Connective Tissue Research最新文献

Purpose: Intervertebral disc degeneration (IDD) is a spinal condition that causes low back pain. Pellino E3 ubiquitin protein ligase 1 (PELI1) expression reportedly correlates with inflammation and cell death. This study aimed to determine its potential role in IDD.

Methods: Cell counting kit-8 assay, 5-ethynyl-2'-deoxyuridine staining, senescence-associated β-galactosidase staining, morphological observation, lactate dehydrogenase (LDH) release assay, quantitative reverse transcriptase polymerase chain reaction, and western blotting were used to examine the effect of PELI1 on tumor necrosis factor alpha (TNF-α)-induced human primary nucleus pulposus cells (hNPCs).

Results: PELI1 was highly expressed in TNF-α-treated hNPCs. TNF-α treatment notably reduced hNPCs viability and proliferation, but enhanced senescence (elevated p16 and p21 expression), extracellular matrix degeneration (reduced collagen II and aggrecan expression and upregulated matrix metallopeptidase-13 and a disintegrin and metalloproteinase with thrombospondin type 1 motifs-5 expression), nucleotide-binding oligomerization domain-like receptor family pyrin domain-containing-3 (NLRP3) inflammasome formation (enhanced NLRP3, apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC), and cleaved caspase 1 expression), pyroptosis (elevated cleaved gasdermin D expression), LDH release, and inflammatory cytokine release (high mobility group box 1, interleukin (IL)-1β, and IL-18). These effects were distinctly reduced by PELI1 silencing but enhanced by its overexpression. Interestingly, the effects triggered by PELI1 silencing were partially reversed by ASC overexpression.

Conclusions: PELI1 May promote IDD progression by expediting nucleus pulposus cell death and participates in the inflammatory response regulated by the NLRP3 inflammasome in nucleus pulposus cells. These suggest PELI1 as a potential therapeutic target for the treatment of IDD.

Introduction: The cervix plays important mechanical roles in pregnancy and regulating the timing of parturition. Dysfunction of the cervix is implicated in disorders of parturition including spontaneous preterm birth, failed induction of labor and post term pregnancies. To address these disorders, it is imperative to first understand the function of the cervix throughout a normal pregnancy. However, current knowledge on the response of the cervix to mechanical fatigue and the underlying microstructural changes throughout a pregnancy is lacking.

Methods: In this study, mechanical fatigue properties were measured at different stages of pregnancy using uniaxial fatigue testing that simulated circumferential hoop stresses in the cervix. Collagen microstructure was quantified using second harmonic generation imaging and three-dimensional orientation analysis.

Results: The stiffness and modulus of the cervix during fatigue testing were dramatically reduced in all stages of pregnancy, and pregnant samples experienced greater peak strain before failure. All mechanical properties recovered postpartum despite persistent changes in cervix size. Microstructural analysis demonstrated increased local collagen alignment in postpartum samples, which may indicate a mechanism that serves to improve material properties after childbirth.

Discussion: Altogether, conclusions from this study enhance our understanding of how properties of the cervix change with pregnancy and lay the foundation for future work investigating how alterations from this healthy function can lead to spontaneous preterm birth and other reproductive complications.

Background: The aim of this study was to assess nerve regeneration in vein grafts filled with olfactory ensheathing cells (OECs) or olfactory stem cells (ONSs) in a 15 mm gap male DA rat sciatic nerve model versus autografts.

Methods: The control group (NG) received a nerve graft reversed and sutured into the 15 mm gap; all of the animals in the five experimental groups received a vein graft to bridge the 15 mm gap filled with extracellular matrix (ECM- MatrigelTM) only (VG); ECM with rat OECs suspended in ECM (VG + rOECs); ECM with human OECs (with Cyclosporin postoperatively to prevent graft rejection) (VG + hONS (c)); ECM only (with Cyclosporin postoperatively as a control for the previous group) (VG (c)); and ECM with rat ONSs within the vein grafts (VG + rONS). After 12 weeks ±4 days, electrophysiological analysis (latency and amplitude) and histological assessment of axon counts (immunohistochemistry with neurofilament [NF] stain) were undertaken.

Results: Group VG + rOECs had the lowest latency results, NG had the highest amplitude results, and groups NG and VG + rOECs had significantly higher axon counts.

Conclusions: The results trended toward the VG + rOECs and NG groups having the most successful electrophysiology results and axon counts. Incorporating OECs into vein grafts may be a viable alternative to nerve grafts for peripheral nerve repair.

Purpose: We probed the roles of SNHG7, miR-146b, PCBP1, and IL-β in the development of osteoarthritis (OA).

