Preclinical Evaluation of Bioactive Small Intestinal Submucosa-PMMA Bone Cement for Vertebral Augmentation.

In vertebroplasty and kyphoplasty, bioinert poly(methyl methacrylate) (PMMA) bone cement is a conventional filler employed for quick stabilization of osteoporotic vertebral compression fractures (OVCFs). However, because of the poor osteointegration, excessive stiffness, and high curing temperature of PMMA, the implant loosens, the adjacent vertebrae refracture, and thermal necrosis of the surrounding tissue occurs frequently. This investigation addressed these issues by incorporating the small intestinal submucosa (SIS) into PMMA (SIS-PMMA). In vitro analyses revealed that this new SIS-PMMA bone cement had improved porous structure, as well as reduced compressive modulus and polymerization temperature compared with the original PMMA. Furthermore, the handling properties of SIS-PMMA bone cement were not significantly different from PMMA. The in vitro effect of PMMA and SIS-PMMA was investigated on MC3T3-E1 cells via the Transwell insert model to mimic the clinical condition or directly by culturing cells on the bone cement samples. The results indicated that SIS addition substantially enhanced the proliferation and osteogenic differentiation of MC3T3-E1 cells. Additionally, the bone cement's biomechanical properties were also assessed in a decalcified goat vertebrae model with a compression fracture, which indicated the SIS-PMMA had markedly increased compressive strength than PMMA. Furthermore, it was proved that the novel bone cement had good biosafety and efficacy based on the International Standards and guidelines. After 12 weeks of implantation, SIS-PMMA indicated significantly more osteointegration and new bone formation ability than PMMA. In addition, vertebral bodies with cement were also extracted for the uniaxial compression test, and it was revealed that compared with the PMMA-implanted vertebrae, the SIS-PMMA-implanted vertebrae had greatly enhanced maximum strength. Overall, these findings indicate the potential of SIS to induce efficient fixation between the modified cement surface and the host bone, thereby providing evidence that the SIS-PMMA bone cement is a promising filler for clinical vertebral augmentation.

Distinguished biomimetic dECM system facilitates early detection of metastatic breast cancer cells.

Breast cancer is the most prevalent malignant tumor affecting women's health. Bone is the most common distant metastatic organ, worsening the quality of life and increasing the mortality of patients. Early detection of breast cancer bone metastasis is urgent for halting disease progression and improving tumor prognosis. Recently, extracellular matrix (ECM) with biomimetic tissue niches opened a new avenue for tumor models in vitro. Here, we developed a biomimetic decellularized ECM (dECM) system to recapitulate bone niches at different situations, bone mimetic dECM from osteoblasts (BM-ECM) and bone tumor mimetic dECM from osteosarcoma cells (OS-ECM). The two kinds of dECMs exhibited distinct morphology, protein composition, and distribution. Interestingly, highly metastatic breast cancer cells tended to adhere and migrate on BM-ECM, while lowly metastatic breast cancer cells preferred the OS-ECM niche. Epithelial-to-mesenchymal transition was a potential mechanism to initiate the breast cancer cell migration on different biomimetic dECMs. Importantly, in the nude mice model, the dECM system captured metastatic breast cancer cells as early as 10 days after orthotopic transplantation in mammary gland pads, with higher signal on BM-ECM than that on OS-ECM. Collectively, the biomimetic dECM system might be a promising tumor model to distinguish the metastatic ability of breast cancer cells in vitro and to facilitate early detection of metastatic breast cancer cells in vivo, contributing to the diagnosis of breast cancer bone metastasis.

RDASNet: Image Denoising via a Residual Dense Attention Similarity Network.

In recent years, thanks to the performance advantages of convolutional neural networks (CNNs), CNNs have been widely used in image denoising. However, most of the CNN-based image-denoising models cannot make full use of the redundancy of image data, which limits the expressiveness of the model. We propose a new image-denoising model that aims to extract the local features of the image through CNN and focus on the global information of the image through the attention similarity module (ASM), especially the global similarity details of the image. Furthermore, dilation convolution is used to enlarge the receptive field to better focus on the global features. Moreover, avg-pooling is used to smooth and suppress noise in the ASM to further improve model performance. In addition, through global residual learning, the effect is enhanced from shallow to deep layers. A large number of experiments show that our proposed model has a better image-denoising effect, including quantitative and visual results. It is more suitable for complex blind noise and real images.

