Model Of Degeneration Research Articles

High-fat and high-sucrose diets induce an experimental rabbit model for age-related macular degeneration (AMD).

Age-related macular degeneration (AMD) is a leading cause of blindness. Metabolic disorders and diets are risk factors. We compared lipid profiles and retinal phenotypes with long-term feeding of four diets in male Chinchilla rabbits. Animals were fed with a normal diet (ND), high-fat (HFD), high-sucrose (HSD), or high-fat and high-sucrose diet (HFSD) for six months. The eyes were examined using multimodal imaging modalities and electroretinogram (ERG). Retinal sections were analyzed using H&E staining, toluidine blue staining, immunostaining, and transmission electron microscopy. Lipids and complement C3 in serum or aqueous humour were measured. RNA sequencing was performed to evaluate the retinal transcriptomes. HFD and HSD had minor effects on lipid profiles but synergistically induced severe dyslipidemia. All diets did not cause obesity. HFSD feeding induced retinal lesions like reticular pseudo-drusen (RPD) and pigmentary abnormalities. The RPD-like lesions were mainly lipid droplets around RPE cells. HFSD induced elevated ocular C3 levels and reduced retinal vessel density. In conclusion, HFD and HSD can synergistically induce normal-weight dyslipidemia and RPD-like retinal lesions. HFSD-fed male Chinchilla rabbits are a good model of early AMD.

Regenerative Outcomes of Combining siCOL1A2 Hydrogel with Acupuncture in a Rat Model of Chronic Intervertebral Disc Degeneration

Intervertebral disc degeneration (IVDD) is a significant cause of chronic pain and disability, necessitating innovative therapeutic strategies. This study investigates the combined effect of a novel siCOL1A2-encapsulated hydrogel and acupuncture on IVDD in a rat model. We developed a hydrogel system, siCOL1A2-encapsulated G5-PBA hydrogel (siCOL1A2@G5-PBA@Gel), designed for sustained siRNA delivery to the degenerated discs and assessed its therapeutic efficacy alongside acupuncture treatment. Key inflammatory genes were identified through RNA-seq analysis, with COL1A2 highlighted as a crucial regulator of inflammatory responses in IVDD. Our in vivo experiments involved treating rats with hydrogel alone, acupuncture alone, and combining both. The treatments were evaluated through behavioral pain assessments, imaging techniques (X-ray and MRI), and histological analyses. Results indicated that the combination therapy significantly alleviated pain, reduced inflammation, and promoted disc regeneration more effectively than individual treatments. The hydrogel proved biocompatible and facilitated targeted gene silencing, while acupuncture enhanced therapeutic outcomes by improving local blood circulation and modulating inflammatory responses. These findings suggest that integrating siCOL1A2 hydrogel with acupuncture offers a promising approach to treating IVDD.

Dynamics of clinical and immunological parameters in retinal pigment epithelium transplantation in the context of combined immunosuppressive therapy in the rabbit model of retinal degeneration

Progressive damage of the retinal pigment epithelium (RPE) underlies the pathogenesis of degenerative retinal diseases such as: age-related macular degeneration (AMD), Stargardt’s disease, retinitis pigmentosa, Best’s disease and others. This group of diseases led by AMD leads to irreversible loss of visual functions, blindness and disability. The possibilities of therapy of late stages of AMD are extremely limited. The most promising approach to replace the damaged retinal pigment epithelium appears to be transplantation of RPE cells derived from induced pluripotent stem cells (iPSC-RPE) into the subretinal space (SRS). Despite immune privilege in the SRS, transplantation of xenogeneic cellular material causes severe inflammation in the posterior segment of the eye and leads to graft rejection in an in vivo experiment in the absence of immunosuppression. The solution to the problem of tissue incompatibility in this case can be the use of combined immunosuppressive therapy (CIT), aimed, on the one hand, at suppression of local inflammation (in the eye) and, on the other hand, at suppression of the systemic transplantation immune response. The aim of the study: clinical and immunological analysis of the results of transplantation of IPSC-RPE suspension on the background of CIT, including single intravitreal intraoperative administration of kenalog and further systemic application of mycophenolate mofetil (MMF), in the model of RPE atrophy in rabbits. Standard and specialized ophthalmological examination was performed at early and distant terms after the intervention in order to clinically assess the posttransplantation process. To analyze the immune status, vitreous humor (VH) and blood serum (BS) of rabbits of the experimental groups were collected. The concentrations of TGF-β1, TGF-β2, and IL-2 were determined in the biomaterial using solid-phase enzyme immunoassay. According to the results of the study, subretinal transplantation of IPSC-RPE, performed against the background of combination of single intravitreal intraoperative administration of kenalog and systemic application of MMF, is a safe method, which provides preservation of the retina and other adjacent structures of the eye and allows to prevent rejection of xenogenic material during its transplantation both in a healthy eye and with pre-formed RPE atrophy, which opens perspectives for full testing of biological effects realized by IPSC-RPE.

