Inflammation and regeneration最新文献

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease characterized by progressive degeneration and loss of upper and lower motor neurons, with approximately 90% of cases being sporadic (sporadic ALS, SALS). A reliable diagnostic biomarker remains an unmet clinical need in SALS, with misdiagnosis and diagnostic delay hindering early management. The mislocalization of the RNA-binding protein TDP-43 (encoded by TARDBP), a pathological hallmark of SALS, could lead to aberrant splicing that produces transcripts with cryptic exons and, consequently, cryptic peptides. This study proposes cryptic peptides in serum extracellular vesicles as a novel candidate diagnostic biomarker of SALS. We included 10 healthy controls and 20 patients with SALS and quantified cryptic peptides predicted from cryptic exon sequences using mass spectrometry-based proteomics. Cryptic peptides from four proteins (RANBP1, IGLON5, ACTN1, ALPK2) were detected in participants, with the IGLON5 cryptic peptide detected significantly more frequently in SALS than in HC (adjusted P = 0.044). The number of detected cryptic peptides classified SALS and healthy controls with acceptable performance (area under the curve = 0.82). In conclusion, cryptic peptides could have diagnostic performance for SALS, warranting further validation.

Background: In tooth development, the Wnt/β-catenin pathway has been shown to play a crucial role in tooth germ formation and tooth differentiation. Regeneration can be considered a replay of development. Understanding and reproducing the involvement of Wnt10a-which is reported to be the causative gene for congenital multiple tooth absence and participates in tooth development with spatiotemporal specificity-in pulp regeneration is essential for realizing dental regenerative medicine. Therefore, this study was initiated based on the concept that evaluating the dynamics of Wnt10a during angiogenesis, which is thought to occur early in dental pulp regeneration, could contribute to a more detailed elucidation of the dental pulp regeneration mechanism.

Methods: Deciduous dental pulp stem cells (SHED) were isolated from human deciduous teeth, and conditioned medium (CM) was collected. In addition, after the induction of vascular differentiation of SHED, the temporal gene expression of Wnt10a, VEGF-A, Tie-2, and β-catenin was analyzed by q-PCR and protein expression by Western blotting and ELISA from 0 to 48 h and 3, 7, 14, and 21 days after the induction.

Results: Canonical Wnt signaling was activated during angiogenesis in regenerated pulp-like tissue induced by ectopic root grafting, and Wnt10a had spatio-temporal specificity. Tie-2 activation occurred during the process of induction of vascular differentiation in SHED.

Conclusions: During angiogenesis in pulp regeneration, when SHED differentiate into blood vessels, Canonical Wnt signaling and VEGF-A are activated to form microvessels, and Tie-2 expression is enhanced to increase vessel circumference. Furthermore, Wnt10a was found to be activated in its early stages and decreased in its mature stages, with spatio-temporal specificity. Because its expression is very low, Wnt10a could be a biomarker to monitor the regenerative status of pulp regeneration.

Background: Otitis media (OM) is an inflammatory disease of the middle ear characterized by mucosal remodeling, effusion, and conductive hearing loss. Although antibiotics and surgical procedures remain standard treatments, their efficacy is often limited by recurrence, antibiotic resistance, and chronic progression. Transcutaneous auricular vagus nerve stimulation (taVNS) is a non-invasive neuromodulatory technique that activates the cholinergic anti-inflammatory pathway through α7 nicotinic acetylcholine receptor (α7nAChR) signaling. This study aimed to evaluate the anti-inflammatory effects of taVNS in a lipopolysaccharide (LPS)-induced mouse model of acute otitis media (AOM) and to assess whether these effects may involve α7nAChR-dependent mechanisms.

