Fibrosis-4 index stratifies risks of hepatocellular carcinoma in patients with chronic hepatitis C.

BACKGROUND AND AIMS: Risk stratification for patients with a higher risk of hepatocellular carcinoma (HCC) is crucial. We aimed to investigate the role of the Fibrosis-4 (FIB-4) index in predicting chronic hepatitis C (CHC)-related HCC. METHODS: A retrospective cohort study consecutively included treatment-naive CHC patients receiving longitudinal follow-up at the National Taiwan University Hospital from 1986 to 2014. The clinical data were collected and traced for HCC development. Multivariable Cox proportional hazard regression analysis was used to investigate the predictors for HCC. RESULTS: A total of 1285 patients in the ERADICATE-C cohort were included. The median age was 54, 56% were females, and 933 had HCV viremia. There were 33%, 38%, and 29% of patients having FIB-4 index <1.45, 1.45-3.25, and ≥3.25, respectively. After a median of 9-year follow-up, 186 patients developed HCC. Multivariable analysis revealed that older age, AFP≥20 ng/mL, cirrhosis, and a higher FIB-4 index were independent predictors for HCC. Compared with patients with FIB-4 index <1.45, those with FIB-4 1.45-3.25 had a 5.51-fold risk (95% confidence interval [CI]: 2.65-11.46), and those with FIB-4 ≥ 3.25 had 7.45-fold risk (95% CI: 3.46-16.05) of HCC. In CHC patients without viremia, FIB-4 index 1.45-3.25 and FIB-4 ≥ 3.25 increased 6.78-fold and 16.77-fold risk of HCC, respectively, compared with those with FIB-4 < 1.45. CONCLUSION: The baseline FIB-4 index can stratify the risks of HCC in untreated CHC patients, even those without viremia. The FIB-4 index should thus be included in the management of CHC.

MicroRNA-137-3p Protects PC12 Cells Against Oxidative Stress by Downregulation of Calpain-2 and nNOS.

The imbalance between excess reactive oxygen species (ROS) generation and insufficient antioxidant defenses contribute to a range of neurodegenerative diseases. High ROS levels damage cellular macromolecules such as DNA, proteins and lipids, leading to neuron vulnerability and eventual death. However, the underlying molecular mechanism of the ROS regulation is not fully elucidated. Recently, an increasing number of studies suggest that microRNAs (miRNAs) emerge as the targets in regulating oxidative stress. We recently reported the neuroprotective effect of miR-137-3p for brachial plexus avulsion-induced motoneuron death. The present study is sought to investigate whether miR-137-3p also could protect PC12 cells against hydrogen peroxide (H2O2) induced neurotoxicity. By using cell viability assay, ROS assay, gene and protein expression assay, we found that PC-12 cells exposed to H2O2 exhibited decreased cell viability, increased expression levels of calpain-2 and neuronal nitric oxide synthase (nNOS), whereas a decreased miR-137-3p expression. Importantly, restoring the miR-137-3p levels in H2O2 exposure robustly inhibited the elevated nNOS, calpain-2 and ROS expression levels, which subsequently improved the cell viability. Furthermore, the suppressive effect of miR-137-3p on the elevated ROS level under oxidative stress was considerably blunted when we mutated the binding site of calpain-2 targted by miR-137-3p, suggesting the critical role of calpain-2 involving the neuroprotective effect of miR-137-3p. Collectively, these findings highlight the neuroprotective role of miR-137-3p through down-regulating calpain and NOS activity, suggesting its potential role for combating oxidative stress insults in the neurodegenerative diseases.

MiR-137-3p rescue motoneuron death by targeting calpain-2.

Brachial plexus root avulsion (BPRA) is a type of injury that leads to motor function loss as a result of motoneurons (MNs) degeneration. Here we identified that the reduced expression of rat miR-137-3p in the ventral horn of spinal cord was associated with MNs death. However, the pathophysiological role of miR-137-3p in root avulsion remains poorly understood. We demonstrated that the calcium-activated neutral protease-2 (calpain-2) was a direct target gene of miR-137-3p with miR-137-3p binding to the 3'-untranslated region of calpain-2. Silencing of calpain-2 suppressed the expression of neuronal nitric oxide synthase (nNOS), a primary source of nitric oxide (NO). After avulsion 2 weeks, up-regulation of miR-137-3p in the spinal cord reduced calpain-2 levels and nNOS expression inside spinal MNs, resulting in an amelioration of the MNs death. These events provide new insight into the mechanism by which upregulation of miR-137-3p can impair MN survival in the BPRA.