Development of a microfluidic system with a bearing micropump for applying fluid shear force in an intervertebral disc degeneration model involving mechanical stimulation

Background: The intervertebral disc (IVD) functions as a shock absorber with viscoelastic tissues, including the gelatinous nucleus pulposus and the collagenous annulus fibrosus (AF). External mechanical loading influences IVD homeostasis but can cause damage and inflammatory reactions, contributing to disc degeneration. There is a lack of in vitro platforms for modeling IVD disease with mechanical stimulation.Methods: This study aimed to create a mechanical stimulation-based model by subjecting AF cells to shear stress using a micropump system. A micropump platform was used to measure pulsation, flow rate, and shear stress. AF cells were exposed to fluid shear stress of 0.5 and 1 dyne/cm², and morphological changes, cytotoxicity, and cell viability were analyzed through a live/dead assay. Perimeter, area, diameter, and elongation were assessed using live cell imaging and imaging analysis software.Results: The micropump platform exhibited optimal rotor characteristics for uniform pulsation and shear stress. Fluid shear stress at 0.5 dyne/cm² showed no significant difference from the control group, while 1 dyne/cm² reduced cell adhesion and survival. Comparing the 0.5 dyne/cm² shear stress group and interleukin-1β group to the control, significant decreases in perimeter, area, and diameter were observed.Conclusion: The study successfully developed a micro-peristaltic pump platform for applying fluid shear stress to cells, identifying an optimal rotor for uniform stress application. The platform effectively modeled IVD disease, revealing reduced cell adhesion and survival under specific shear stress conditions. This platform has potential promise for discovering biomarkers and exploring cellular responsiveness to external stimuli.

QM107, a novel CD148 (RTP Type J) activating peptide therapy for treating neovascular age-related macular degeneration.

Angiogenesis is a pathological component of neovascular age-related macular degeneration. Current therapies, although successful, are prone to high levels of patient non-response and a loss of efficacy over time, indicating the need to explore other therapeutic avenues. We have shown that an interaction between syndecan-2 and the tyrosine phosphatase receptor CD148 (RTP Type J) results in the ablation of angiogenesis. Here we exploit this pathway to develop a peptide activator of CD148 as a therapy for neovascular age-related macular degeneration. We tested a peptide (QM107) derived from syndecan-2 in a variety of angiogenesis models and a pre-clinical model of neovascular age-related macular degeneration. We assessed the toxicological and inflammatory profiles of QM107 and its stability in vitreous humour. QM107 inhibits angiogenesis in ex vivo sprouting assays and disrupts endothelial microcapillary formation via inhibition of cell migration. QM107 acts through CD148, leading to changes in GSK3A phosphorylation and β1 integrin activation. QM107 elicits a negligible inflammatory response and exhibits limited toxicity in cultured cells, and is stable in vitreous humour. Finally, we show proof of concept that QM107 blocks angiogenesis in vivo using a model of neovascular age-related macular degeneration. We have developed a CD148 activating peptide which shows promise in inhibiting angiogenesis in models of neovascular age-related macular degeneration. This treatment could either represent an alternative or augment existing therapies, and owing to its distinct mode of action be used in patients who do not respond to existing treatments.

Deprivation of visual input alters specific subset of inhibitory neurons and affect thalamic afferent terminals in V1 of rd1 mouse.

