlight that AKD leads to early cardiac and vascular dysfunctions. How these dysfunctions could be managed to prevent cardiovascular events deserves further study.

Osteopontin Promotes Cholangiocyte Secretion of Chemokines to Support Macrophage Recruitment and Fibrosis in MASH.

Osteopontin (OPN) promotes the ductular reaction and is a major driver of chronic liver disease (CLD) progression. Although CLD is characterised by the accumulation of inflammatory cells including macrophages around the peri-portal regions, the influence of OPN on recruitment is unclear. We investigated the role of OPN in cholangiocyte chemokine production and macrophage recruitment by combining invivo, invitro, and in silico approaches. The effects of OPN on cholangiocyte chemokine production and macrophage migration were assessed in culture, alongside RNA-sequencing to identify genes and pathways affected by OPN depletion. Murine liver injury models were used to assess liver chemokine expression and liver macrophage/monocyte recruitment. OPN and chemokine expression were analysed in liver tissue and plasma from biopsy-proven metabolic dysfunction-associated alcoholic steatohepatitis (MASH) patients. OPN-knockdown in cholangiocytes reduced chemokine secretion. RNA-sequencing showed OPN-related effects clustered around immunity, chemotaxis and chemokine production. Macrophage exposure to cholangiocyte-conditioned media showed OPN-supported migration via chemokines chemokine (C-C motif) ligand (CCL)2, CCL5 and chemokine (C-X-C motif) ligand (CXCL)1. These effects were related to NF-κB signalling. Murine liver fibrosis was accompanied by upregulated liver OPN, CCL2, CCL5 and CXCL1 mRNA, and accumulation of liver cluster of differentiation (CD)11b/F4/80+CC chemokine receptors (CCR2)high macrophages but treatment with OPN-specific neutralising aptamers reduced fibrosis, chemokine mRNAs and accumulation of liver CD11b/F4/80+CCR2high/lymphocyte antigen 6 complexhigh inflammatory monocytes. In human MASH, liver OPN correlated with chemokines CCL2 and IL8 in association with portal injury and fibrosis. Plasma OPN, serum CCL2 and IL8 also increased with fibrosis stage. OPN promotes cholangiocyte chemokine secretion and the accumulation of pro-inflammatory monocytes. These data support neutralisation of OPN as an anti-inflammatory and anti-fibrotic strategy.

Bidirectional Impact of Varying Severity of Acute Kidney Injury on Calcium Oxalate Stone Formation.

Acute Kidney Injury (AKI) is a prevalent renal disorder. The occurrence of AKI may promote the formation of renal calcium oxalate stones by exerting continuous effects on renal tubular epithelial cells. We aimed to delineate the molecular interplay between AKI and nephrolithiasis. A mild (20 min) and severe (30 min) renal ischemia-reperfusion injury model was established in mice. Seven days after injury, calcium oxalate stones were induced using glyoxylate (Gly) to evaluate the impact of AKI on the formation of kidney stones. Transcriptome sequencing was performed on tubular epithelial cells (TECs) to elucidate the relationship between AKI severity and kidney stones. Key transcription factors (TF) regulating differential gene transcription levels were identified using motif analysis, and pioglitazone, ginkgetin, and fludarabine were used for targeted therapy to validate key transcription factors as potential targets for kidney stone treatment. Severe AKI led to increased deposition of calcium oxalate crystals in renal, impaired kidney function, and upregulation of kidney stone-related gene expression. In contrast, mild AKI was associated with decreased crystal deposition, preserved kidney function, and downregulation of similar gene expression. Transcriptomic analysis revealed that genes associated with inflammation and cell adhesion pathways were significantly upregulated after severe AKI, while genes related to energy metabolism pathways were significantly upregulated after mild AKI. An integrative bioinformatic analysis uncovered a TF regulatory network within TECs, pinpointing that PKNOX1 was involved in the upregulation of inflammation-related genes after severe AKI, and inhibiting PKNOX1 function with Pioglitazone could simultaneously reduce the increase of calcium oxalate crystals after severe AKI in kidney. On the other hand, motif analysis also revealed the protective role of STAT1 in the kidneys after mild AKI, enhancing the function of STAT1 with Ginkgetin could reduce kidney stone formation, while the specific inhibitor of STAT1, Fludarabine, could eliminate the therapeutic effects of mild AKI on kidney stones. Inadequate repair of tubular epithelial cells after severe AKI increases the risk of kidney stone formation, with the upregulation of inflammation-related genes regulated by PKNOX1 playing a role in this process. Inhibiting PKNOX1 function can reduce kidney stone formation. Conversely, after mild AKI, effective cell repair through upregulation of STAT1 expression can protect TEC function, reduce stone formation, and activating STAT1 function can also achieve the goal of treating kidney stones.

Toll-Like Receptor 4 (TLR4) Promotes DRG Regeneration and Repair after Sciatic Nerve Injury via the ERK-NF-kB Pathway.