Propofol Protects Against H2O2-Induced Oxidative Injury in Differentiated PC12 Cells via Inhibition of Ca(2+)-Dependent NADPH Oxidase.

Propofol (2,6-diisopropylphenol) is a widely used general anesthetic with anti-oxidant activities. This study aims to investigate protective capacity of propofol against hydrogen peroxide (H2O2)-induced oxidative injury in neural cells and whether the anti-oxidative effects of propofol occur through a mechanism involving the modulation of NADPH oxidase (NOX) in a manner of calcium-dependent. The rat differentiated PC12 cell was subjected to H2O2 exposure for 24 h to mimic a neuronal in vitro model of oxidative injury. Our data demonstrated that pretreatment of PC12 cells with propofol significantly reversed the H2O2-induced decrease in cell viability, prevented H2O2-induced morphological changes, and reduced the ratio of apoptotic cells. We further found that propofol attenuated the accumulation of malondialdehyde (biomarker of oxidative stress), counteracted the overexpression of NOX core subunit gp91(phox) (NOX2) as well as the NOX activity following H2O2 exposure in PC12 cells. In addition, blocking of L-type Ca(2+) channels with nimodipine reduced H2O2-induced overexpression of NOX2 and caspase-3 activation in PC12 cells. Moreover, NOX inhibitor apocynin alone or plus propofol neither induces a significant downregulation of NOX activity nor increases cell viability compared with propofol alone in the PC12 cells exposed to H2O2. These results demonstrate that the protective effects of propofol against oxidative injury in PC12 cells are mediated, at least in part, through inhibition of Ca(2+)-dependent NADPH oxidase.

Time-specific microRNA changes during spinal motoneuron degeneration in adult rats following unilateral brachial plexus root avulsion: ipsilateral vs. contralateral changes.

BACKGROUND: Spinal root avulsion induces multiple pathophysiological events consisting of altered levels of specific genes and proteins related to inflammation, apoptosis, and oxidative stress, which collectively result in the death of the affected motoneurons. Recent studies have demonstrated that the gene changes involved in spinal cord injury can be regulated by microRNAs, which are a class of short non-coding RNA molecules that repress target mRNAs post-transcriptionally. With consideration for the time course of the avulsion-induced gene expression patterns within dying motoneurons, we employed microarray analysis to determine whether and how microRNAs are involved in the changes of gene expression induced by pathophysiological events in spinal cord motoneurons. RESULTS: The expression of a total of 3,361 miRNAs in the spinal cord of adult rats was identified. Unilateral root-avulsion resulted in significant alterations in miRNA expression. In the ipsilateral half compared to the contralateral half of the spinal cord, on the 3rd day after the injury, 55 miRNAs were upregulated, and 24 were downregulated, and on the 14th day after the injury, 36 miRNAs were upregulated, and 23 were downregulated. The upregulation of miR-146b-5p and miR-31a-3p and the downregulation of miR-324-3p and miR-484 were observed. Eleven of the miRNAs, including miR-21-5p, demonstrated a sustained increase; however, only miR-466c-3p presented a sustained decrease 3 and 14 days after the injury. More interestingly, 4 of the miRNAs, including miR-18a, were upregulated on the 3rd day but were downregulated on the 14th day after injury.Some of these miRNAs target inflammatory-response genes in the early stage of injury, and others target neurotransmitter transport genes in the intermediate stages of injury. The altered miRNA expression pattern suggests that the MAPK and calcium signaling pathways are consistently involved in the injury response. CONCLUSIONS: This analysis may facilitate the understanding of the time-specific altered expression of a large set of microRNAs in the spinal cord after brachial root avulsion.

Lithium enhances survival and regrowth of spinal motoneurons after ventral root avulsion.