Retinitis Pigmentosa (RP) is a heterogenous group of inherited disorder, and its progression not only affects the retina but also the primary visual cortex. This manifests imbalances in the excitatory and inhibitory neurotransmission. Here, we investigated if changes in cortical functioning is linked to alterations in GABAergic population of neurons and its two important subsets, somatostatin (SST) and parvalbumin (PV) neuron in rd1 model of retinal degeneration (RD). We demonstrate marked decrease in the proportion of SST neurons in different layers of cortex whereas PV neurons were less affected. Moreover, we found reduced expression of glutamatergic thalamic afferents (VGLUT2) due to lack of visual activity. These results suggest PV neurons are likely recruited by the cortical circuitry to increase the inhibitory drive and compensate the disrupted inhibition-excitation balance. However, reduced SST expression perhaps results in weakening of stimulus selectivity. Delineating their functional role during RD will provide insights for acquisition of high-resolution vision thereby improving current state of vision restoration.

Host-Graft Synapses Form Functional Microstructures and Shape the Host Light Responses After Stem Cell-Derived Retinal Sheet Transplantation.

Retinitis pigmentosa represents a leading cause of blindness in developed countries, yet effective treatments for the disease remain unestablished. Previous studies have demonstrated the potential of stem cell-derived retinal organoid (SC-RO) sheet transplantation to form host-graft synapses and to improve light responsiveness in animal models of retinal degeneration. However, the detailed microstructures of these de novo synapses and their functional contribution have not been well elucidated. This study aims to (1) elucidate the microstructures of the host-graft synapse, and (2) investigate the overall distribution and contribution of these synapses to host retinal light responses. We identified host-graft synapses using a reporter system in mouse SC-RO and rd1 mice, a well-established model of end-stage retinal degeneration. Correlative array tomography was used to reveal the microstructure of host-graft synapses. Furthermore, we developed a semi-automated algorithm that robustly detects the host-graft photoreceptor synapses in the overall grafted area using the same reporter system in flat-mount retinas. We then integrated the spatial distribution of the host-graft synapses with light responses detected by multi-electrode array recording. Correlative array tomography revealed that host-graft synapses recapitulate the developmental process of photoreceptor synapse formation involving horizontal cells first and then rod bipolar cells. By integrating the spatial distribution of host-graft synapse and multi-electrode array recording, we showed that the number of light-responsive host retinal ganglion cells is positively correlated with the local density of host-graft synapses. De novo host-graft synapses recapitulate the developmental microstructure of the photoreceptor synapse, and their formation contributes to the light responsiveness after SC-RO transplantation.

Magnetic resonance imaging classification in a percutaneous needle injury rat model of intervertebral disc degeneration

BackgroundDuring the development of disease-modifying intervertebral disc degeneration (IDD) drugs, the rat model of IDD is frequently used for disease progression assessment. The aim of this study was to describe a magnetic resonance (MRI) scoring system for the assessment of different disc conditions in puncture-induced IDD, allowing standardization and comparison of results obtained by different investigators.MethodsA total of 36 Sprague-Dawley rats were utilized in the present study. The animals were divided into two groups: a sham group and an IDD group caused by puncture. The rats in the IDD group were subsequently divided into six categories based on time frames, with five rats in each category. The sham group was divided into two sub-groups (n = 3) for 28 and 56 days, respectively. T2-weighted images of rats consecutively studied with MRI of the coccygeal discs were classified according to the time course using the corresponding histological data. Additional scoring of the micro-CT was employed to identify the progression of bone destruction of the rat model of IDD.ResultsA comparison of the MRI results between the sham group and the IDD group revealed a significant reduction in NP height, area, T2WI value, and DHI in the latter group (P < 0.05). The micro-CT results demonstrated that following acupuncture, there was a notable decline in the BV, Tb.N, and height of the coccygeal vertebra, while the BS/BV and Tb.Sp exhibited a significant increase (P < 0.05). The histological results were analogous to the MRI results, indicating a progressive exacerbation of IDD and a corresponding increase in NP score (P < 0.05). The results of the MRI were found to be consistent with those of the micro-CT and histological analyses (P < 0.05). The results of the study demonstrate a robust correlation between MRI analysis and histological findings. Live animals are employed for MRI analysis to improve experiment comparability. The reliability of the MRI scoring system ensures assessment of disease progression in live animals, while promoting cost savings and animal welfare by avoiding the sacrifice of animals at different times.ConclusionsThe described scoring paradigm has quantitatively been found to differentiate IDD disease progression in an in vivo rat model. Hence, we suggest employing it to evaluate the rat IDD model and assess the effects of treatments in this model.