Materials and methods: OA models were established using anterior cruciate ligaments, and chondrocytes were obtained from mouse cartilage tissue. Cells were treated with 10 ng/ml Il-1β. RT-qPCR was used to detect the expression of SNHG7, miR-146b, PCBP1, and IL-β in tissues and cells. Safranin-O/Fast Green staining was performed to analyze the cartilage damage in each group of mice.

Results: SNHG7 and PCBP1 expressions were down-regulated, and miR-146b expression was up-regulated in OA tissue and IL-1β-treated chondrocytes compared to normal cartilage tissue and chondrocytes. Forced SNHG7 expression improved cartilage structure, enhanced proliferative viability of chondrocytes, and inhibited apoptosis and IL-1β release in IL-1β-treated chondrocytes in OA mice. In contrast, miR-146b upregulation decreased proliferative viability and promoted apoptosis and IL-1β release in chondrocytes. Rescue assays showed that miR-146b attenuated the protective effects of SNHG7 on apoptosis and inflammation in IL-1β-treated chondrocytes, and activation of PCBP1 expression significantly inhibited the cytotoxic effects of miR-146b. Mechanistically, SNHG7 acted as a competitive endogenous RNA by targeting miR-146b to promote the expression of PCBP1.

Conclusions: This study confirms that SNHG7 inhibits IL-1β-mediated inflammatory responses in chondrocytes via the miR-146b/PCBP1 axis, thereby suppressing IL-1β-induced OA.

Background: Various forms of decellularized extracellular matrix (dECM), including patches, powders, and hydrogels, have been applied to tissue engineering. Due to a broad need for alternatives to dECM, mostly derived from animal sources, human amniotic membrane (AM) and umbilical cord (UC) as disposable birthing materials can be suitable candidates. The present study developed hydrogels from AM and UC hydrogels and compared their physicochemical and biological properties.

Materials and methods: The decellularized and powdered AM and UC tissues were solubilized with pepsin to form pre-gel solutions. The developed hydrogels underwent biological and physicochemical assessments using techniques such as western blot, scanning electron microscopy, immunohistochemistry, and histopathology.

Results: UC hydrogel demonstrated a higher elastic modulus and shorter gelation time. Although the western blot results did not show significant differences in concentration of the main ECM components, specific staining showed a higher content of mucopolysaccharides in UC hydrogel as well as collagen fibers in AM hydrogel. Both hydrogels induced a fibroblast-like morphology in the cytoplasm of mesenchymal stromal cells (MSCs). Both hydrogels are suitable for 3D culture systems and support in vivo myogenic differentiation of MSCs. Finally, the hydrogels were found to be biocompatible in vivo and showed infiltration and colonization by host cells in mice.

Conclusion: This study highlights significant bio-physicochemical variations between human UC and AM hydrogels, emphasizing the need for careful consideration in their application for tissue reconstruction, in vitro culture systems, and cell-delivery techniques.

Purpose: This study aimed to evaluate the early effects of N-acetylcysteine, which has antioxidant, inflame-modulatory, and cytoprotective properties, on tendon healing.

Materials and methods: Thirty-five male Wistar Hannover rats were divided into five groups: first-week treatment (Group 1T), first-week control (Group 1C), third-week treatment (Group 3T), third-week control (Group 3C), and native tendons (Group N). Bilateral Achilles tenotomy was performed on all rats except Group N. After tenotomy, 150 mg/kg N-acetylcysteine was administered daily intraperitoneally to treatment groups, while isotonic saline was given to the control groups. Tendons were evaluated histopathologically, immunohistochemically, and biomechanically after sacrifice in the first and third weeks.

Results: No significant differences were observed in the first week (p > 0.05). Movin and Bonar scores (lower scores reflect improved histologic healing) were significantly lower in Group 3T than in Group 3C (p = 0.002). Collagen type-I/type-III ratios were higher in Group 3T compared to Group 3C (p = 0.001). Fmax (N) values were similar across Group 3T, Group 3C, and Group N (p = 0.772). However, cross-sectional areas (mm²) were significantly smaller in Group 3T than in Group 3C (p = 0.001), with the smallest areas observed in native tendons. Thus, tensile strength (MPa, load per unit area) and toughness (J/10³ mm³, energy absorbed per unit volume) were significantly higher in Group 3T than in Group 3C (p = 0.001).

Conclusion: N-acetylcysteine supplied some improved results on early markers of tendon healing. Although our findings support the potential of NAC as a therapeutic adjunct in tendon injuries, further studies are needed to evaluate the long-term effects and underlying mechanisms.

