The novel potential therapeutic target PSMP/MSMP promotes acute kidney injury via CCR2.

Acute kidney injury (AKI) is a major worldwide health concern that currently lacks effective medical treatments. PSMP is a damage-induced chemotactic cytokine that acts as a ligand of CCR2 and has an unknown role in AKI. We have observed a significant increase in PSMP levels in the renal tissue, urine, and plasma of patients with AKI. PSMP deficiency improved kidney function and decreased tubular damage and inflammation in AKI mouse models induced by kidney ischemia-reperfusion injury, glycerol, and cisplatin. Single-cell RNA sequencing analysis revealed that Ly6Chi or F4/80lo infiltrated macrophages (IMs) were a major group of proinflammatory macrophages with strong CCR2 expression in AKI. We observed that PSMP deficiency decreased CCR2+Ly6Chi or F4/80lo IMs and inhibited M1 polarization in the AKI mouse model. Moreover, overexpressed human PSMP in the mouse kidney could reverse the attenuation of kidney injury in a CCR2-dependent manner, and this effect could be achieved without CCL2 involvement. Extracellular PSMP played a crucial role, and treatment with a PSMP-neutralizing antibody significantly reduced kidney injury in vivo. Therefore, PSMP might be a therapeutic target for AKI, and its antibody is a promising therapeutic drug for the treatment of AKI.

CCR2 is a potential therapeutic target in peri-implantitis.

AIM: CCR2 (C-C chemokine receptor type 2) plays a crucial role in inflammatory and bone metabolic diseases; however, its role in peri-implantitis remains unclear. This study aimed to explore whether CCR2 contributes to peri-implantitis and the treatment effects of cenicriviroc (CVC) on peri-implant inflammation and bone resorption. MATERIALS AND METHODS: The expression of CCR2 was studied using clinical tissue analysis and an in vivo peri-implantitis model. The role of CCR2 in promoting inflammation and bone resorption in peri-implantitis was evaluated in Ccr2-/- mice and wild-type mice. The effect of CVC on peri-implantitis was evaluated using systemic and local dosage forms. RESULTS: Human peri-implantitis tissues showed increased CCR2 and CCL2 levels, which were positively correlated with bone loss around the implants. Knocking out Ccr2 in an experimental model of peri-implantitis resulted in decreased monocyte and macrophage infiltration, reduced pro-inflammatory cytokine generation and impaired osteoclast activity, leading to reduced inflammation and bone loss around the implants. Treatment with CVC ameliorated bone loss in experimental peri-implantitis. CONCLUSIONS: CCR2 may be a potential target for peri-implantitis treatment by harnessing the immune-inflammatory response to modulate the local inflammation and osteoclast activity.

CCL2 is a key regulator and therapeutic target for periodontitis.

AIM: Our previous study revealed that the C-C motif chemokine receptor 2 (CCR2) is a promising target for periodontitis prevention and treatment. However, CCR2 is a receptor with multiple C-C motif chemokine ligands (CCLs), including CCL2, CCL7, CCL8, CCL13 and CCL16, and which of these ligands plays a key role in periodontitis remains unclear. The aim of the present study was to explore the key functional ligand of CCR2 in periodontitis and to evaluate the potential of the functional ligand as a therapeutic target for periodontitis. MATERIALS AND METHODS: The expression levels and clinical relevance of CCR2, CCL2, CCL7, CCL8, CCL13 and CCL16 were studied using human samples. The role of CCL2 in periodontitis was evaluated by using CCL2 knockout mice and overexpressing CCL2 in the periodontium. The effect of local administration of bindarit in periodontitis was evaluated by preventive and therapeutic medication in a mouse periodontitis model. Microcomputed tomography, haematoxylin and eosin staining, tartrate-resistant acid phosphatase staining, real-time quantitative polymerase chain reaction, enzyme-linked immunosorbent assay, bead-based immunoassays and flow cytometry were used for histomorphology, molecular biology and cytology analysis. RESULTS: Among different ligands of CCR2, only CCL2 was significantly up-regulated in periodontitis gingival tissues and was positively correlated with the severity of periodontitis. Mice lacking CCL2 showed milder inflammation and less bone resorption than wild-type mice, which was accompanied by a reduction in monocyte/macrophage recruitment. Adeno-associated virus-2 vectors overexpressing CCL2 in Ccl2-/- mice gingiva reversed the attenuation of periodontitis in a CCR2-dependent manner. In ligation-induced experimental periodontitis, preventive or therapeutic administration of bindarit, a CCL2 synthesis inhibitor, significantly inhibited the production of CCL2, decreased the osteoclast number and bone loss and reduced the expression levels of proinflammatory cytokines TNF-α, IL-6 and IL-1ß. CONCLUSIONS: CCL2 is a pivotal chemokine that binds to CCR2 during the progression of periodontitis, and targeting CCL2 may be a feasible option for controlling periodontitis.