Mast cells promote choroidal neovascularization in a model of age-related macular degeneration

‘Wet’ age-related macular degeneration (AMD) is characterized by pathologic choroidal neovascularization (CNV) that destroys central vision. Abundant evidence points to inflammation and immune cell dysfunction in the progression of CNV in AMD. Mast cells are resident immune cells that control the inflammatory response. Mast cells accumulate and degranulate in the choroid of patients with AMD, suggesting they play a role in CNV. Activated mast cells secrete various biologically active mediators, including inflammatory cytokines and proteolytic enzymes such as tryptase. We investigated the role of mast cells in AMD using a model of CNV. Conditioned media from activated mast cells exerts proangiogenic effects on choroidal endothelial cells and choroidal explants. Laser-induced CNV in vivo was markedly attenuated in mice genetically depleted of mast cells (KitW−sh/W−sh) and in wild-type mice treated with mast cell stabilizer, ketotifen fumarate. Tryptase was found to elicit pronounced choroidal endothelial cell sprouting, migration and tubulogenesis; while tryptase inhibition diminished CNV. Transcriptomic analysis of laser-treated RPE/choroid complex revealed collagen catabolism and extracellular matrix (ECM) reorganization as significant events correlated in clusters of mast cell activation. Consistent with these analyses, compared to wildtype mice choroids of laser-treated mast cell-deficient mice displayed less ECM remodelling evaluated using collagen hybridizing peptide tissue binding. Findings herein provide strong support for mast cells as key players in the progression of pathologic choroidal angiogenesis and as potential therapeutic targets to prevent pathological neovascularization in ‘wet’ AMD.

Modulation of cannabinoid receptor 2 alters neuroinflammation and reduces formation of alpha-synuclein aggregates in a rat model of nigral synucleinopathy

Research into the disequilibrium of microglial phenotypes has become an area of intense focus in neurodegenerative disease as a potential mechanism that contributes to chronic neuroinflammation and neuronal loss in Parkinson’s disease (PD). There is growing evidence that neuroinflammation accompanies and may promote progression of alpha-synuclein (Asyn)-induced nigral dopaminergic (DA) degeneration. From a therapeutic perspective, development of immunomodulatory strategies that dampen overproduction of pro-inflammatory cytokines from chronically activated immune cells and induce a pro-phagocytic phenotype is expected to promote Asyn removal and protect vulnerable neurons. Cannabinoid receptor-2 (CB2) is highly expressed on activated microglia and peripheral immune cells, is upregulated in the substantia nigra of individuals with PD and in mouse models of nigral degeneration. Furthermore, modulation of CB2 protects against rotenone-induced nigral degeneration; however, CB2 has not been pharmacologically and selectively targeted in an Asyn model of PD. Here, we report that 7 weeks of peripheral administration of CB2 inverse agonist SMM-189 reduced phosphorylated (pSer129) Asyn in the substantia nigra compared to vehicle treatment. Additionally, SMM-189 delayed Asyn-induced immune cell infiltration into the brain as determined by flow cytometry, increased CD68 protein expression, and elevated wound-healing-immune-mediator gene expression. Additionally, peripheral immune cells increased wound-healing non-classical monocytes and decreased pro-inflammatory classical monocytes. In vitro analysis of RAW264.7 macrophages treated with lipopolysaccharide (LPS) and SMM-189 revealed increased phagocytosis as measured by the uptake of fluorescence of pHrodo E. coli bioparticles. Together, results suggest that targeting CB2 with SMM-189 skews immune cell function toward a phagocytic phenotype and reduces toxic aggregated species of Asyn. Our novel findings demonstrate that CB2 may be a target to modulate inflammatory and immune responses in proteinopathies.