Methods: AOM was induced by transtympanic injection of LPS in BALB/c mice, followed by taVNS applied to the auricular concha using biphasic square pulses (0.3 mA, 20 Hz, 200 μs). Methyllycaconitine, a selective α7nAChR antagonist, was administered to assess receptor involvement. Auditory function, tympanic membrane morphology, and mucosal changes were assessed through auditory brainstem response testing, otoscopic imaging, and histological analysis. Cytokine levels in middle ear effusion and serum were quantified by ELISA, while inflammatory gene and protein expression were analyzed using qPCR and Western blotting. Statistical analyses were performed using one-way ANOVA or Kruskal-Wallis tests with post hoc comparisons.

Results: taVNS significantly improved hearing thresholds and reduced mucosal thickening and goblet cell hyperplasia in AOM mice. It markedly decreased tumor necrosis factor-α, interleukin (IL)-1β, and IL-6 levels in middle ear effusion, and mechanistically, this anti-inflammatory effect was associated with suppression of NF-κB activation without altering TLR4 or MYD88 expression. These effects were abolished by methyllycaconitine pretreatment. taVNS also reduced spleen enlargement and systemic cytokine concentrations, indicating modulation of both local and systemic inflammation.

Conclusions: taVNS was associated with attenuation of LPS-induced acute otitis media and reduced NF-κB activation and downstream cytokine expression in a manner consistent with the involvement of α7nAChR-related signaling. By attenuating excessive inflammatory responses, taVNS was associated with improved auditory function and reduced middle ear injury, suggesting its potential as a non-invasive therapeutic strategy for LPS-induced acute otitis media.

Background: Super-enhancers (SEs), characterized by dense clusters of enhancer elements enriched with transcriptional activator binding sites, are involved in cell differentiation. However, little is known about SE-mediated regulation of adipogenic genes. The aim of this study was to elucidate the functional role of the KLF6-proximal SE during the adipogenesis of human adipose-derived stem cells (hADSCs).

Methods: Adipogenic induction medium (AIM) was used for differentiation of hADSCs into adipocytes, which were evaluated for adipogenic gene expression and adipogenesis using quantitative PCR and Oil Red O (ORO) staining, respectively. The effects of SE inhibitors, locked nucleic acid-mediated enhancer RNA (eRNA) knockdown, and small interfering RNA-mediated knockdown of KLF6 on adipogenesis and adipogenic gene levels were evaluated. Chromatin immunoprecipitation assays were performed to identify transcriptional regulators binding to the promoter regions of KLF6 and the adipogenesis-inhibitory Delta-like non-canonical Notch ligand 1 (DLK1) gene during adipogenesis.

Results: In silico screening identified KLF6 as an obesity-susceptibility gene associated with single-nucleotide polymorphisms and located within the domain of SE_00159, which was activated in adipocytes. AIM-cultured hADSCs exhibited time-dependent increases in KLF6 mRNA and protein expression. During adipogenesis, the transcription factor peroxisome proliferator-activated receptor gamma (PPARγ) bound to the KLF6 promoter. Treatment with the SE inhibitor JQ1 resulted in a dose-dependent decrease in KLF6 mRNA expression and reduced ORO staining. Knockdown of eRNA expressed from the SE_00159 domain decreased KLF6 levels during adipogenesis. Consistently, KLF6 knockdown during hADSC adipogenesis downregulated the adipogenic genes PPARG and CEBPA, while upregulating DLK1. Additionally, KLF6 together with histone deacetylase (HDAC)3 bound to the DLK1 promoter and concomitantly caused dissociation of histone acetyltransferase p300 during adipogenesis.

Conclusions: SE activation upregulates KLF6 through PPARγ/p300- and eRNA-mediated transcriptional induction during hADSC adipogenesis. KLF6, in turn, represses DLK1 expression through recruitment of HDAC3 to the promoter and p300 dissociation, thereby facilitating adipocyte differentiation. These findings support a working model in which the epigenetic regulation of KLF6 and DLK1 as a potential therapeutic axis in human obesity.