BACKGROUND: During the clinical treatment of the brachial plexus root avulsion (BPRA), reimplantation surgery can not completely repair the motor function of the hand because the axonal growth velocity of the spinal motoneurons (MNs) is too slow to re-innervate the intrinsic hand muscles before muscle atrophy. Here, we investigated whether lithium can enhance the regenerative capacity of the spinal MNs in a rat model of BPRA. RESULTS: The avulsion and immediate reimplantation of the C7 and C8 ventral roots were performed and followed with daily intraperitoneal administration of a therapeutic concentrationof LiCl. After a 20 week long-term rehabilitation, the motor function recovery of the injured forepaw was studied by a grasping test. The survival and regeneration of MNs were checked by choline acetyltransferase (ChAT) immunofluorescence and by Fluoro-Gold (FG) retrograde labeling through the median and ulnar nerves of the ventral horn MNs. The number and diameter of the nerve fibers in the median nerve were assessed by toluidine blue staining. Our results showed that lithium plus reimplantation therapy resulted in a significantly higher grasping strength of the digits of the injured forepaw. Lithium plus reimplantation allowed 45.1% ± 8.11% of ChAT-positive MNs to survive the injury and increased the number and diameter of nerve fibers in the median nerve. The number of FG-labeled regenerative MNs was significantly elevated in all of the reimplantation animals. Our present data proved that lithium can enhance the regenerative capacity of spinal MNs. CONCLUSIONS: These results suggest that immediate administration of lithium could be used to assist reimplantation surgery in repairing BPRA injuries in clinical treatment.

Injectable chondroitin sulfate-grafted self-antioxidant hydrogels ameliorate nucleus pulposus degeneration against overactive inflammation.

Overactive inflammatory cascade accompanied by oxidative stress in the nucleus pulposus exacerbates intervertebral disc degeneration (IVDD). Hydrogels have been demonstrated to be promising in treating IVDD, yet they remain less efficacious in the case of anti-inflammation associated with antioxidation. In this study, we designed an injectable self-antioxidant hydrogel (HA/CS) with enhanced inflammation inhibitory performance for delivering chondroitin sulfate (CS) with well-documented anti-inflammatory property to treat IVDD. The hydrogel was rapidly formed via dynamic boronate ester bonding between furan/phenylboronic acid and furan/dopamine-modified hyaluronic acid (HA), and mechanically enhanced by Diels-Alder reaction-induced secondary crosslinking, partial dopamine groups of which contribute to grafting phenylboronic acid-modified CS (CS-PBA). This hydrogel exhibits favorable injectability, mechanical property, and pH-responsive delivery behavior. The dopamine moiety endows the hydrogel with efficient antioxidative property. By sustained delivery of CS, the HA/CS hydrogel is well competent to inhibit inflammatory cytokine expression and maintain anabolic/catabolic balance in an inflammation-simulated environment. Most importantly, the HA/CS hydrogel significantly ameliorates degeneration in a puncture-induced IVDD rat model. The self-antioxidant HA/CS hydrogel designed in this work may serve as a novel and promising therapeutic platform for IVDD.

Parathyroid hormone treatment partially reverses endplate remodeling and attenuates low back pain in animal models of spine degeneration.

Low back pain (LBP) is one of the most prevalent diseases affecting quality of life, with no disease-modifying therapy. During aging and spinal degeneration, the balance between the normal endplate (EP) bilayers of cartilage and bone shifts to more bone. The aged/degenerated bony EP has increased porosity because of osteoclastic remodeling activity and may be a source of LBP due to aberrant sensory innervation within the pores. We used two mouse models of spinal degeneration to show that parathyroid hormone (PTH) treatment induced osteogenesis and angiogenesis and reduced the porosity of bony EPs. PTH increased the cartilaginous volume and improved the mechanical properties of EPs, which was accompanied by a reduction of the inflammatory factors cyclooxygenase-2 and prostaglandin E2. PTH treatment furthermore partially reversed the innervation of porous EPs and reversed LBP-related behaviors. Conditional knockout of PTH 1 receptors in the nucleus pulposus (NP) did not abolish the treatment effects of PTH, suggesting that the NP is not the primary source of LBP in our mouse models. Last, we showed that aged rhesus macaques with spontaneous spinal degeneration also had decreased EP porosity and sensory innervation when treated with PTH, demonstrating a similar mechanism of PTH action on EP sclerosis between mice and macaques. In summary, our results suggest that PTH treatment could partially reverse EP restructuring during spinal regeneration and support further investigation into this potentially disease-modifying treatment strategy for LBP.