Protective effects of alpinetin against interleukin-1β-exposed nucleus pulposus cells: Involvement of the TLR4/MyD88 pathway in a cellular model of intervertebral disc degeneration

Intervertebral disc degeneration (IDD) causes a variety of symptoms such as low back pain, disc herniation, and spinal stenosis, which can lead to high social and economic costs. Alpinetin has an anti-inflammatory potential, but its effect on IDD is unclear. Herein, we investigated the effect of alpinetin on IDD. To mimic an in vitro model of IDD, nucleus pulposus cells (NPCs) were exposed to interleukin 1β (IL-1β). The viability of NPCs was assessed by CCK-8 assay. The expression of Toll-like receptor 4 (TLR4), myeloid differentiation primary response protein 88 (MyD88), aggrecan, collagen-2, and matrix metalloproteinase-3 (MMP-3) was examined by qRT-PCR and western blotting. The protein levels of B cell lymphoma-2 (Bcl-2), Bcl-2-associated protein X (Bax), and cleaved caspase-3 were scrutinized by western blotting. The flow cytometry assay was performed to assess apoptosis of NPCs. The contents of inflammatory factors were examined by ELISA kits. Results showed that alpinetin repressed IL-1β-tempted activation of the TLR4/MyD88 pathway and apoptosis in NPCs. Alpinetin alleviated IL-1β-tempted inflammatory responses and oxidative stress in NPCs. Moreover, alpinetin lessened IL-1β-tempted extracellular matrix (ECM) degeneration in NPCs by enhancing the expression of aggrecan and collagen-2 and reducing the expression of MMP-3. The effects of alpinetin on IL-1β-exposed NPCs were neutralized by TLR4 upregulation. In conclusion, alpinetin repressed IL-1β-tempted apoptosis, inflammatory responses, oxidative stress, and ECM degradation in NPCs through the inactivation of the TLR4/MyD88 pathway.

Natural Compounds That Activate the KEAP1/Nrf2 Signaling Pathway as Potential New Drugs in the Treatment of Idiopathic Parkinson's Disease.

Recently, a single-neuron degeneration model has been proposed to understand the development of idiopathic Parkinson's disease based on (i) the extremely slow development of the degenerative process before the onset of motor symptoms and during the progression of the disease and (ii) the fact that it is triggered by an endogenous neurotoxin that does not have an expansive character, limiting its neurotoxic effect to single neuromelanin-containing dopaminergic neurons. It has been proposed that aminochrome is the endogenous neurotoxin that triggers the neurodegenerative process in idiopathic Parkinson's disease by triggering mitochondrial dysfunction, oxidative stress, neuroinflammation, dysfunction of both lysosomal and proteasomal protein degradation, endoplasmic reticulum stress and formation of neurotoxic alpha-synuclein oligomers. Aminochrome is an endogenous neurotoxin that is rapidly reduced by flavoenzymes and/or forms adducts with proteins, which implies that it is impossible for it to have a propagative neurotoxic effect on neighboring neurons. Interestingly, the enzymes DT-diaphorase and glutathione transferase M2-2 prevent the neurotoxic effects of aminochrome. Natural compounds present in fruits, vegetables and other plant products have been shown to activate the KEAP1/Nrf2 signaling pathway by increasing the expression of antioxidant enzymes including DT-diaphorase and glutathione transferase. This review analyzes the possibility of searching for natural compounds that increase the expression of DT-diaphorase and glutathione transferase through activation of the KEAP1/Nrf2 signaling pathway.

Deciphering the Effect of Hyaluronic Acid/Collagen Hydrogel for Pain Relief and Tissue Hydration in a Rat Model of Intervertebral Disc Degeneration.

