Injectable conductive hydrogel remodeling microenvironment and mimicking neuroelectric signal transmission after spinal cord injury.

Severe spinal cord injury (SCI) leads to dysregulated neuroinflammation and cell apoptosis, resulting in axonal die-back and the loss of neuroelectric signal transmission. While biocompatible hydrogels are commonly used in SCI repair, they lack the capacity to support neuroelectric transmission. To overcome this limitation, we developed an injectable silk fibroin/ionic liquid (SFMA@IL) conductive hydrogel to assist neuroelectric signal transmission after SCI in this study. The hydrogel can form rapidly in situ under ultraviolet (UV) light. The mechanical supporting and neuro-regenerating properties are provided by silk fibroin (SF), while the conductive capability is provided by the designed ionic liquid (IL). SFMA@IL showed attractive features for SCI repair, such as anti-swelling, conductivity, and injectability. In vivo, SFMA@IL hydrogel used in rats with complete transection injuries was found to remodel the microenvironment, reduce inflammation, and facilitate neuro-fiber outgrowth. The hydrogel also led to a notable decrease in cell apoptosis and the achievement of scar-free wound healing, which saved 45.6 ± 10.8 % of spinal cord tissue in SFMA@IL grafting. Electrophysiological studies in rats with complete transection SCI confirmed SFMA@IL's ability to support sensory neuroelectric transmission, providing strong evidence for its signal transmission function. These findings provide new insights for the development of effective SCI treatments.

A multi-functional double cross-linked chitosan hydrogel with tunable mechanical and antibacterial properties for skin wound dressing.

Chitosan hydrogels with essential antibacterial properties and biocompatibility have great potential in tissue engineering and regeneration medicine. However, pure chitosan hydrogel could be limited by insufficient mechanical properties. In this work, we designed a multi-functional chitosan hydrogel based on the combination of chitosan methacrylate (CTSMA) and sulfhydrated chitosan (CTSSH), which is cross-linked simultaneously by free-radical polymerization reaction and Thiol-ene reaction. The CTSMA/CTSSH (CMS) hydrogels displayed superior tissue adhesive and mechanical properties when compared to pure CTSMA hydrogel. Additionally, the resulting hydrogels exhibited potent antimicrobial effects against both E. coli and S. aureus. Besides, the CMS hydrogels exhibited good biocompatibility as demonstrated by cytotoxicity and cell proliferation experiments using fibroblasts cells (L929) and adipose-derived stem cells (ADSCs). In vivo experiment, the repairing effect of hydrogels on full-thickness skin defect model in rats was studied. Histological and immunohistochemical staining results showed that CMS hydrogels promoted angiogenesis, dermal repair and epidermal regeneration. Overall, the study highlights the potential of the CMS hydrogels as a promising biomaterial in wound healing applications.

Porous microneedle patch with sustained delivery of extracellular vesicles mitigates severe spinal cord injury.

The transplantation of mesenchymal stem cells-derived secretome, particularly extracellular vesicles is a promising therapy to suppress spinal cord injury-triggered neuroinflammation. However, efficient delivery of extracellular vesicles to the injured spinal cord, with minimal damage, remains a challenge. Here we present a device for the delivery of extracellular vesicles to treat spinal cord injury. We show that the device incorporating mesenchymal stem cells and porous microneedles enables the delivery of extracellular vesicles. We demonstrate that topical application to the spinal cord lesion beneath the spinal dura, does not damage the lesion. We evaluate the efficacy of our device in a contusive spinal cord injury model and find that it reduces the cavity and scar tissue formation, promotes angiogenesis, and improves survival of nearby tissues and axons. Importantly, the sustained delivery of extracellular vesicles for at least 7 days results in significant functional recovery. Thus, our device provides an efficient and sustained extracellular vesicles delivery platform for spinal cord injury treatment.

Controlled delivery of a neurotransmitter-agonist conjugate for functional recovery after severe spinal cord injury.