Single Impact Injury of Vertebral Endplates Without Structural Disruption, Initiates Disc Degeneration Through Piezo1 Mediated Inflammation and Metabolism Dysfunction.

STUDY DESIGN: In vitro experimental study. OBJECTIVE: To establish an axial impact injury model of intervertebral disc (IVD) and to investigate if a single impact injury without endplate structural disruption could initiate intervertebral disc degeneration (IDD), and what is the roles of Piezo1 in this process. SUMMARY OF BACKGROUND DATA: Although IDD process has been confirmed to be associated with structural failures such as endplate fractures, whether a single impact injury of the endplates without structural disruption could initiate IDD remains controversial. Previous studies reported that Piezo1 mediated inflammation participated in the progression of IDD induced by mechanical stretch; however, the roles of Piezo1 in IVD impact injury remain unknown. METHODS: Rats spinal segments were randomly assigned into Control, Low, and High Impact groups, which were subjected to pure axial impact loading using a custom-made apparatus, and cultured for 14 days. The degenerative process was investigated by using histomorphology, real-time Polymerase Chain Reaction(PCR), western-blot, immunofluorescence, and energy metabolism of IVD cell. The effects of Piezo1 were investigated by using siRNA transfection, real-time PCR, western-blot, and immunofluorescence. RESULTS: The discs in both of the impact groups presented degenerative changes after 14 days, which showed significant up-regulation of Piezo1, NLRP3 inflammasome, the catabolic (MMP-9, MMP-13), and pro-inflammatory gene (IL-1ß) expression than that of the control group (P < 0.05), accompanied by significantly increased release of ATP, lactate, nitric oxide (NO), and glucose consumption of IVD cells at first 7 days. Silencing Piezo1 reduced the activation of NLRP3 inflammasome and IL-1ß expression in the nucleus pulposus induced by impact injury. CONCLUSION: It demonstrated that not only fracture of the endplate but also a single impact injury without structural impairment could also initiate IDD, which might be mediated by activation of Piezo1 induced inflammation and abnormal energy metabolism of IVD cells.Level of Evidence: N/A.

Application of electrophysiological measures in degenerative cervical myelopathy.

Degenerative cervical myelopathy (DCM) is one of the leading causes of progressive spinal cord dysfunction in the elderly. Early diagnosis and treatment of DCM are essential to avoid permanent disability. The pathophysiology of DCM includes chronic ischemia, destruction of the blood-spinal cord barrier, demyelination, and neuronal apoptosis. Electrophysiological studies including electromyography (EMG), nerve conduction study (NCS), motor evoked potentials (MEPs) and somatosensory evoked potentials (SEPs) are useful in detecting the presymptomatic pathological changes of the spinal cord, and thus supplementing the early clinical and radiographic examinations in the management of DCM. Preoperatively, they are helpful in detecting DCM and ruling out other diseases, assessing the spinal cord compression level and severity, predicting short- and long-term prognosis, and thus deciding the treatment methods. Intra- and postoperatively, they are also useful in monitoring neurological function change during surgeries and disease progression during follow-up rehabilitation. Here, we reviewed articles from 1979 to 2021, and tried to provide a comprehensive, evidence-based review of electrophysiological examinations in DCM. With this review, we aim to equip spinal surgeons with the basic knowledge to diagnosis and treat DCM using ancillary electrophysiological tests.

Elimination of Senescent Cells by Senolytics Facilitates Bony Endplate Microvessel Formation and Mitigates Disc Degeneration in Aged Mice.