Intervertebral disc (IVD) degeneration is one of the primary causes of low back pain, causing disability; hence, there is no regenerative nature of the current treatments. Hyaluronic acid (HA) was reported to facilitate tissue repair and alleviate pain. Herein, we determined the therapeutic effect of HA and type II collagen (COLII) hydrogel for tissue repair targeting pain in IVD degeneration. We implanted HA/COLII hydrogel following surgically induced disc injury at coccygeal levels in the rat tail model of pain. We assessed the efficacy of the HA/COLII hydrogel in reducing pain behaviour by using the von Frey assessment, protein expression of growth-associated protein (GAP) 43 for sensory nerve innervation, and disc hydration by magnetic resonance imaging (MRI). We observed the anti-nociceptive effect of the HA/COLII hydrogel in alleviating mechanical allodynia in rats. There was an inhibition of sensory hyperinnervation indicated by the GAP43 protein in the treatment group. We revealed an increase in T1ρ mapping of MRI, indicating that the hydrogel restored disc hydration in vivo. Our findings suggest the HA/COLII hydrogel alleviates pain behaviour, inhibits hyperinnervation and promotes disc hydration for tissue repair, implying that it is a potential candidate for the treatment of degenerative disc-associated low back pain.

An exploratory study of cervical disc degeneration model and mechanism of acupuncture therapy in rabbits

Acupuncture is an important therapy method in traditional Chinese medicine for treating intervertebral disc degeneration (IVDD), offering a wide range of applications. It is based on the theory of Chinese veterinary medicine and combines the stage of the disease course and individual differences for syndrome differentiation and treatment. However, there are few studies on the acupuncture treatment of cervical disc degeneration (CDD) in rabbits. Treatment based on syndrome differentiation is the basic principle of Chinese veterinary treatment. The selection of acupoints for external treatment should be based on individual etiology and pathogenesis. Nevertheless, most current studies do not follow this guideline. In this study, we established the CDD model and explored the mechanism of acupuncture treatment in alleviating CDD in rabbits by selecting a group of main acupoints including cervical Jiaji, Fengchi, Tianzhu, Naohu, Dazhui, and Houxi acupoints, combined with Western medicine's understanding of the pathogenesis of cervical spondylosis, from the anti-inflammatory, analgesic, and tissue-repairing perspectives. Magnetic resonance imaging (MRI) confirmed the successful establishment of the rabbit CDD model. Acupuncture stimulation reduced the increase of average and maximum neck temperature due to CDD in rabbits. The acupuncture treatment relieved the spinal disc damage in the neck of the rabbit, which also decreased the expression level of pro-apoptotic factor Bax and increased the expression level of anti-apoptotic factor Bcl-2. In addition, it can alleviate the abnormal apoptosis of rabbit intervertebral disc, decrease the expression level of inflammatory factors such as TNF-α, IL-1β, IL-2, and PGE2α, and alleviate the intense inflammation and pain response caused by CDD in rabbits. In conclusion, Acupuncture treatment can slow down the CDD of rabbits by regulating the inflammatory response and abnormal apoptosis of intervertebral disc tissue.

Neuroprotective effects of ellorarxine in neuronal models of degeneration.

Retinoic acid (RA) was first recognised to be important for the central nervous system (CNS) in its developmental regulatory role and, given this action, it has been proposed in the adult CNS to regulate plasticity and promote regeneration. These types of roles have included support of neurogenesis, induction of neurite outgrowth, and protection from neuronal death. These functions are predominantly mediated by the retinoic acid receptor (RAR) transcription factor, and hence agonists for the RARs have been tested in a variety of models of neurodegeneration. This present study employs several in vitro models less explored for the action of RAR agonists to reverse neurodegeneration. A series of assays are used in which neuronal cells are placed under the types of stress that have been linked to neurodegeneration, in particular amyotrophic lateral sclerosis (ALS), and the neuroprotective influence of a new potent agonist for RAR, ellorarxine, is tested out. In these assays, neuronal cells were subjected to excitotoxic stress induced by glutamate, proteostasis disruption caused by epoxomicin, and oxidative stress leading to stress granule formation triggered by sodium arsenite. Ellorarxine effectively reversed neuronal death in excitotoxic and proteostasis disruption assays and mitigated stress granule formation induced by sodium arsenite. This study also highlights for the first time the novel observation of RAR modulation of stress granules, although it is unknown whether this change in stress granules will be neuroprotective or potentially regenerative. Furthermore, the distribution of RAR agonists following intraperitoneal injection was assessed in mice, revealing preferential accumulation in the central nervous system, particularly in the spinal cord, compared to the liver. Gene expression studies in the spinal cord demonstrated that ellorarxine induces transcriptional changes at a low dose (0.01 mg/kg). These findings underscore the therapeutic potential of RAR agonists, such as ellorarxine, for ALS and potentially other neurodegenerative diseases.