Despite considerable unmet medical needs, effective pharmacological treatments that promote functional recovery after spinal cord injury remain limited. Although multiple pathological events are implicated in spinal cord injuries, the development of a microinvasive pharmacological approach that simultaneously targets the different mechanisms involved in spinal cord injury remains a formidable challenge. Here we report the development of a microinvasive nanodrug delivery system that consists of amphiphilic copolymers responsive to reactive oxygen species and an encapsulated neurotransmitter-conjugated KCC2 agonist. Upon intravenous administration, the nanodrugs enter the injured spinal cord due to a disruption in the blood-spinal cord barrier and disassembly due to damage-triggered reactive oxygen species. The nanodrugs exhibit dual functions in the injured spinal cord: scavenging accumulated reactive oxygen species in the lesion, thereby protecting spared tissues, and facilitating the integration of spared circuits into the host spinal cord through targeted modulation of inhibitory neurons. This microinvasive treatment leads to notable functional recovery in rats with contusive spinal cord injury.

Single-Cell Sequencing Reveals the Optimal Time Window for Anti-Inflammatory Treatment in Spinal Cord Injury.

Though the occurrence of neuroinflammation after spinal cord injury (SCI) is essential for antigen clearance and tissue repair, excessive inflammation results in cell death and axon dieback. The effect of anti-inflammatory medicine used in clinical treatment remains debatable owing to the inappropriate therapeutic schedule that does not align with the biological process of immune reaction. A better understanding of the immunity process is critical to promote effective anti-inflammatory therapeutics. However, cellular heterogeneity, which results in complex cellular functions, is a major challenge. This study performs single-cell RNA sequencing by profiling the tissue proximity to the injury site at different time points after SCI. Depending on the analysis of single-cell data and histochemistry observation, an appropriate time window for anti-inflammatory medicine treatment is proposed. This work also verifies the mechanism of typical anti-inflammatory medicine methylprednisolone sodium succinate (MPSS), which is found attributable to the activation inhibition of cells with pro-inflammatory phenotype through the downregulation of pathways such as TNF, IL2, and MIF. These pathways can also be provided as targets for anti-inflammatory treatment. Collectively, this work provides a therapeutic schedule of 1-3 dpi (days post injury) to argue against classical early pulse therapy and provides some pathways for target therapy in the future.

Downregulation of UBE4B promotes CNS axon regrowth and functional recovery after stroke.

The limited intrinsic regrowth capacity of corticospinal axons impedes functional recovery after cortical stroke. Although the mammalian target of rapamycin (mTOR) and p53 pathways have been identified as the key intrinsic pathways regulating CNS axon regrowth, little is known about the key upstream regulatory mechanism by which these two major pathways control CNS axon regrowth. By screening genes that regulate ubiquitin-mediated degradation of the p53 proteins in mice, we found that ubiquitination factor E4B (UBE4B) represses axonal regrowth in retinal ganglion cells and corticospinal neurons. We found that axonal regrowth induced by UBE4B depletion depended on the cooperative activation of p53 and mTOR. Importantly, overexpression of UbV.E4B, a competitive inhibitor of UBE4B, in corticospinal neurons promoted corticospinal axon sprouting and facilitated the recovery of corticospinal axon-dependent function in a cortical stroke model. Thus, our findings provide a translatable strategy for restoring corticospinal tract-dependent functions after cortical stroke.

Extracellular vesicles from colorectal cancer cells promote metastasis via the NOD1 signalling pathway.

Pattern-recognition receptors (PRRs) have been shown to promote tumour metastasis via sensing tumour cell-derived small extracellular vesicles (EVs). Nucleotide-binding oligomerisation domain 1 (NOD1), a cytoplasmic PRR, plays a role in colorectal cancer (CRC) by detecting bacterial products. However, the precise mechanisms underlying the effects of NOD1, following identification of CRC cell-derived EVs (CRC-EVs), to potentiate CRC liver metastasis (CRC-LM), remain poorly understood. Here, we demonstrate that CRC-EVs activate NOD1 in macrophages to initiate secretion of inflammatory cytokines and chemokines. NOD1-activated macrophages also promote CRC cell migration, while in a murine model of liver metastasis (LM), NOD1-deficient mice exhibit reduced metastasis following CRC-EV treatment. Furthermore, cell division cycle 42 (CDC42), a small Rho guanosine-5'-triphosphate (GTP)ase, is delivered by CRC-EVs into macrophages where it activates NOD1. In addition, EVs from the plasma of patients with CRC-LM mediate NOD1 activation in human peripheral blood mononuclear cells. Moreover, high NOD1 expression in tumour tissues is associated with poor prognosis of CRC-LM. Our findings suggest that CRC-EVs activate NOD1 to promote tumour metastasis, thus, NOD1 may serve as a potential target in the diagnosis and treatment of CRC-LM.