Senolytics are a class of drugs that selectively eliminate senescent cells and ameliorate senescence-associated disease. Studies have demonstrated the accumulation of senescent disc cells and the production of senescence-associated secretory phenotype decrease the number of functional cells in degenerative tissue. It has been determined that clearance of senescent cell by senolytics rejuvenates various cell types in several human organs, including the largest avascular structure, intervertebral disc (IVD). The microvasculature in the marrow space of bony endplate (BEP) are the structural foundation of nutrient exchange in the IVD, but to date, the anti-senescence effects of senolytics on senescent vascular endothelial cells in the endplate subchondral vasculature remains unclear. In this study, the relationships between endothelial cellular senescence in the marrow space of the BEP and IVD degeneration were investigated using the aged mice model. Immunofluorescence staining was used to evaluate the protein expression of P16, P21, and EMCN in vascular endothelial cells. Senescence-associated ß-galactosidase staining was used to investigate the senescence of vascular endothelial cells. Meanwhile, the effects of senolytics on cellular senescence of human umbilical vein endothelial cells were investigated using a cell culture model. Preliminary results showed that senolytics alleviate endothelial cellular senescence in the marrow space of BEP as evidenced by reduced senescence-associated secretory phenotype. In the aged mice model, we found decreased height of IVD accompanied by vertebral bone mass loss and obvious changes to the endplate subchondral vasculature, which may lead to the decrease in nutrition transport into IVD. These findings may provide evidence that senolytics can eliminate the senescent cells and facilitate microvascular formation in the marrow space of the BEP. Targeting senescent cellular clearance mechanism to increase nutrient supply to the avascular disc suggests a potential treatment value of senolytics for IVD degenerative diseases.

Cocktail of isobavachalcone and curcumin enhance eradication of Staphylococcus aureus biofilm from orthopedic implants by gentamicin and alleviate inflammatory osteolysis.

Orthopedic device-related infection (ODRI) caused by Staphylococcus aureus, especially methicillin-resistant S. aureus (MRSA) biofilm may lead to persist infection and severe inflammatory osteolysis. Previous studies have demonstrated that both isobavachalcone and curcumin possess antimicrobial activity, recent studies also reveal their antiosteoporosis, anti-inflammation, and immunoregulatory effect. Thus, this study aims to investigate whether the combination of isobavachalcone and curcumin can enhance the anti-S. aureus biofilm activity of gentamicin and alleviate inflammatory osteolysis in vivo. EUCAST and a standardized MBEC assay were used to verify the synergy between isobavachalcone and curcumin with gentamicin against planktonic S. aureus and its biofilm in vitro, then the antimicrobial and immunoregulatory effect of cocktail therapy was demonstrated in a femoral ODRI mouse model in vivo by µCT analysis, histopathology, quantification of bacteria in bone and myeloid-derived suppressor cell (MDSC) in bone marrow. We tested on standard MSSA ATCC25923 and MRSA USA300, 5 clinical isolated MSSA, and 2 clinical isolated MRSA strains and found that gentamicin with curcumin (62.5-250 µg/ml) and gentamicin with isobavachalcone (1.56 µg/ml) are synergistic against planktonic MSSA, while gentamicin (128 µg/ml) with curcumin (31.25-62.5, 250-500 µg/ml) and gentamicin (64-128 µg/ml) with isobavachalcone (1.56-12.5 µg/ml) exhibit synergistic effect against MSSA biofilm. Results of further study revealed that cocktail of 128 µg/ml gentamicin together with 125 µg/ml curcumin +6.25 µg/ml isobavachalcone showed promising biofilm eradication effect with synergy against USA300 biofilm in vitro. Daily intraperitoneal administration of 20 mg/kg/day isobavachalcone, 20 mg/kg/day curcumin, and 20 mg/kg/day gentamicin, can reduce inflammatory osteolysis and maintain microarchitecture of trabecular bone during orthopedic device-related MRSA infection in mice. Cocktail therapy also enhanced reduction of MDSC M1 polarization in peri-implant tissue, suppression of MDSC amplification in bone marrow, and Eradication of USA300 biofilm in vivo. Together, these results suggest that the combination of isobavachalcone and curcumin as adjuvants administrated together with gentamicin significantly enhances its antimicrobial effect against S. aureus biofilm, and can also modify topical inflammation in ODRI and protect bone microstructure in vivo, which may serve as a potential treatment strategy, especially for S. aureus induced ODRI.

Bone tissue engineering scaffolds with HUVECs/hBMSCs cocultured on 3D-printed composite bioactive ceramic scaffolds promoted osteogenesis/angiogenesis.