REGULATION OF RETINOGENESIS IN PRENATAL ONTOGENESIS AND ITS DISRUPTION IN AGE-RELATED MACULAR DEGENERATION

One of the main causes of blindness in the elderly in developed countries is age-related macular degeneration (AMD) [1, 2, 3]. Active research is being conducted to identify putative molecular targets for the development of an effective strategy for conservative treatment of both atrophic and neovascular forms of this disease [4, 5, 6], but at the present stage, pharmacotherapy is effective only for some patients, and is insufficient to stop the progression of the disease or eliminate the damage that has already occurred. The most promising and promising potential treatment options are considered to be the introduction of cellular technologies with retinal cell replacement and targeted pharmacotherapy. The aim of this study was to investigate the role of the retinal development inducer, SIRT1 [7, 8, 9], in retinogenesis to justify pharmacological inhibition of this protein as a possible treatment option for neovascular AMD. Using immunohistochemistry and immunocytochemistry, we assessed the expression and subcellular localization of SIRT1 and its innate inhibitor DBC1 in the retina of fetuses, adults, and mice. We also studied SIRT1 in retinal progenitor cells derived from mice and humans, the former having been used in small interfering RNA studies. SIRT1 is widely expressed in the developing and adult retina and is a regulator of key genes in retinal development, namely PAX6, Nestin, and CRX [10, 11, 12]. Moreover, we found that photoreceptor progenitor cells were among the smallest cells in a heterogeneous mouse retinal progenitor cell population [13, 14, 15]. Collectively, these results provide a basis for manipulating SIRT1 expression in small retinal progenitors as a means to increase the yield of photoreceptors for transplantation in retinal degeneration models [16, 17]. Furthermore, SIRT1 was found to be highly expressed in human-derived choroidal neovascular membranes, allowing its activity to be pharmacologically inhibited by the drug nicotinamide as a potent regulator of angiogenic and hypoxic signaling in a human retinal pigment epithelial cell line at both the protein level using angiogenesis arrays and at the RNA level using whole genome microarrays [18, 19]. These results point to the SIRT1 inhibitor, Nicotinamide, as a possible agent for treatment of neovascular AMD. Further studies of Nicotinamide are warranted in animal models of AMD. To the best of our knowledge, this is the first time that a detailed analysis of SIRT1 as a regulator of both retinal development and choroidal neovascularization has been reported.

Therapeutic CRISPR epigenome editing of inflammatory receptors in the intervertebral disc

Low back pain (LBP) ranks among the leading causes of disability worldwide and generates a tremendous socioeconomic cost. Disc degeneration, a leading contributor to LBP, can be characterized by the breakdown of the extracellular matrix of the intervertebral disc (IVD), disc height loss, and inflammation. The inflammatory cytokine TNF-α has multiple signaling pathways, including proinflammatory signaling through Tumor Necrosis Factor Receptor 1 (TNFR1), and has been implicated as a primary mediator of disc degeneration. We tested our ability to regulate the TNFR1 signaling pathway in vivo, utilizing Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) epigenome editing to slow the progression of disc degeneration in rats. Sprague-Dawley rats were treated with TNF-α and CRISPR interference (CRISPRi)-based epigenome-editing therapeutics targeting TNFR1, showing decreased behavioral pain in a disc degeneration model. Surprisingly, while treatment with the vectors alone was therapeutic, the TNF-α injection became therapeutic after TNFR1 modulation. These results suggest direct inflammatory receptor modulation as a potent strategy for treating disc degeneration.

Preserving the Immune-Privileged Niche of the Nucleus Pulposus: Safeguarding Intervertebral Discs from Degeneration after Discectomy with Synthetic Mucin Hydrogel Injection.