Critical roles for CCR2 and the therapeutic potential of cenicriviroc in periodontitis: A pre-clinical study.

AIM: CCR2 plays important roles in many inflammatory and bone metabolic diseases, but its specific role in periodontitis is unknown. In the present study, we aimed to explore the role of CCR2 in the progression of periodontitis and evaluate the effect of cenicriviroc (CVC) on periodontitis. MATERIALS AND METHODS: The expression of CCR2 was studied in patients with periodontitis and in ligation-induced murine model of periodontitis. The role of CCR2 in promoting inflammation and bone resorption in periodontitis was evaluated in Ccr2-/- mice and wild-type mice. The effect of CVC in the prevention and treatment of periodontitis was evaluated by systemic and local medication. Microcomputed tomography, haematoxylin and eosin staining, tartrate-resistant acid phosphatase staining, quantitative real-time polymerase chain reaction, enzyme-linked immunosorbent assay, and flow cytometry were used for histomorphology, molecular biology, and cytology analysis, respectively. RESULTS: In this study, we demonstrated that CCR2 was highly expressed in human and murine periodontitis and that CCR2 deficiency was associated with decreased inflammatory monocyte and macrophage infiltration and inflammatory mediators, osteoclast number and alveolar bone resorption. Prevention and treatment with CVC significantly reduced the severity of periodontitis, regardless of whether it was administered systemically or locally. CONCLUSIONS: CCR2 plays an important role in the development and progression of periodontitis, and CVC is a potential drug for the prevention and treatment of periodontitis.

In vitro detection of circulating tumor cells using the nicking endonuclease-assisted lanthanide metal luminescence amplification strategy.

The in vitro detection of circulating tumor cells (CTCs) has been proven as a vital method for early diagnosis and evaluation of cancer metastasis, since the existence and number fluctuation of CTCs have shown close correlation with clinical outcomes. However, it remains difficult and technically challenging to realize accurate CTCs detection, due to the rarity of CTCs in the blood samples with complex components. Herein, we reported a CTCs in vitro detection strategy, utilizing a loop amplification strategy based on DNA tetrahedron and nicking endonuclease reaction, as well as the anti-background interference based on lanthanide metal luminescence strategy. In this work, a detection system (ATDN-MLLPs) composed of an aptamer-functionalized tetrahedral DNA nanostructure (ATDN) and magnetic lanthanide luminescent particles (MLLPs) was developed. ATDN targeted the tumor cells via aptamer-antigen recognition and extended three hybridizable target DNA segments from the apex of a DNA tetrahedron to pair with probe DNA on MLLPs. Then, the nicking endonuclease (Nt.BbvCI) recognized the formed double-strand DNA and nicked the probe DNA to release the target DNA for recycling, and the released TbNps served as a high signal-to-noise ratio fluorescence signal source for CTCs detection. With a detection limit of 5 cells/mL, CTCs were selectively screened throughout a linear response range of low orders of magnitude. In addition, the ATDN-MLLPs system was attempted to detect possible existence of CTCs in biological samples in vitro.

The involvement of epidural fat in ossification of the ligamentum flavum: From the perspective of exosomal proteome.

Ossification of the ligamentum flavum (OLF) is the primary etiology of thoracic spinal stenosis. The functional properties of epidural fat (EF), an adipose tissue located in close proximity to ligamentum flavum (LF), have been scarcely investigated. The metabolic state of adipocytes significantly influences their functionality, and exosomes play a pivotal role in intercellular communication. This study aimed to investigate the role of EF-derived exosomes in OLF and characterize their protein profile by proteomics analysis. Our findings demonstrate that exosomes obtained from EF adjacent to OLF possess the ability to enhance osteogenesis of fibroblasts in vitro. Furthermore, proteomics analysis revealed metabolic dysfunction in EF adipocytes and identified lactate dehydrogenase A (LDHA) as a potential mediator involved in the development of OLF. This study provides new insights into the pathogenic mechanism underlying OLF and offers a theoretical basis for preventing and treating ligament ossification.

Magnetic lanthanide sensor with self-ratiometric time-resolved luminescence for accurate detection of epithelial cancerous exosomes.