Previously, we found that the expression of Toll-like receptor 4 (TLR4) is altered after sciatic nerve injury, and its differential expression plays a key role in recovery. However, the mechanisms by which TLR4 affects neuronal function in the dorsal root ganglion (DRG) have not been completely evaluated. The objective is to determine TLR4 expression in DRG tissues after sciatic neural injury and exploring the effects of TLR4 knockdown and overexpression in the DRG on neuronal function and nerve regeneration in rats in vivo and in vitro. We established a model of nerve injury and utilized molecular biology and cell biology experiments to explore the molecular mechanisms by which TLR4 in the DRG affects sciatic nerve restoration and regeneration after injury. Verified the localization of TLR4 in DRG neurons. Investigated pathways that related to apoptosis or nerve regeneration by which TLR4 regulates the function of DRG neurons. TLR4 expression was upregulated in the DRG tissues of rats after sciatic nerve injury. TLR4 overexpression promoted axon regeneration and inhibited apoptosis in DRG neurons. TLR4 promoted the regeneration of axons and the recovery of motor and sensory functions in the sciatic nerve after injury in vivo, and the data showed that TLR4 may regulate the function of DRG neurons and promote nerve repair and regeneration through the ERK and NF-κB signaling pathways in vivo and ex vivo. The study suggests that TLR4 may regulate the function of DRG neurons and promote nerve regeneration by affecting the ERK and NF-κB signaling pathways.

The Occurrence of Senescence in the Arteriovenous Fistula in the Rat.

Maturational failure of dialysis arteriovenous fistulas (AVFs) not uncommonly occurs and is of considerable and timely importance. Our prior studies demonstrate that senescence, a phenotypic process that promotes vascular and other diseases, occurs in the murine AVF. In the present study, we examined whether senescence also occurs in the rat AVF model and the effect of compounds that inhibit or accelerate senescence. The rat AVF was created in the femoral vessels by an end vein-side artery anastomosis. We assessed in the AVF the expression of critical drivers of senescence, specifically, the cell cycle inhibitors p16Ink4a and p21Cip1, and such indices of a senescence phenotype as senescence-associated β-galactosidase (SA-β-gal) activity, SA-β-gal staining, and a senescence-associated secretory phenotype (SASP). We examined the effects of compounds that retard or accelerate senescence on AVF blood flow. The AVF evinced upregulation of p16Ink4a and p21Cip1 when assessed 3 days after AVF creation. The AVF also demonstrated increased SA-β-gal activity in the artery and vein; staining for SA-β-gal in the AVF artery, anastomosis, and vein; and a prominent SASP. Fisetin, an established senolytic that is protective in other models of vascular injury, when administered for 3 weeks, increased AVF blood flow and outward remodeling. Hemin, when administered for 3 weeks, decreased AVF blood flow. We demonstrate that hemin is a novel inducer of a senescence phenotype in endothelial cells, as reflected by several senescence indices. However, when administered relatively acutely (for 5 days) hemin increased AVF blood flow via HO-dependent mechanisms, as the latter was entirely prevented by a competitive inhibitor of HO activity. The rat AVF exhibits senescence within 3 days of its creation. Chronic administration of a senolytic compound (fisetin) increases AVF blood flow, whereas chronic administration of a pro-senescence compound (hemin) decreases AVF blood flow.

Transcriptome-Optimized Hydrogel Design of a Stem Cell Niche for Enhanced Tendon Regeneration.

Bioactive hydrogels have emerged as promising artificial niches for enhancing stem cell-mediated tendon repair. However, a substantial knowledge gap remains regarding the optimal combination of niche features for targeted cellular responses, which often leads to lengthy development cycles and uncontrolled healing outcomes. To address this critical gap, an innovative, data-driven materiomics strategy is developed. This approach is based on in-house RNA-seq data that integrates bioinformatics and mathematical modeling, which is a significant departure from traditional trial-and-error methods. It aims to provide both mechanistic insights and quantitative assessments and predictions of the tenogenic effects of adipose-derived stem cells induced by systematically modulated features of a tendon-mimetic hydrogel (TenoGel). The knowledge generated has enabled a rational approach for TenoGel design, addressing key considerations, such as tendon extracellular matrix concentration, uniaxial tensile loading, and in vitro pre-conditioning duration. Remarkably, our optimized TenoGel demonstrated robust tenogenesis in vitro and facilitated tendon regeneration while preventing undesired ectopic ossification in a rat tendon injury model. These findings shed light on the importance of tailoring hydrogel features for efficient tendon repair. They also highlight the tremendous potential of the innovative materiomics strategy as a powerful predictive and assessment tool in biomaterial development for regenerative medicine.