A Dilated Residual Network for Turbine Blade ICT Image Artifact Removal.

Artifacts are divergent strip artifacts or dark stripe artifacts in Industrial Computed Tomography (ICT) images due to large differences in density among the components of scanned objects, which can significantly distort the actual structure of scanned objects in ICT images. The presence of artifacts can seriously affect the practical application effectiveness of ICT in defect detection and dimensional measurement. In this paper, a series of convolution neural network models are designed and implemented based on preparing the ICT image artifact removal datasets. Our findings indicate that the RF (receptive field) and the spatial resolution of network can significantly impact the effectiveness of artifact removal. Therefore, we propose a dilated residual network for turbine blade ICT image artifact removal (DRAR), which enhances the RF of the network while maintaining spatial resolution with only a slight increase in computational load. Extensive experiments demonstrate that the DRAR achieves exceptional performance in artifact removal.

ATBF1 is a potential diagnostic marker of histological grade and functions via WNT5A in breast cancer.

BACKGROUND: Histological grade has been demonstrated to be an important factor of breast cancer outcome and is associated with cell differentiation and is currently being evaluated via H&E-stained sections. Molecular biomarkers are essential to improve the accuracy of histological grading. ATBF1, a large transcription factor, has been considered a tumor suppressor gene with frequent mutations or deletions in multiple cancers. In breast cancer, ATBF1 was reported to function in cell differentiation and mammary development. However, its role in the clinic has rarely been reported. METHODS: Breast cancer tissues (BCTs) and adjacent noncancerous tissues (ANCTs) were collected to analyze the expression of ATBF1 at the mRNA and protein levels. Three anti-ATBF1 antibodies recognizing independent peptides of ATBF1 (N-terminal end, middle region and C-terminal end) were applied for IHC staining. Small interfering RNA (siRNA) was used to silence ATBF1 expression and to investigate the roles of ATBF1 in MCF7 cells. Microarrays were introduced to analyze the differentially expressed genes, enriched GO terms and KEGG terms regulated by ATBF1 and its potential downstream genes, which were further confirmed in vitro and in clinical samples. RESULTS: The expression of ATBF1 was reduced in BCTs at both the mRNA and protein levels compared with that in ANCTs. ATBF1 protein was predominantly localized in the nucleus of ANCTs but in the cytoplasm of BCTs. Both the mRNA and protein levels of ATBF1 were significantly correlated with histological grade. Consistently, knockdown of ATBF1 increased stemness marker expression and reduced differentiation markers in vitro. Further analysis identified WNT5A as an essential downstream gene of ATBF1 in breast cancer cells. Treatment of WNT5A disrupted cell proliferation induced by ATBF1 silencing. In BCTs, a significant correlation was observed between the expression of WNT5A and ATBF1. CONCLUSION: The results indicated that ATBF1 expression might be a useful diagnostic marker associated with histological grade and breast cancer malignancy. WNT5A and its signaling pathway are novel mechanisms by which ATBF1 contributes to breast cancer tumorigenesis.

Enhancement of Tendon Repair Using Tendon-Derived Stem Cells in Small Intestinal Submucosa via M2 Macrophage Polarization.

(1) Background: Reconstruction of Achilles tendon defects and prevention of postoperative tendon adhesions were two serious clinical problems. In the treatment of Achilles tendon defects, decellularized matrix materials and mesenchymal stem cells (MSCs) were thought to address both problems. (2) Methods: In vitro, cell adhesion, proliferation, and tenogenic differentiation of tendon-derived stem cells (TDSCs) on small intestinal submucosa (SIS) were evaluated. RAW264.7 was induced by culture medium of TDSCs and TDSCs-SIS scaffold groups. A rat Achilles tendon defect model was used to assess effects on tendon regeneration and antiadhesion in vivo. (3) Results: SIS scaffold facilitated cell adhesion and tenogenic differentiation of TDSCs, while SIS hydrogel coating promoted proliferation of TDSCs. The expression of TGF-ß and ARG-1 in the TDSCs-SIS scaffold group were higher than that in the TDSCs group on day 3 and 7. In vivo, the tendon regeneration and antiadhesion capacity of the implanted TDSCs-SIS scaffold was significantly enhanced. The expression of CD163 was significantly highest in the TDSCs-SIS scaffold group; meanwhile, the expression of CD68 decreased more significantly in the TDSCs-SIS scaffold group than the other two groups. (4) Conclusion: This study showed that biologically prepared SIS scaffolds synergistically promote tendon regeneration with TDSCs and achieve antiadhesion through M2 polarization of macrophages.