Intervertebral disc (IVD) herniation is a prevalent spinal disorder, often necessitating surgical intervention such as microdiscectomy for symptomatic relief and nerve decompression. IVDs comprise a gel-like nucleus pulposus (NP) encased by an annulus fibrosus (AF), and their avascular nature renders them immune-privileged. Microdiscectomy exposes the residual NP to the immune system, precipitating an immune cell infiltration and attack that exacerbates IVD degeneration. While many efforts in the tissue engineering field are directed toward IVD regeneration, the inherently limited regenerative capacity due to the avascular and low-cellularity nature of the disc and the challenging mechanical environment of the spine often impedes success. This study, aiming to prevent IVD degeneration post-microdiscectomy, utilizes mucin-derived gels (Muc-gels) that form a gel at the surgical site, inspired by the natural mucin coating on living organisms to evade immune reorganization. It is shown that type I macrophages are present in severely degenerated human discs. Encapsulating IVDs within Muc-gels prevents fibrous encapsulation and macrophage infiltration in a mouse subcutaneous model. The injection of Muc-gels prevents IVD degeneration in a rat tail IVD degeneration model up to 24 weeks post-operation. Mechanistic investigations indicate that Muc-gels attenuate immune cell infiltration into NPs, offering durable protection against immune attack post-microdiscectomy.

Proton-activated chloride channel increases endplate porosity and pain in a mouse spine degeneration model.

Chronic low back pain (LBP) can severely affect daily physical activity. Aberrant osteoclast-mediated resorption leads to porous endplates, which allow the sensory innervation that causes LBP. Here, we report that expression of the proton-activated chloride (PAC) channel was induced during osteoclast differentiation in the porous endplates via a RANKL/NFATc1 signaling pathway. Extracellular acidosis evoked robust PAC currents in osteoclasts. An acidic environment of porous endplates and elevated PAC activation-enhanced osteoclast fusion provoked LBP. Furthermore, we found that genetic knockout of the PAC gene Pacc1 significantly reduced endplate porosity and spinal pain in a mouse LBP model, but it did not affect bone development or homeostasis of bone mass in adult mice. Moreover, both the osteoclast bone-resorptive compartment environment and PAC traffic from the plasma membrane to endosomes to form an intracellular organelle Cl channel had a low pH of approximately 5.0. The low pH environment activated the PAC channel to increase sialyltransferase St3gal1 expression and sialylation of TLR2 in the initiation of osteoclast fusion. Aberrant osteoclast-mediated resorption is also found in most skeletal disorders, including osteoarthritis, ankylosing spondylitis, rheumatoid arthritis, heterotopic ossification, and enthesopathy. Thus, elevated Pacc1 expression and PAC activity could be a potential therapeutic target for the treatment of LBP and osteoclast-associated pain.

Exploring the protective effects of Qiju Granule in a rat model of dry age-related macular degeneration

AimThe aim of this study was to evaluate the potential protective effect of Qiju Granule in a rat model of age-related macular degeneration (AMD) and investigate the underlying mechanisms involved. MethodsRats were injected intravenously with 40 mg/kg of sodium iodate (SI) to induce a dry AMD model. The rats in the treatment group received three different doses of Qiju Granule once a day via gavage, while the rats in the control group were given an equal volume of physiological saline. On day 14 and day 28 following the intervention, various methods were employed to evaluate retinal function and structure, including electroretinography (ERG), optical coherence tomography (OCT), and histological examination. The expression of glial fibrillary acidic protein (GFAP), basic fibroblast growth factor (bFGF), brain-derived neurotrophic factor (BDNF), and ciliary neurotrophic factor (CNTF) was assessed via immunofluorescence. Beyond immunofluorescence, the mRNA levels of bFGF, BDNF, and CNTF were quantitatively determined using real-time polymerase chain reaction (qRT-PCR). ResultsRats treated with Qiju Granule exhibited significant improvements in both retinal function and structure compared to the model group. The most noteworthy effects were observed at a high dose of Qiju Granule. Furthermore, the expression levels of bFGF, BDNF, and CNTF were significantly unregulated in the treated groups compared to the model group. ConclusionsQiju Granule demonstrated a protective effect on the retina in the SI-induced rat model of AMD. The protective mechanism may be attributed to the upregulation of retinal neurotrophic factors expression.