Two-photon excitation microscopy has become an important technique in dermatologic research, providing high-resolution imaging of living skin and its immune responses. Unlike conventional histology, two-photon microscopy allows direct observation of living tissues in real time at cellular resolution. This method enables the visualization of keratinocyte organization, skin appendages such as hair follicles and sebaceous glands, and the vascular, lymphatic, and neural networks within the dermis. It has also revealed dynamic immune processes in the skin. Clinical application is still limited by safety concerns and the high cost of equipment. Nonetheless, strategies based on autofluorescence, melanin scattering, and second harmonic generation (SHG) have been explored to visualize collagen and fibrosis in the dermis. This review summarizes the structural, functional, and translational aspects of two-photon skin imaging. We outline the technical principles, applications in animal and human studies, and potential implications for dermatologic research and clinical diagnosis.

Inflammatory bowel disease (IBD) encompasses chronic, relapsing inflammatory disorders of the gastrointestinal tract, which are driven by intricate interactions between the host immune system and intestinal microbiota. Recent studies have revealed that microbiota-derived D-amino acids (D-AAs), once considered biologically inert, play critical roles in maintaining mucosal homeostasis and modulating immune responses. These metabolites, which are increasingly classified as postbiotics, directly influence epithelial barrier integrity, immune cell activity, and microbial ecology. In this review, we summarize the current insights into the biosynthesis, bacterial functions, and immunological implications of D-AAs in the gut, with a particular focus on their involvement in IBD pathogenesis. Specific D-AAs, such as D-alanine, contribute to bacterial cell wall integrity and quorum sensing and interact with host immune cells, alter microbial communities, and regulate mucosal barrier function. Evidence from both human studies and murine models highlights how disrupted D-AAs' metabolism through dysbiosis or impaired host sensing via enzymes such as D-amino acid oxidase (DAO) exacerbates inflammation. Finally, we discuss the translational potential of D-AAs as non-invasive biomarkers and therapeutic targets in IBD, emphasizing the need for integrative multi-omics approaches that connect microbial metabolism with host immune regulation and disease outcomes.

Background: Induced pluripotent stem cell (iPSC)-derived T cells offer a renewable source for off-the-shelf immunotherapy. With the advent of the artificial thymic organoid (ATO) method, the in vitro differentiation of CD4⁺ T cells from iPSCs has also become feasible. CD4⁺ T cells have shown superior longevity, resistance to exhaustion, and helper functions in primary settings, but whether iPSC-derived CD4⁺ T cells retain these features remains unclear.

Methods: In this study, CD4⁺ T cells were differentiated from human iPSCs using the ATO system. Primary T cells served as controls to evaluate the phenotypic and activation features of iPSC-derived CD4⁺ and CD8⁺ T cells. To assess antitumor function, we generated CD19-BBζ CAR-iPSC-T cells and employed a hematologic malignancy model using NALM6 acute lymphoblastic leukemia (ALL) cells. Both short-term and long-term cytotoxicity assays were conducted to compare iPSC-derived CD4⁺ and CD8⁺ T cells in terms of killing efficiency, cytokine secretion, persistence, exhaustion phenotype, and proliferative capacity. The helper function of iPSC-derived CD4⁺ T cells toward CD8⁺ T cells was further evaluated by Ki-67 staining and proliferation assays. Statistical analyses were performed using GraphPad Prism.

Results: Our study demonstrated that iPSC-derived CD4⁺ T cells exhibited both helper- and cytotoxic-like features. Compared with iPSC-derived CD8⁺ T cells or CD4⁺/CD8⁺ mixtures, iPSC-derived CD4⁺ T cells showed superior proliferation, cytokine secretion, and sustained cytotoxicity following CAR transduction. They also promoted the expansion of iPSC-derived CD8⁺ T cells and displayed helper-like functions with increased resistance to exhaustion.

Conclusions: Although not identical to primary CD4⁺ T cells, iPSC-derived CD4⁺ T cells recapitulated key functional advantages, especially sustained antitumor activity, supporting their value as a renewable, off-the-shelf source for next-generation CAR-T therapies.