Exosome-based delivery nanoplatforms: next-generation theranostic platforms for breast cancer.

Breast cancer is the most frequent type of malignancy, and the leading cause of cancer-related death in women across the globe. Exosomes are naturally derived 50-150 nm nanovesicles with a variety of bioactive molecules to regulate the complex intracellular pathways involved in all stages of breast cancer development. Exosomes are also considered as a potential new generation of natural nanocarriers due to their intriguing endogenous functionalities. Recently, the development of exosome-based delivery nanoplatforms that combine the inherent unique advantages of exosomes with advanced nanotechnology has emerged as a promising area. In the present review, we first declare the fundamental principles of the relationship between exosomes and breast cancer, ranging from the initiation and progression of breast cancer, to drug resistance. More efforts are made to present a comprehensive overview of the recent advances of exosome nanotechnology for breast cancer therapy, including natural exosomes from different cell types, engineered exosomes with cargo loading and membrane modification, and artificial bionic exosomes with more stable and scalable properties. Based on the recent advanced nanotechnologies, exosome-based delivery nanoplatforms have been considered as the next-generation theranostic platforms, which sheds the light on the achievement of the clinical translation of exosomes for breast cancer therapy.

BMSCs and Osteoblast-Engineered ECM Synergetically Promotes Osteogenesis and Angiogenesis in an Ectopic Bone Formation Model.

Bone mesenchymal stem cells (BMSCs) have been extensively used in bone tissue engineering because of their potential to differentiate into multiple cells, secrete paracrine factors, and attenuate immune responses. Biomaterials are essential for the residence and activities of BMSCs after implantation in vivo. Recently, extracellular matrix (ECM) modification with a favorable regenerative microenvironment has been demonstrated to be a promising approach for cellular activities and bone regeneration. The aim of the present study was to evaluate the effects of BMSCs combined with cell-engineered ECM scaffolds on osteogenesis and angiogenesis in vivo. The ECM scaffolds were generated by osteoblasts on the small intestinal submucosa (SIS) under treatment with calcium (Ca)-enriched medium and icariin (Ic) after decellularization. In a mouse ectopic bone formation model, the SIS scaffolds were demonstrated to reduce the immune response, and lower the levels of immune cells compared with those in the sham group. Ca/Ic-ECM modification inhibited the degradation of the SIS scaffolds in vivo. The generated Ca/Ic-SIS scaffolds ectopically promoted osteogenesis according to the results of micro-CT and histological staining. Moreover, BMSCs on Ca/Ic-SIS further increased the bone volume percentage (BV/TV) and bone density. Moreover, angiogenesis was also enhanced by the Ca/Ic-SIS scaffolds, resulting in the highest levels of neovascularization according to the data ofCD31 staining. In conclusion, osteoblast-engineered ECM under directional induction is a promising strategy to modify biomaterials for osteogenesis and angiogenesis. BMSCs synergetically improve the properties of ECM constructs, which may contribute to the repair of large bone defects.

Initial assessment of treatment of talar posterior process fractures with open reduction and percutaneous fixation.