Rationally Designed, Self-Assembling, Multifunctional Hydrogel Depot Repairs Severe Spinal Cord Injury.

Following severe spinal cord injury (SCI), dysregulated neuroinflammation causes neuronal and glial apoptosis, resulting in scar and cystic cavity formation during wound healing and ultimately the formation of an atrophic microenvironment that inhibits nerve regrowth. Because of this complex and dynamic pathophysiology, a systemic solution for scar- and cavity-free wound healing with microenvironment remodeling to promote nerve regrowth has rarely been explored. A one-step solution is proposed through a self-assembling, multifunctional hydrogel depot that punctually releases the anti-inflammatory drug methylprednisolone sodium succinate (MPSS) and growth factors (GFs) locally according to pathophysiology to repair severe SCI. Synergistically releasing the anti-inflammatory drug MPSS and GFs in the hydrogel depot throughout SCI pathophysiology protects spared tissues/axons from secondary injury, promotes scar boundary- and cavity-free wound healing, and results in permissive bridges for remarkable axonal regrowth. Behavioral and electrophysiological studies indicate that remnants of spared axons, not regenerating axons, mediate functional recovery, strongly suggesting that additional interventions are still required to render the rebuilt neuronal circuits functional. These findings pave the way for the development of a systemic solution to treat acute SCI.

Nonpeptidic quinazolinone derivatives as dual nucleotide-binding oligomerization domain-like receptor 1/2 antagonists for adjuvant cancer chemotherapy.

Nucleotide-binding oligomerization domain-containing protein 1 and 2 (NOD1/2) receptors are potential immune checkpoints. In this article, a quinazolinone derivative (36b) as a NOD1/2 dual antagonist was identified that significantly sensitizes B16 tumor-bearing mice to paclitaxel treatment by inhibiting both nuclear factor κB (NF-κB) and mitogen-activated protein kinase inflammatory signaling that mediated by NOD1/2.

Identification of benzofused five-membered sultams, potent dual NOD1/NOD2 antagonists in vitro and in vivo.

Nucleotide-binding oligomerization domain-containing proteins 1 and 2 play important roles in immune system activation. Recently, a shift has occurred due to the emerging knowledge that preventing nucleotide-binding oligomerization domains (NODs) signaling could facilitate the treatment of some cancers, which warrants the search for dual antagonists of NOD1 and NOD2. Herein, we undertook the synthesis and identification of a new class of derivatives of dual NOD1/NOD2 antagonists with novel benzofused five-membered sultams. Compound 14k was finally demonstrated to be the most potent molecule that inhibits both NOD1-and NOD2-stimulated NF-κB and MAPK signaling in vitro and in vivo.

Discovery of Potent Inhibitors against P-Glycoprotein-Mediated Multidrug Resistance Aided by Late-Stage Functionalization of a 2-(4-(Pyridin-2-yl)phenoxy)pyridine Analogue.

SIS3 is a specific inhibitor of Smad3 that inhibits the TGFß1-induced phosphorylation of Smad3. In this article, a variety of SIS3 derivatives were designed and synthesized to discover potential inhibitors against P-glycoprotein-mediated multidrug resistance aided by late-stage functionalization of a 2-(4-(pyridin-2-yl)phenoxy)pyridine analogue. A novel class of potent P-gp reversal agents were investigated, and a lead compound 37 was identified as a potent P-gp reversal agent with strong bioactivity and outstanding affinity for P-gp.

Promoter Hypomethylation Is Responsible for Upregulated Expression of HAI-1 in Hepatocellular Carcinoma.

An upregulated expression of hepatocyte growth factor activator inhibitor type 1 (HAI-1) in hepatocellular carcinomas (HCC) associates with poor prognosis, but the underlying mechanism for expression regulation has not been elucidated. HAI-1 was expressed in HCC cell line Hep3B cells at a high level but absent or has a low level in other HCC cell lines HepG2 and SMMC7721 and immortal normal liver cell line L02 at transcriptional and translational levels, respectively. A dual-luciferase reporter assay showed that transcriptional activity of HAI-1 in the promoter region (-452 bp to -280 bp from the mRNA start site) was strongly enhanced in Hep3B and SMMC7721. Bisulfite genomic sequencing results of the HAI-1 promoter region showed an inverse correlation between levels of promoter methylation and expression in HCC cells. The expression level of HAI-1 in SMMC7721, HepG2, and L02 cells was elevated after 5-Aza-2'-deoxycytidine treatment. Hypomethylation of the HAI-1 promoter region contributed to the elevated HAI-1 expression in HCC tissues. In addition, the hypomethylation of the HAI-1 promoter region correlated with poor differentiation status of HCC tissues. Our findings indicate that promoter hypomethylation is an important mechanism for aberrant HAI-1 expression regulation in HCC.