Background: /Objective: Tissue engineering involves scaffolds, cells and growth factors, among which growth factors have limited applications due to potential safety risks and high costs. Therefore, an alternative approach to exogenously induce osteogenesis is desirable. Considering that osteogenesis and angiogenesis are coupled, a system of human umbilical vein endothelial cells (HUVECs) and human bone mesenchymal stem cells (hBMSCs) coculture is more biologically adapted to the microenvironment in vivo and can mediate osteogenesis and angiogenesis via paracrine signalling. Hence, in this study, a HUVECs/hBMSCs coculture system with appropriate cell and medium proportions was established. The substrate for the coculture system was a 3D-printed composite bioceramic scaffold (ß-TCP/CaSiO3) based on a previous study. The aim of this study was to explore the potential of this system for bone tissue engineering. Methods: Bioactive ceramic scaffolds for tissue engineering were fabricated via a 3D Bioplotter™ system. The coculture system for in vitro and in vivo studies consisted of direct contact between HUVECs and hBMSCs cultured on the 3D-printed scaffolds. Results: The proportions of HUVECs/hBMSCs and medium components were determined by cell viability, and the coculture system showed negligible cytotoxicity. CD31 secreted by HUVECs formed strings, and cells tended to aggregate in island chain-like arrays. Real-time cell tracking showed that HUVECs were recruited by hBMSCs, and the integrin expression by HUVECs was upregulated. Ultimately, osteogenic and angiogenic marker gene expression and protein secretion were upregulated. Moreover, the obtained bone tissue engineering scaffolds could induce early osteogenic protein secretion and capillary tube formation in nude rats. Conclusion: These bone tissue engineering scaffolds without exogenous growth factors exhibited the ability to promote osteogenesis/angiogenesis. Translational potential of this article: The fabricated 3D-printed bioactive ceramic scaffolds could provide mechanical, biodegradable and bioadaptive support for personalized bone regeneration. In addition, the bone tissue engineering scaffolds exhibited the ability to promote osteogenesis/angiogenesis without the addition of exogenous growth factors, thus mitigating safety risks. Although application of the HUVECs/hBMSCs coculture system might be a time-consuming process, further development of cord blood storage could be beneficial for multicell coculture.

Panax notoginseng saponins attenuate intervertebral disc degeneration by reducing the end plate porosity in lumbar spinal instability mice.

Although painkillers could alleviate some of the symptoms, there are no drugs that really cope with the intervertebral disc degeneration (IDD) at present, so it is urgent to find a cure that could prevent or reverse the progression of IDD. During the development of IDD, the cartilaginous end plates (EPs) become hypertrophic and porous by the increase of osteoclast activities, which hinder the penetration of nutrition. The compositional and structural degeneration of the EP may cause both nutritional as well as mechanical impairment to the nucleus pulposus (NP) so that developing drugs that target the degenerating EP may be another option in addition to targeting the NP. In the lumbar spine instability mouse model, we found increased porosity in the cartilaginous EP, accompanied by the decrease in total intervertebral disc volume. Panax notoginseng saponins (PNS), a traditional Chinese patent drug with anti-osteoclastogenesis effect, could alleviate IDD by inhibiting aberrant osteoclast activation in the porous EP. Further in vitro experiment validated that PNS inhibit the receptor activator of nuclear factor kappa-Β ligand-induced osteoclast differentiation, while the transcriptional activation of PAX6 may be involved in the mechanism, which had been defined as an inhibitory transcription factor in osteoclastogenesis. These findings may provide a novel therapeutic strategy for IDD.

Cationic Polymer Brush-Modified Carbon Nanotube-Meditated eRNA LINC02569 Silencing Attenuates Nucleus Pulposus Degeneration by Blocking NF-κB Signaling Pathway and Alleviate Cell Senescence.

Enhancer RNAs (eRNAs) are noncoding RNAs that synthesized at active enhancers. eRNAs have important regulatory characteristics and appear to be significant for maintenance of cell identity and information processing. Series of functional eRNAs have been identified as potential therapeutic targets for multiple diseases. Nevertheless, the role of eRNAs on intervertebral disc degeneration (IDD) is still unknown yet. Herein, we utilized the nucleus pulposus samples of patients and identified a key eRNA (LINC02569) with the Arraystar eRNA Microarray. LINC02569 mostly locates in nucleus and plays an important role in the progress of IDD by activating nuclear factor kappa-B (NF-κB) signaling pathway. We used a cationic polymer brush coated carbon nanotube (oCNT-pb)-based siRNA delivery platform that we previously designed, to transport LINC02569 siRNA (si-02569) to nucleus pulposus cells. The siRNA loaded oCNT-pb accumulated in nucleus pulposus cells with lower toxicity and higher transfection efficiency, compared with the traditional siRNA delivery system. Moreover, the results showed that the delivery of si-02569 significantly alleviated the inflammatory response in the nucleus pulposus cells via inhibiting P65 phosphorylation and preventing its transfer into the nucleus, and meanwhile alleviated cell senescence by decreasing the expression of P21. Altogether, our results highlight that eRNA (LINC02569) plays important role in the progression of IDD and could be a potential therapeutic target for alleviation of IDD.