The purpose of this study was to provide an initial assessment of treatment for talar posterior process fractures using open reduction and internal fixation (ORIF) through posteromedial approach and percutaneous screw fixation. From January 2014 to December 2018, 12 cases with displaced fracture of talar posterior process were treated in our department. The clinical and radiological results were assessed after 4 and 12 months of operation with Visual Analog Scale (VAS) pain and American Orthopedic Foot and Ankle Society (AOFAS) scores. ORIF was performed in four of the cases and percutaneous screw fixation was performed in eight of the cases. The average follow-up period was 13 months. Complications such as wound infection, nerve injury, screw loosening, malunion or nonunion of fracture were absent. For clinical assessment, considerable mprovements were observed for the AOFAS and VAS scores at 4 and 12 months postoperatively for both techniques. There was no significant difference for AOFAS scores and VAS scores between the two techniques (p > 0.05). Both techniques showed good functional outcome and were performed for posterior talar process fracture following the fracture displacement guidelines. Percutaneous screw fixation treatment with computer-assisted three-dimensional evaluation shortened the operation time and reduced incidences of surgical complications.

Osteoblast/fibroblast coculture derived bioactive ECM with unique matrisome profile facilitates bone regeneration.

Extracellular matrix (ECM) with mimetic tissue niches was attractive to facilitate tissue regeneration in situ via recruitment of endogenous cells and stimulation of self-healing process. However, how to engineer the complicate tissue specific ECM with unique matrisome in vitro was a challenge of ECM-based biomaterials in tissue engineering and regenerative medicine. Here, we introduced coculture system to engineer bone mimetic ECM niche guided by cell-cell communication. In the cocultures, fibroblasts promoted osteogenic differentiation of osteoblasts via extracellular vesicles. The generated ECM (MN-ECM) displayed a unique appearance of morphology and biological components. The advantages of MN-ECM were demonstrated with promotion of multiple cellular behaviors (proliferation, adhesion and osteogenic mineralization) in vitro and bone regeneration in vivo. Moreover, proteomic analysis was used to clarify the molecular mechanism of MN-ECM, which revealed a specific matrisome signature. The present study provides a novel strategy to generate ECM with tissue mimetic niches via cell-cell communication in a coculture system, which forwards the development of tissue-bioactive ECM engineering along with deepening the understanding of ECM niches regulated by cells for bone tissue engineering.

ATBF1 Participates in Dual Functions of TGF-ß via Regulation of Gene Expression and Protein Translocalization.

TGF-ß is a critical cytokine to regulate multiple pathophysiological functions. For tumor development and progression, TGF-ß was reported to play dual functions as a tumor suppressor and epithelial-mesenchymal transition (EMT) inducer. The mechanism of the TGF-ß signaling pathway is essential for TGF-ß/Smad-targeted therapy in clinic. Here, ATBF1 was demonstrated to participate in dual functions of TGF-ß via different ways. On one hand, ATBF1 expression level was associated with EMT and migration induced by TGF-ß. After TGF-ß treatment, ATBF1 expression was reduced in a dose- and time-dependent manner, along with the alteration of cell morphology and EMT marker expression. Knockdown of ATBF1 by siRNA further promoted EMT progression and cell migration. On the other hand, ATBF1 localization was associated with cell proliferation inhibited by TGF-ß. The number of cells with nucleus localization of ATBF1 in TGF-ß activation group was much higher than that in control group. After that, knockdown of ATBF1 by siRNA rescued the inhibition of cell proliferation affected by TGF-ß. These data revealed that ATBF1 is a key gene for the dual roles of TGF-ß, which may contribute to future therapy.

[Research progress of decellularized extracellular matrix hydrogel in regenerative medicine].

Decellularized extracellular matrix (dECM) has been widely used as a scaffold for regenerative medicine due to its high biomimetic and excellent biocompatibility. As a functional polymer material with high water content and controlled fluidity, hydrogel is very promising for some minimally invasive surgery in clinical practice. In recent years, with the rapid development of hydrogel theory and technology, dECM hydrogel has gradually become a research hotspot in the field of regenerative medicine. In this paper, the related researches in recent years are reviewed regarding the preparation of dECM hydrogel and its preclinical application. The future clinical use is also prospected.

In-Situ Mutation Detection by Magnetic Beads-Probe Based on Single Base Extension and Its Application in Genotyping of Hepatitis B Virus Pre-C Region 1896nt Locus Single Nucleotide Polymorphisms.