[Downregulation of proteinase activated receptor 4 inhibits migration of SW620 human colorectal cancer cells].

OBJECTIVE: To establish the human colorectal cancer cell model SW620/PAR4D with inducible suppression of proteinase activated receptor 4 (PAR4) expression, and investigate the role PAR4 plays in the proliferation and migration of cancer cells. METHODS: A human colorectal cancer cell line with tetracycline-inducible expression regulatory system, namely SW620/Tet-on, was established; inducible expression lentiviral vector with artificial microRNA targeting PAR4, pLVX-Tight-Puro-PAR4-miR, was constructed and transfected into SW620/Tet-on to make an inducible PAR4-suppressed cell model SW620/PAR4D. Western blotting was used to confirm the suppression of PAR4 expression after the doxycycline (DOX) treatment. CCK-8 assay was used to evaluate the impact of suppressed PAR4 expression on cell proliferation, and wound-healing assay was used to analyze the migration of the cells. RESULTS: The SW620/PAR4D cell model was established successfully. Suppression of PAR4 expression by DOX treatment had no significant impact on the growth/proliferation of SW620/PAR4D cells, but markedly inhibited the cell migration. CONCLUSION: Suppression of PAR4 expression has no significant effect on the proliferation of SW620 cells, but can inhibit the migration of the cells.

Nuclear distribution of eIF3g and its interacting nuclear proteins in breast cancer cells.

Eukaryotic translation initiation factor 3 subunit g (eIF3g) is a core subunit of the eukaryotic translation initiation factor 3 complex, and is important in the initiation of translation. It is also involved in caspase-mediated apoptosis, and is upregulated in multidrug-resistant cancer cells. In the present study, the nuclear distribution of eIF3g was determined by performing co-immunoprecipitation of proteins that potentially interact with eIF3g in the nucleus. Mass spectrometry characterization showed that three proteins, heterogeneous nuclear ribonucleoprotein U/scaffold attachment factor A, HSZFP36/zinc finger protein 823 and ß­actin, were among the candidate eIF3g­interacting proteins in the nucleus. The protein­protein interaction was further confirmed by cross­linking and a glutathione S­transferase pull­down assay, followed by western blotting. The co­localization of these proteins was determined by confocal microscopy. These findings provide novel insight into the possible functions of eIF3g in the nucleus and serves as an important first step for further investigation of the roles of eIF3g in cancer development.

Fetal liver cell-containing hybrid organoids improve cell viability and albumin production upon transplantation.

Cell transplantation is a potential alternative for orthotopic liver transplantation because of the chronic donor shortage. Functional liver tissue is needed for cell transplantations. However, large functional liver tissue is difficult to construct because of the high oxygen consumption of hepatocytes. In our previous study, we developed a novel method to generate hybrid organoids. In this study, we used fetal liver cells (FLCs) to construct a hybrid organoid. Nucleus numbers, angiogenesis, and albumin production were measured in transplanted samples. Higher cell viability and larger liver tissue was found in FLC-containing samples than in hepatocyte-containing samples. Furthermore, the therapeutic efficiency of FLC-containing samples was evaluated by transplantation into Nagase analbuminemia rats. As a result, an increase in albumin concentration was found in rat blood. In summary, transplantation of a FLC-containing hybrid organoid is a potential approach for cell transplantation.

Temporal trends and risk assessment of polychlorinated biphenyls and heavy metals in a solid waste site in Taizhou, China.