Citrus extraction provides neuroprotective effect in optic nerve crush injury mice through P53 signaling pathway

Optic nerve injury induces the loss of retinal ganglion cells and optic nerve atrophy. Our previous study demonstrated the neuroprotective effects of Citrus in the optic nerve crush (ONC) injury model. To explore the underlying neuroprotective mechanism, we focus on the changes in gene expression profile pre- and post-Citrus treatment in the ONC injury model. The results suggested that the Citrus regulated the chemokine signaling pathway and P53 signaling pathway, etc. Four genes were identified as potential target genes (Melk, Mki67, Ccnb1, and Cenpf). Subsequent qRT-PCR and western blot confirmed that Citrus regulated cell apoptosis and inflammation-related gene expression, and inhibited P53 activation-induced cell death. The present study demonstrated that the neuroprotective effect of Citrus was achieved through the regulation of the P53/BCL-2/BAX pathway and the expression regulation of four critical genes, which provide novel therapeutic targets for the therapy of optic nerve injury.

Trans-chalcone ameliorates CCl4-induced acute liver injury by suppressing endoplasmic reticulum stress, oxidative stress and inflammation

BackgroundAcute liver injury serves as a crucial marker for detecting liver damage due to toxic, viral, metabolic, and autoimmune exposures. Due to the response to adverse external stimuli and various cellular homeostasis, Endoplasmic reticulum stress (ERS), Oxidative stress, and Inflammation have great potential for treating liver injury. Trans-chalcones (TC) is a polyphenolic compound derived from a natural plant with anti-oxidative and anti-inflammatory abilities. Here, TC was aimed to attenuate liver injury by triggering ER stress, oxidative stress, inflammation, and apoptosis. A single dose of carbon tetrachloride (CCl4) 1 mL/kg was administered intraperitoneally into C57BL6 mice to construct an in vivo NAFLD model, whereas AML12 cells were treated with lipopolysaccharides (LPS) to construct an in vitro NAFLD model. The mice used in the experiment were randomly assigned to two groups: a 12-hour set and a 24-hour set. Forty-nine mice were randomly divided into seven groups, the control group (Group I), TC group (Group II) 10 mg/kg TC, negative control group (Group III) CCl4, TC + CCl4 groups (Groups IV−VI), mice were subcutaneously treated with (5, 10, and 20) mg/kg of TC for three consecutive days before the CCl4 injection and the positive control group (Group VII) received 10 mg/kg Silymarin. After the experiment, serum transaminase, liver histological pathology, hepatic expression levels ERS, oxidative stress, and inflammation-related markers were assessed. TC pre-treatment significantly alleviates the expression of ER stress, oxidative stress, inflammatory cytokines, and apoptosis in both in vivo and in vitro models of liver injury. TC treatment significantly reduced serum transaminase levels (ALT and AST), and improved liver histopathological scores. TC administration also led to a reduction in MDA levels and the suppression of ROS generated by CCl4 in hepatic tissue, which contributed to an increase in GSH levels. The protective effect of TC on the liver injury mouse model was achieved by inhibiting hepatocyte apoptosis. Moreover, TC pre-treatment dramatically decreased the protein levels of ER stress indicators such as CHOP, Bip, Ero-Lα, IRE1α, PERK, Calnexin, and PDI when compared to the CCl4-only treated group. TC exerts hepatoprotective effects against CCl4-induced acute liver injuries in mice by modulating ERS, oxidative stress, and inflammation. These results suggest that TC pre-treatment at a dose of (20 mg/kg BW) was as effective as silymarin (10 mg/kg) in preventing CCl4-induced acute liver injury. Further investigations are necessary to elucidate the precise molecular mechanisms underlying the hepatoprotective effects of TC and to explore its therapeutic potential in clinical trials.

A microfluidic model of the first sensory synapse for analgesic target discovery.

The synaptic connections between dorsal root ganglia (DRG) and dorsal horn (DH) neurons are a crucial relay point for the transmission of painful stimuli. To delineate how synaptic plasticity may modulate the excitability of DH neurons, we have devised a microfluidic co-culture model that recapitulates the first sensory synapse using postnatal mouse sensory neurons. We show that DRG-DH co-cultures characterize salient features of the in vivo physiology of sensory neurons. Immunocytcochemical experiments of the cultured DH neurons show a co-localization of MAP2 with VGLUT2 and of MAP2 with Synapsin 1, corroborating the glutamatergic identity of the DH neurons and further suggesting the formation of active synapses in this neuronal set. Fluorometric imaging experiments demonstrate the elicitation of calcium responses in DH neurons following the stimulation of DRG cell bodies or axons. Selective NMDA and AMPA receptor blockade appreciably silences DH neuron responses, suggesting that glutamatergic signaling is maintained in vitro. Last, a surrogate model of peripheral nerve injury is introduced in the form of an axotomy, which results in elevated and prolonged calcium responses of DH neurons. Overall, the microfluidic mouse co-cultures provide a method advancement in the study of periphery-to-center pain signaling, where the potential of utilizing the platform for drug target identification is underscored.