Hepatitis B virus (HBV) is closely related to occurrence and development of viral hepatitis. A mutation of 1896nt locus in its pre-C region can promote replication of HBV DNA and improve stability of pre-genome RNA structure, and can even help HBV evade immune clearance. In this study, magnetic beads-probe (MBs@probe) method, combined with single base extension (SBE) technology, was developed for in-situ mutation detection of HBV pre-C region 1896nt locus. Before successfully completing the genotyping of 165 HBV samples, the crucial reaction conditions were first optimized, such as SBE temperature, MBs size and amount, and probe concentration on the surface of MBs. Experimental results showed that these conditions had significant effects on MBs@probe in-situ mutation detection. Comprehensive considerations, such as 58 °C of SBE temperature, high fluorescence intensity and signal-to-noise ratios (SNRs) were obtained when MBs@probe complex was made by 100 µg of 300 nm-MBs and 3.0 µM of probes in the system. Finally, 1896nt locus mutation in pre-C region of 165 HBV samples was successfully genotyped, among which 71 HBV samples were wild types and the remaining 94 samples were mutant types. Meanwhile, 14 randomly chosen samples were taken to further analyze fluorescence intensity and SNRs respectively, and sequencing results for the first two samples were consistent with results from the MBs@probe in-situ mutation detection method. Compared with two-color fluorescence hybridization (TCFH) genotyping technology, this method generally improves the SNRs to more than 10 (which is more than 2-fold), has higher reliability and is more suitable to detect SNPs for known sites.

ECM coating modification generated by optimized decellularization process improves functional behavior of BMSCs.

Bone mesenchymal stem cells (BMSCs) have been widely applied in tissue engineering and regenerative medicine. However, small number of BMSCs and loss of stem cell characteristics after expansion in vitro limited clinical use of BMSCs. In the present study, osteoblasts were cultured to lay down extracellular matrix (ECM) and then the cells were removed (decellularization) to generate ECM coating substrates. The decellularization process was optimized to maximally remove cells and cellular components, along with integrated ECM retained which was demonstrated to be beneficial for BMSCs expansion in vitro. After decellularization, only less than 2% of residual DNA and cellular proteins were detected in TFFF-ECM (decellularized by triton X-100 (T) and three freeze/thaw cycles (FFF)), which was much less than that in TN-ECM generated by traditional decellularization method (triton X-100 (T) and NH4OH (N)). Meanwhile, ECM components and structure were preserved best after decellularization by TFFF method. More ECM proteins were detected, and structure proteins (fibronectin and collagen) exhibited as classic network fibers in TFFF-ECM. Functionally, all kinds of decellularized ECM (dECM) were demonstrated to promote BMSCs proliferation and osteogenic differentiation capacity, thus maintain the stemness of BMSCs. Importantly, cells cultured on TFFF-ECM grew faster than the cells on other kinds of dECM at early stage and TFFF-ECM was beneficial to preserve stemness of BMSCs with high expression of OCT4 and NANOG when cultured in vitro. Proteomic analysis showed the proteins in ECM functioned in multiple biological activities and signaling pathways, which contributed to stemness maintenance of BMSC. Thus, the mild decellularization process optimized in this study enhanced the effectiveness of dECM for BMSCs culture in vitro and maybe further applied to BMSCs based tissue repair.

Isoquercitrin promotes the osteogenic differentiation of osteoblasts and BMSCs via the RUNX2 or BMP pathway.

AIM: Isoquercitrin is widely present in fruits, vegetables and medicinal herbs. As a natural phytoestrogen, isoquercitrin has been considered a possible osteoporosis prevention option to avoid the risk of hormone therapy. MATERIALS AND METHODS: The cell proliferation of osteoblasts and bone mesenchymal stem cells (BMSCs) was examined by cell counting kit-8 (CCK-8). The osteogenic differentiation was evaluated by real-time qPCR, ALP staining and Alizarin Red S staining. Small interfering RNA (siRNA) was used to knockdown the expression of runt-related transcription factor 2 (RUNX2). RESULTS: The cell proliferation of osteoblasts and BMSCs was promoted by isoquercitrin at low concentrations. High concentrations of isoquercitrin promoted the osteogenic differentiation via RUNX2 expression in osteoblasts and via the bone morphogenetic protein (BMP) pathway in BMSCs. Inhibition of RUNX2 expression in osteoblasts by siRNA or addition of noggin to the culture medium of BMSCs reduced the effects of osteogenic differentiation induced by isoquercitrin. CONCLUSIONS: These data suggest that isoquercitrin is a natural potential osteoinductive compound and might be valuable for the prevention/treatment of bone disorders.