Fluorescence-based LB (liquid biopsy) offers a rapid means of detecting cancer non-invasively. However, the widespread issue of sample loss during purification steps will diminish the accuracy of detection results. Therefore, in this study, we introduce a magnetic lanthanide sensor (MLS) designed for sensitive detection of the characteristic protein, epithelial cell adhesion molecule (EpCAM), on epithelial tumor exosomes. By leveraging the inherent multi-peak emission and time-resolved properties of the sole-component lanthanide element, combined with the self-ratiometric strategy, MLS can overcome limitations imposed by manual operation and/or sample complexity, thereby providing more stable and reliable output results. Specifically, terbium-doped NaYF4 nanoparticles (NaYF4:Tb) and deformable aptamers terminated with BHQ1 were sequentially introduced onto superparamagnetic silica-decorated Fe3O4 nanoparticles. Prior to target binding, emission from NaYF4:Tb at 543 nm was partially quenched due to the fluorescence resonance energy transfer (FRET) from NaYF4:Tb to BHQ1. Upon target binding, changes in the secondary structure of aptamers led to the fluorescence intensity increasing since the deconfinement of distance-dependent FRET effect. The characteristic emission of NaYF4:Tb at 543 nm was then utilized as the detection signal (I1), while the less changed emission at 583 nm served as the reference signal (I2), further reporting the self-ratiometric values of I1 and I2 (I1/I2) to illustrate the epithelial cancerous features of exosomes while ignoring possible sample loss. Consequently, over a wide range of exosome concentrations (2.28 × 102-2.28 × 108 particles per mL), the I1/I2 ratio exhibited a linear increase with exosome concentration [Y(I1/I2) = 0.166 lg (Nexosomes) + 3.0269, R2 = 0.9915], achieving a theoretical detection limit as low as 24 particles per mL. Additionally, MLS effectively distinguished epithelial cancer samples from healthy samples, showcasing significant potential for clinical diagnosis.

Accurate and noninvasive diagnosis of epithelial cancers through AND gate photoluminescence on tumor-derived small extracellular vesicles.

Noninvasive detection of small extracellular vesicles (sEVs) has become one of the most promising liquid biopsy methodologies for effective and timely cancer diagnosis and prognostic monitoring. Currently, accurate and sensitive detection of tumor-derived sEVs is compromised by their heterogeneous nature, and the tissue origin and parent cell cycle change may significantly affect the tumor-associated information (e.g., phenotypic proteins) of sEVs. Accordingly, many of the single-marker dependent detections on sEVs may not provide comprehensive information about the tumor, and their reliability and clinical applicability cannot be guaranteed. Herein, a strategy for constructing AND gate photoluminescence on tumor-derived sEVs is proposed. Briefly, only after co-recognition of the two epithelial phenotypic proteins (EpCAM and MUC1) on tumor-derived sEVs simultaneously, can our designed lanthanide luminescence probe precursors then assemble to form the AND gate for photoluminescence detection. Consequently, the generated AND gate photoluminescence provided time-resolved luminescence for a wide cancerous sEV linear detection range of 6.0 × 104-6.0 × 109 particles per mL, with a calculated detection limitation of 1.42 × 102 particles per mL. Furthermore, the AND gate photoluminescence can significantly distinguish epithelial cancer patients from healthy controls, displaying its great potential for accurate and noninvasive cancer diagnosis.

Floating Immunomagnetic Microspheres for Highly Efficient Circulating Tumor Cell Isolation under Facile Magnetic Manipulation.

Among circulating tumor cell enrichment strategies, immunomagnetic beads (IMBs) have received great attention due to their excellent performance. However, traditional strategies using IMBs normally require an additional mechanical stirring device to fully mix the IMBs and specimens, and this step may cause mechanical cellular damage. In this study, by changing the architecture and motion trajectory control strategy of the IMBs, floating immunomagnetic microspheres (FIMMs) and their matching rotary magnetic manipulation device were proposed to achieve highly efficient CTC capture under a cell-friendly condition. Generally, the FIMMs were prepared through layer-by-layer assembly of the individual functional components, and their stress state governed by either buoyancy or magnetic force was tuned by the rotary magnetic manipulation device. Consequently, recognition of FIMMs and target cells as well as CTC recovery can be simply realized through external magnetic manipulation. Accordingly, satisfactory enrichment efficiencies for CTCs with varied epithelial expression levels were obtained as 92.93 ± 3.23% for MCF-7, 79.93 ± 3.31% for A549, and 92.57 ± 5.22% for HepG2. Besides, an extremely low detection limitation of 5 cells mL-1 can be achieved from complex sample conditions, even the whole blood. In addition, FIMMs successfully enriched 23-56 CTCs from 1.5 mL of blood samples from cancer patients.