The solid wastes generated during the production of chemicals are important sources of polychlorinated biphenyls (PCBs) and heavy metals. However, few studies have been conducted regarding long-term monitoring of the risks and states of PCBs and heavy metal pollution from these sources. Herein, we reported the concentrations and risks posed by these pollutants at a chemical solid waste storage site in Taizhou, China, based on data collected before (in 2006) and after clearing the solid waste (in 2013). We examined the concentrations of 24 PCBs, including ten dioxin-like-PCB (DL-PCB) congeners (PCB77, 105, 114, 118, 123, 126, 156, 167, 169, and 189). Our data showed that the mean ∑24PCB concentrations in the soil, water, and plant samples were 6902.90 ng/g, 1637.58 ng/L, and 33.95 ng/g, respectively, in 2013. Furthermore, Cr was the most prevalent contaminant. The hazard quotient (HQ) values showed that Pb posed the highest risk in the soil samples, followed by Hg. The results of the reporter gene assay showed that soil extracts from S1, S2, S3, S4, S5, S6, and S9 exhibited potential estrogenic activities. A comparison of the data showed that the PCB pollution in some sites of this area was still serious. The data provided here are fundamentally useful for policy makers to regulate this type of storage site.

Hybrid organoids consisting of extracellular matrix gel particles and hepatocytes for transplantation.

Hepatocyte transplantation is a potential therapy for treating various liver diseases. However, oxygen shortage leading to loss of hepatocyte function becomes a limitation following hepatocyte transplantation. To overcome this problem, we developed a hybrid organoid, consisting of growth factor (GF)-immobilizable gel particles combined with hepatocytes. Benefits of the hybrid organoid were evaluated in three groups: (i) hybrid organoid consisting of cells and GF-immobilizable gel particles (HG-C); (ii) hybrid organoid consisting of cells and gel particles (G-C); and (iii) cells suspended in collagen (C-C). We found liver-specific functions of HG-C were maintained longer than in the other conditions during in vitro culture. Furthermore, after transplantation, HG-C was effective in maintaining viability of transplanted hepatocytes and promoting angiogenesis around the hepatocytes. In summary, transplantation of HG-C is a potential method for future liver tissue engineering.

Cell-specific expression of artificial microRNAs targeting essential genes exhibit potent antitumor effect on hepatocellular carcinoma cells.

To achieve specific and potent antitumor effect of hepatocyte carcinoma cells, replication defective adenoviral vectors, namely rAd/AFP-amiRG, rAd/AFP-amiRE and rAd/AFP-amiRP, were constructed which were armed with artificial microRNAs (amiRs) targeting essential functional genes glyceraldehyde-3-phosphate dehydrogenase, eukaryotic translation initiation factor 4E and DNA polymerase α respectively under the control of a recombinant promoter comprised of human α-fetoprotein enhancer and basal promoter. The AFP enhancer/promoter showed specific high transcription activity in AFP-positive HCC cells Hep3B, HepG2 and SMMC7721, while low in AFP-negative cell Bcap37. All artificial microRNAs exhibited efficient knockdown of target genes. Decreased ATP production and protein synthesis was observed in rAd/AFP-amiRG and rAd/AFP-amiRE treated HCC cells. All three recombinant adenoviruses showed efficient blockage of cell cycle progression and significant suppression of HCC cells in vitro. In nude mice model bearing Hep3B xenograft, administration of rAd/AFP-amiRG showed potent antitumor effect. The strategy of tumor-specific knockdown of genes essential for cell survival and proliferation may suggest a novel promising approach for HCC gene therapy.

Hepatocyte growth factor activator inhibitor type­1 in cancer: advances and perspectives (Review).

Cancer is one of the most common diseases, with high morbidity and mortality rates. Large­scale efforts have been made to understand the pathogenesis of the disease, particularly in the advanced stages, in order to develop effective therapeutic approaches. Hepatocyte growth factor activator inhibitor type-1 (HAI-1), also known as serine protease inhibitor Kunitz type 1, inhibits the activity of several trypsin-like serine proteases. In particular, HAI-1 suppresses hepatocyte growth factor (HGF) activator and matriptase, resulting in subsequent inhibition of HGF/scatter factor and macrophage­stimulating protein (MSP). HGF and MSP are involved in cancer development and progression, via the receptors Met receptor tyrosine kinase (RTK) and Ron RTK, respectively. Therefore, HAI-1-mediated downregulation of HGF and MSP signaling may suppress tumorigenesis and progression in certain types of cancers. Abnormal HAI-1 expression levels have been observed in various types of human cancer. The exact function of HAI-1 in cancer pathogenesis, however, has not been fully elucidated. In this review, the focus is on the potential impact of aberrant HAI-1 expression levels on tumorigenesis and progression, the underlying mechanisms, and areas that require further investigation to clarify the precise role of HAI-1 in cancer.