Activity-dependent Synaptic Recruitment of Neuroligin 1 in Spinal Dorsal Horn Contributed to Inflammatory Pain.

Neuroligin 1 (NLGN1), a cell adhesion molecule present at excitatory glutamatergic synapses, has been shown to be critical for synaptic specialization and N-methyl-d-aspartate (NMDA)-subtype glutamate receptor-dependent synaptic plasticity. Whether and how NLGN1 is engaged in nociceptive behavioral sensitization remains largely unknown. In this study, we found an activity-dependent regulation of NLGN1 synaptic expression in pain-related spinal cord dorsal horns of mice. The enhancement of neuronal activity by pharmacological activation of NMDA receptor (NMDAR) or removal of GABAergic inhibition in intact mice significantly increased NLGN1 concentration at synaptosomal membrane fraction. Intraplantar injection of complete Freund's adjuvant (CFA) also increased the NLGN1 expression at synapses. NMDAR might act through Ca2+/calmodulin-dependent protein kinase II (CaMKII) and Src-family protein tyrosine kinase member Fyn to induce the synaptic redistribution of NLGN1. We also found that one of the important roles of NLGN1 was to facilitate the clustering of NMDAR at synapses. The NLGN1-targeting siRNA suppressed the synaptic expression of GluN2B-containing NMDAR in CFA-injected mice and meanwhile, attenuated the inflammatory mechanical allodynia and thermal hypersensitivity. These data suggested that tissue injury-induced synaptic redistribution of NLGN1 was involved in the development of pain hypersensitivity through facilitating the synaptic incorporation of NMDARs.

Inhibition of protein tyrosine phosphatase 1B in spinal cord dorsal horn of rats attenuated diabetic neuropathic pain.

Protein tyrosine phosphatase 1B (PTP1B) has been shown to dephosphorylate and inactivate insulin receptors, which contributes to the pathogenesis of diabetes. Neuropathic pain is one of the severe complications that results from diabetic neuropathy. However, whether PTP1B was involved in the development of diabetic neuropathic pain is largely unknown. The current study illustrated that PTP1B was located in spinal cord dorsal horn neurons of Sprague-Dawley rats. Western blot analysis demonstrated that the diabetic neuropathic pain induced by intraperitoneal injection of streptozotocin was associated with an increased protein expression and a dynamic redistribution of spinal PTP1B into excitatory glutamatergic synapses. We found that PTP1B operated to stimulate Src kinase and enhance the tyrosine phosphorylation of N-methyl-D-aspartate (NMDA) subtype of glutamate receptors. The siRNA-mediated knockdown of PTP1B in streptozotocin-injected rats repressed Src activity, decreased NMDA receptor phosphorylation and alleviated the thermal hyperalgesia and mechanical allodynia. A similar analgesia against diabetic neuropathic pain was also achieved when PTP1B activity was manipulated by a chemical PTP Inhibitor or PTP1B(C215S) mutant. These data revealed a regulated expression of PTP1B in spinal cord dorsal horn of rats after diabetic neuropathy, and demonstrated that inhibition of PTP1B was beneficial for the treatment of pain hypersensitivity related to diabetes.

A Cell-Engineered Small Intestinal Submucosa-Based Bone Mimetic Construct for Bone Regeneration.

Extracellular matrix (ECM)-ornamented biomaterials have attracted attention due to their high potential to improve the biofunctionality of original materials. It is thought that ECM with a bone mimetic microenvironment generated by the specific induction of osteoblasts would be more beneficial for bone regeneration than a regular ECM. In this study, we developed an osteogenic and mineralized ECM construct (Os/M-ECM-SIS) under the guidance of osteoblasts on a small intestinal submucosa (SIS) scaffold cotreated with icariin and calcium. The generated Os/M-ECM-SIS scaffolds exhibited similar morphology and inorganic components as natural bone and higher mechanical strength than ECM-SIS. Cell adhesion, proliferation, and differentiation of osteoblasts and fibroblasts were also enhanced in the cells cultured on the Os/M-ECM-SIS scaffolds. The Os/M-ECM-SIS scaffolds even promoted transdifferentiation of fibroblasts with an upregulation of osteogenic differentiation markers. In a calvarial defect model, new bone formation was greatly enhanced in defects implanted with the Os/M-ECM-SIS scaffolds compared with ECM-SIS scaffolds. Further study showed that the Os/M-ECM-SIS scaffolds promoted bone regeneration in vitro and in vivo via the Bmp/Smad-signaling pathway. Thus, this work proposes a valuable method for generating a mineralized bone mimetic scaffold with SIS as off-the-shelf bone graft substitute that provides an excellent osteogenic microenvironment, making it suitable for application in bone tissue engineering.