Simultaneous incorporation of PTH(1-34) and nano-hydroxyapatite into Chitosan/Alginate Hydrogels for efficient bone regeneration.

Tissue regeneration based on the utilization of artificial soft materials is considered a promising treatment for bone-related diseases. Here, we report cranial bone regeneration promoted by hydrogels that contain parathyroid hormone (PTH) peptide PTH(1-34) and nano-hydroxyapatite (nHAP). A combination of the positively charged natural polymer chitosan (CS) and negatively charged sodium alginate led to the formation of hydrogels with porous structures, as shown by scanning electron microscopy. Rheological characterizations revealed that the mechanical properties of the hydrogels were almost maintained upon the addition of nHAP and PTH(1-34). In vitro experiments showed that the hydrogel containing nHAP and PTH(1-34) exhibited strong biocompatibility and facilitated osteogenic differentiation of rat bone marrow mesenchymal stem cells (rBMSCs) via the Notch signaling pathway, as shown by the upregulated expression of osteogenic-related proteins. We found that increasing the content of PTH(1-34) in the hydrogels resulted in enhanced osteogenic differentiation of BMSCs. Implantation of the complex hydrogel into a rat cranial defect model led to efficient bone regeneration compared to the rats treated with the hydrogel alone or with nHAP, indicating the simultaneous therapeutic effect of nHAP and PTH during the treatment process. Both the in vitro and in vivo results demonstrated that simultaneously incorporating nHAP and PTH into hydrogels shows promise for bone regeneration, suggesting a new strategy for tissue engineering and regeneration in the future.

Fabrication of a Polylactide-Glycolide/Poly-ε-Caprolactone/Dextran/Plastrum Testudinis Extract Composite Anti-Inflammation Nanofiber Membrane via Electrospinning for Annulus Fibrosus Regeneration.

Tissue engineering is a promising approach for the treatment of chronic lower back pain (LBP) caused by intervertebral disc degeneration (IDD) resulting from degeneration and inflammation of annulus fibrosus (AF) tissue. However, scaffold with an anti-inflammatory effect on AF cells has not been reported. In this study, we fabricated a polylactide-glycolide (PLGA)/poly-ε-caprolactone (PCL)Zdextran (DEX) composite membrane loaded with plastrum testudinis extract (PTE), a Traditional Chinese Medicine herbal extract, via electrospinning. The membranes were characterized by mechanical measurements and scanning electron microscopy (SEM). Using an in vitro inflammation model induced by interleukin (IL)-1ß, the cytocompatibility and anti-inflammatory effects of the composites were investigated by CCK-8 assay and flow cytometry. Potential regulatory mechanisms were examined by RT-qPCR and Western blotting. The results showed that the P10P8D2 (PLGA 10 g, PCL 8 g, DEX 2 g) composite nanofiber membrane exhibited the most uniform diameter distribution, best mechanical properties, a moderate degradation rate, and the best cytocompatibility characteristics. The optimal concentration of PTE was 120 µg/mL. Importantly, P10P8D2 combined with PTE exhibited anti-inflammatory and cell proliferation promotion effects. Moreover, the NF-κBB/NLRP3/IL-ß signaling pathway was inactivated. Our findings suggested that the nanofiber membrane composed of P10P8D2 and PTE has anti-inflammatory and pro-proliferation effects on AF cells. It may provide an effective strategy for AF tissue regeneration.

Single-Cell RNA-Seq Analysis Reveals Macrophage Involved in the Progression of Human Intervertebral Disc Degeneration.