The Long Noncoding RNA LINC00963 Inhibits Corneal Fibrosis Scar Formation by Targeting miR-143-3p.

Corneal fibrosis is a complication of severe corneal injury, one of the major causes of vision loss. The formation of myofibroblasts has emerged as a key stimulative factor of corneal fibrosis. In the current study, we focused on the role of LINC00963 in regulating corneal fibrosis. Transforming growth factor ß1 (TGF-ß1) was used to induce human corneal stromal cells differentiating into corneal myofibroblasts, and the significant increase of α-smooth muscle actin (α-SMA) was verified by quantitative real-time PCR (qRT-PCR), western blot, and immunofluorescence, respectively. LINC00963 was identified to be one-half decreased compared with nonstimulated human corneal stromal cells, indicating that it might play a role in corneal fibrosis. Interestingly, overexpression of LINC00963 resulted in decreased formation of myofibroblasts indicating that it might exhibit an inhibiting effect. Moreover, bioinformatics tool was applied to predict the downstream target of LINC00963. We investigated that LINC00963 suppressed α-SMA induced by TGF-ß1 in corneal fibroblasts, at least in part, by downregulating the expression of miR-143-3p. In addition, either LINC00963 promotion or miR-143-3p inhibition could significantly decrease myofibroblast contractility and collagen I and III secretion, which are the key to contribute to corneal fibrosis. Taken together, our study identified LINC00963 as a promising therapeutic target.

A circular RNA from NFIX facilitates oxidative stress-induced H9c2 cells apoptosis.

Myocardial infarction is the leading cause of death worldwide, and cardiomyocyte apoptosis during myocardial infarction and reperfusion is a significant factor of poor prognosis. As important regulatory molecules, biofunctions of circRNAs in the pathogenesis of myocardial infarction remain elusive. To confirm the expression level and biological function of circNFIX in cardiomyocytes upon oxidative stress. Divergent polymerase chain reaction and Sanger sequencing were performed to verify the circular structure. The stability of circNFIX was confirmed by RNase R treatment and actinomycin D assay. In order to simulate oxidative stress during myocardial infarction, H9c2 cells were subjected to hydrogen peroxide and hypoxia stimulation. In vivo, mouse models of myocardial ischemia were established. The biological function of circNFIX in cardiomyocytes was investigated through loss- and gain-of-function assays, and cardiomyocyte apoptosis level was detected by the terminal deoxyribonucleotidyl transferase-mediated TdT-mediated dUTP nick end labeling assay and Western blot. CircNFIX is abundant, conserved, and stable in H9c2 cells. The expression of circNFIX was significantly downregulated in cardiomyocytes subjected to oxidative stress. Enforced CircNFIX promotes H9c2 cells apoptosis induced by hydrogen peroxide, in sharp contrast to circNFIX knockdown. In this study, we found that circNFIX served as a pro-apoptosis factor in cardiomyocyte apoptosis. CircNFIX possesses potential to be the biomarker and therapeutic target in myocardial infarction.

Analysis of circular RNA-associated competing endogenous RNA network in breast cancer.

As the most common type of cancer in female patients, the morbidity and mortality rates of breast cancer (BC) are high, and its incidence is gradually increasing worldwide. However, the underlying molecular and genetic mechanisms involved in the etiopathogenesis of BC remain unclear. Circular RNAs (circRNAs) are a novel type of non-coding RNAs that have been verified to serve a crucial role in tumorigenesis. However, the majority of functions and mechanisms of circRNAs remain unknown. The present study identified 47 differentially expressed circRNAs in a dataset from Gene Expression Omnibus. Using the cancer-specific circRNA database, the potential microRNA (miRNA) response elements, RNA-binding proteins and open reading frames of the candidate circRNAs were predicted. Combing the predictions of miRNAs and target mRNAs, a competing endogenous RNA network was constructed, which may serve as the theoretical basis for further research. Furthermore, the analyses conducted using Gene Ontology terms and Kyoto Encyclopedia of Genes and Genomes pathways indicated that candidate circRNAs may serve a role in transcriptional regulation. Moreover, 20 BC tissue specimens and their paired adjacent normal tissue specimens were used to evaluate the expression levels of the screened circRNAs. Thus, the analyses of the raw microarray data conducted in the present study offer perspectives on the exploration of mechanisms associated with BC tumorigenesis with regard to the circRNA-miRNA-mRNA network.