Adenosine A1 receptor potentiated glycinergic transmission in spinal cord dorsal horn of rats after peripheral inflammation.

Adenosine is present at the extracellular space within spinal cord dorsal horn and engaged in the processing of nociceptive sensory signals. Systemic or spinal administration of exogenous adenosine produces a potent analgesia against pathological pain. Here we found that inhibitory glycinergic neurotransmission was an important target for adenosine regulation. In spinal cord slices from intact rats, adenosine increased the inhibitory postsynaptic currents mediated by glycine receptors (GlyRs). In spinal slices from Complete Freund's Adjuvant-injected rats, adenosine potentiated glycinergic transmission to a more degree than in control rats. This synaptic potentiation was dependent on the activation of adenosine A1 receptor (A1R), and attributed to the modification of postsynaptic GlyRs function. The Gi protein-coupled A1R typically signals through Gαi/cAMP-dependent protein kinase (PKA) and Gßγ pathways. We found that blockade of either Gαi/PKA or Gßγ signaling attenuated the ability of adenosine to increase glycinergic synaptic responses in inflamed rats. To identify which GlyRs subunit was subjected to A1R regulation, we recorded glycine-evoked whole-cell currents in HEK293T cells co-transfected with A1R and distinct GlyRs subunit. We found that α1, the most abundant functional GlyRs subunit in adult spinal cord, was insensitive to A1R activation. However, when GlyRs α3 subunit or α1ins subunit, a longer α1 isoform, was co-expressed with A1R, adenosine caused a significant increase of glycinergic currents. Inhibition of PKA and Gßγ abolished the stimulatory effects of A1R on α3 and α1ins, respectively. These data suggested that A1R might potentiate glycinergic transmission through Gαi/PKA/α3 and Gßγ/α1ins pathways in inflamed rat.

A Novel Approach to Utilize Icariin as Icariin-Derived ECM on Small Intestinal Submucosa Scaffold for Bone Repair.

Icariin (Ic) has been demonstrated as a potent osteoinductive compound for bone tissue engineering. However, toxic side effects of the drug and poor biocompatibility of drug delivery systems (DDSs) still limit its application for bone repair in the clinic. To overcome these disadvantages and utilize the osteoinductivity of Ic, we developed a novel method to utilize Ic as an Ic-derived osteoinductive extracellular matrix (ECM) on small intestinal submucosa (SIS) (Ic-ECM-SIS). The generated Ic-ECM-SIS scaffolds, as a natural construct, exhibited much better biocompatibility (including cell adhesion, cell survival and cell proliferation) than Ic-SIS scaffolds generated by traditional DDSs. Meanwhile, osteogenic differentiation was promoted by Ic-ECM-SIS with higher expression of alkaline phosphatase, bone sialoprotein and osteocalcin than ECM-SIS, which was same as Ic-SIS. BMP-4 expression was further increased in the cells on Ic-ECM-SIS compared to that on Ic-SIS. A mouse calvarial defect model was introduced to evaluate the function of Ic-ECM-SIS on bone regeneration in vivo. The bone regeneration was enhanced in the defects implanted with Ic-ECM-SIS, with a higher new bone formation ratio (BV/TV) than the defects implanted with ECM-SIS or Ic-SIS. Angiogenesis was also promoted by Ic-ECM-SIS implantation when compared with ECM-SIS or Ic-SIS. Thus, this work proposes a novel method for applying a drug as a drug-derived ECM-modified scaffold for bone tissue engineering.