Intervertebral disc degeneration (IDD) has been considered as the primary pathological mechanism that underlies low back pain. Understanding the molecular mechanisms underlying human IDD is imperative for making strategies to treat IDD-related diseases. Herein, we report the molecular programs, lineage progression patterns, and paths of cellular communications during the progression of IDD using single-cell RNA sequencing (scRNA-seq) on nucleus pulposus (NP) cells from patients with different grades of IDD undergoing discectomy. New subtypes of cells and cell-type-specific gene signatures of the metabolic homeostatic NP cells (Met NPC), adhesive NP cells (Adh NPC), inflammatory response NP cells (IR NPC), endoplasmic reticulum stress NP cells (ERS NPC), fibrocartilaginous NP cells (Fc NPC), and CD70 and CD82+ progenitor NP cells (Pro NPC) were identified. In the late stage of IDD, the IR NPC and Fc NPC account for a large proportion of NPC. Importantly, immune cells including macrophages, T cells, myeloid progenitors, and neutrophils were also identified, and further analysis showed that significant intercellular interaction between macrophages and Pro NPC occurred via MIF (macrophage migration inhibitory factor) and NF-kB signaling pathways during the progression of IDD. In addition, dynamic polarization of macrophage M1 and M2 cell subtypes was found in the progression of IDD, and gene set functional enrichment analysis suggested a significant role of the macrophage polarization in regulating cell metabolism, especially the Pro NPC. Finally, we found that the NP cells in the late degenerative stage were mainly composed of the cell types related to inflammatory and endoplasmic reticulum (ER) response, and fibrocartilaginous activity. Our results provided new insights into the identification of NP cell populations at single-cell resolution and at the relatively whole-transcriptome scale, accompanied by cellular communications between immune cells and NP cells, and discriminative markers in relation to specific cell subsets. These new findings present clues for effective and functional manipulation of human IDD-related bioremediation and healthcare.

Regulatory Roles of Bone in Neurodegenerative Diseases.

Osteoporosis and neurodegenerative diseases are two kinds of common disorders of the elderly, which often co-occur. Previous studies have shown the skeletal and central nervous systems are closely related to pathophysiology. As the main structural scaffold of the body, the bone is also a reservoir for stem cells, a primary lymphoid organ, and an important endocrine organ. It can interact with the brain through various bone-derived cells, mostly the mesenchymal and hematopoietic stem cells (HSCs). The bone marrow is also a place for generating immune cells, which could greatly influence brain functions. Finally, the proteins secreted by bones (osteokines) also play important roles in the growth and function of the brain. This article reviews the latest research studying the impact of bone-derived cells, bone-controlled immune system, and bone-secreted proteins on the brain, and evaluates how these factors are implicated in the progress of neurodegenerative diseases and their potential use in the diagnosis and treatment of these diseases.

Panax Notoginseng Saponins Prevent Bone Loss by Promoting Angiogenesis in an Osteoporotic Mouse Model.

With the aging of the population and the extension of life expectancy, osteoporosis is becoming a global epidemic. Although there are several drugs used to treat osteoporosis in clinical practice, such as parathyroid hormone or bisphosphonates, they all have some serious side effects. Therefore, a safer drug is called for osteoporosis, especially for the prevention in the early stage of the disease, not only the treatment in the later stage. Panax notoginseng saponin (PNS), a traditional Chinese herb, has been used as anti-ischemic drug due to its function on improving vascular circulation. In order to verify whether Panax notoginseng saponins (PNS) could be used to prevent osteoporosis, ovariectomy (OVX) was induced in female C57BL/C6J mice, followed by orally administration with 40 mg/kg/d, 80 mg/kg/d, and 160 mg/kg/d of three different dosages of PNS for 9 weeks. Serum biochemical analysis, micro-CT, histological evaluation, and immunostaining of markers of osteogenesis and angiogenesis were performed in the sham, osteoporotic (OVX), and treatment (OVX+PNS) groups. Micro-CT and histological evaluation showed that compared to sham group, the bone mass of OVX group reduced significantly, while it was significantly restored in the moderate-dose PNS (40 mg/kg and 80 mg/kg) treatment groups. The expression of CD31 and osteocalcin (OCN) in the bone tissue of treatment group also increased, suggesting that PNS activated osteogenesis and angiogenesis, which subsequently increased the bone mass. These results confirmed the potential function of PNS on the prevention of osteoporosis. However, in the high dose of PNS (160 mg/kg) group, the antiosteoportic effect had been eliminated, which also suggested the importance of proper dose of PNS for the prevention and treatment of osteoporosis in postmenopausal women.