High-throughput single-cell analysis and screening have become essential tools in life science research. Imaging flow cytometry, in particular, enables large-scale image-based profiling of heterogeneous cell populations, allowing statistical analysis of cellular morphology, subcellular features, and functional responses. However, its analytical capability is often limited by the use of conventional two-dimensional (2D) image sensors. In this review, we highlight recent advances in single-pixel imaging flow cytometry, which replaces 2D image sensors with single-pixel photodetectors. This approach offers advantages in sensitivity, flexibility, and speed in imaging system design and has been implemented in various optical configurations to achieve high-throughput single-cell imaging. We first introduce its key techniques, then outline representative biomedical applications, including cancer and COVID-19 research, and finally discuss current limitations and prospects for future developments. Single-pixel imaging flow cytometry is expected to serve as a versatile platform supporting both basic and translational studies in diverse biomedical applications.

Bond-selective vibrational imaging techniques, such as Raman spectroscopy, are opening up many applications that were previously considered impossible or inaccessible by other means, such as fluorescence imaging. In particular, vibrational microscopy offers unique advantages, such as the ability to perform highly multiplexed, label-free imaging. Indeed, recent advances in optical and chemical technologies have made it possible to image biological phenomena at the cellular level with high sensitivity, high resolution, and high specificity. Applications of vibrational microscopy both in biological research and in medicine, including the detection of pathological lesions, are expanding rapidly. Here, we provide a general overview of Raman microscopy, and we review recent progress in cutting-edge applications, including label-free imaging and the development of small Raman tags, Raman probes enabling highly sensitive ultra-multiplexed observation, and functional Raman probes.

Background: Anti-survival of motor neuron (SMN) antibodies have recently been identified in rheumatic and musculoskeletal diseases (RMDs), notably mixed connective tissue disease (MCTD). However, their immunological characteristics, prevalence, and clinical relevance beyond MCTD remain poorly understood. This study aimed to elucidate the clinical significance of anti-SMN antibodies in a wide spectrum of RMDs.

Methods: We assessed anti-SMN antibodies and antibody-producing cells using Western blotting and immunofluorescence staining with recombinant SMN complexes. Serum anti-SMN antibody titers were measured using a recombinant SMN complex-bound magnetic bead assay in 906 serum samples from patients with 16 types of RMDs and healthy controls. Clinical associations and treatment responses were analyzed.

Results: Western blotting using patients' sera confirmed SMN-specific antibodies. Immunofluorescence staining identified anti-SMN antibody-producing plasma cells in an MCTD patient's lymph node. Anti-SMN antibodies were detected in 36.7% of MCTD, 10.6% of systemic lupus erythematosus (SLE), and 2.4% of systemic sclerosis patients, while none of the healthy controls were positive. Antibody titers were higher against the SMN complex than individual components, highlighting the importance of the complex structure. In MCTD, antibody positivity was strongly correlated with interstitial lung disease (90.9% vs. 36.8%, P = .013). In SLE, antibody-positive patients had significantly lower white blood cell counts and complement levels, higher anti-ds-DNA antibody titer, and higher prevalence of serositis (35.0% vs. 11.3%), gastrointestinal involvement (15.0% vs. 2.4%), nephritis (70.0% vs. 30.4%), and higher median SLE Disease Activity Index scores (19.5 vs. 5.0) compared to antibody-negative patients (all P < .05). Antibody titers decreased after treatment (- 71.5% at 3 months, P = .010) and increased upon relapse.

Conclusions: Anti-SMN antibodies are prevalent in MCTD and SLE. Consistent with prior studies, their titers in MCTD were associated with distinct clinical features. Importantly, we newly demonstrate that in SLE, anti-SMN antibody levels correlate with immune complex-related manifestations and disease activity, providing a novel and clinically significant insight. These findings highlight their potential as biomarkers for disease stratification, organ involvement, and monitoring disease progression in RMDs.