Background: Methyltransferase-like 3 (METTL3) is implicated in human diseases, including osteoporosis (OP). In this study, we aimed to explore the functions and mechanisms of METTL3 in OP using bone marrow mesenchymal stem cells (BMSCs).

Methods: The identification of BMSCs-derived exosomes was conducted by transmission electron microscope (TEM), Nanoparticle Tracking Analysis (NTA) and western blot. The osteogenic differentiation of osteoblasts (hFOB1.19) was analyzed by Alizarin red staining assay, Alkaline phosphatase (ALP) staining assay and western blot. The relationship between METTL3 and SMAD family member 5 (SMAD5) was analyzed by Methylated RNA Immunoprecipitation (MeRIP) assay and dual-luciferase reporter assay.

Results: BMSCs-derived exosomes (BMSC-Exos) promoted the osteogenic differentiation and elevated METTL3 expression in hFOB1.19 cells. Exosomal METTL3 knockdown repressed the osteogenic differentiation in hFOB1.19 cells. METTL3 could stabilize and regulate SMAD5 expression by N6-methyladenosine (m6A) modification. Moreover, SMAD5 overexpression restored exosomal METTL3 knockdown-mediated effect on the osteogenic differentiation in hFOB1.19 cells.

Conclusion: BMSCs-derived exosomal METTL3 mediated the m6A methylation of SMAD5 to facilitate osteogenic differentiation of hFOB1.19 cells.

Purpose: Orthodontic interventions such as maxillary expansion are pivotal in correcting malocclusions; however, the intracellular mechanisms of bone remodeling during this process are not well understood. This study investigated the role of the mitogen-activated protein kinase (MAPK) pathway in bone remodeling during maxillary expansion and relapse in rats.

Materials and methods: Thirty male Wistar rats were randomly divided into three groups: Control (Ctrl), Expansion only (EO), and Expansion with MEK inhibitor U0126 (EO  +  INH). Customized expanders applied 100 g force for seven days, followed by natural relapse. Tissue changes within the mid-palatal suture were assessed via micro-computed tomography, histology, and immunohistochemistry. In vitro, primary bone marrow mesenchymal stem cells (BMSCs) were exposed to cyclic tensile stress with or without MAPK inhibition, followed by evaluation of protein expression, alkaline phosphatase activity, and Alizarin red staining.

Results: The EO group showed a significant increase in maxillary arch width compared to the EO  +  INH group, a difference that remained significant after relapse. This group also had higher levels of phosphorylated mitogen-extracellular kinase (p-MEK), phosphorylated extracellular signal-regulated kinase 1/2 (p-ERK1/2), and phosphorylated Ets-like transcription factor 1 (p-ELK1), along with increased osteoblast markers and bone resorption. Conversely, MAPK inhibition impeded bone remodeling, indicated by decreased osteogenic markers and fewer TRAP-positive cells. In vitro, tensile stress enhanced osteogenic differentiation, which was attenuated with MAPK inhibition.

Conclusions: Mechanical activation of MEK-ERK1/2-ELK1 pathway is essential for effective maxillary expansion. Thus, inhibiting this pathway significantly impairs bone remodeling, underscoring its potential as a therapeutic target to enhance bone formation in orthodontic treatments.

Background: Steroid-induced osteonecrosis of the femoral head (SONFH) is a metabolic disorder that leads to structural changes, collapse of the femoral head, and joint dysfunction. This study investigates the role of interferon regulatory factor 8 (IRF8) in osteocyte apoptosis in SONFH, so as to find new targets for the treatment of SONFH.

Methods: Murine long bone osteocyte-Y4 cells were cultured and treated with dexamethasone to establish SONFH cell models. si-IRF8 was transfected into the cells. The expression levels of IRF8, B cell leukemia/lymphoma 2 (Bcl-2), BCL2 associated X (Bax), zinc finger protein 667 (ZNF667), and miR-181a-5p were detected. Cell apoptosis and viability were detected. The enrichment of IRF8 on the miR-181a-5p promoter was assayed. The binding relationship between IRF8 and miR-181a-5p promoter, and between miR-181a-5p and ZNF667 3'UTR sequence was verified. Combined experiments with miR-181a-5p knockdown or ZNF667 overexpression were performed to observe the changes in cell apoptosis.

Results: IRF8 and ZNF667 were increased in SONFH cells and miR-181a-5p was decreased. Inhibition of IRF8 increased SONFH cell viability and reduced apoptosis. Mechanistically, IRF8 was enriched in the miR-181a-5p promoter to inhibit miR-181a-5p and miR-181a-5p targeted and inhibited ZNF667. miR-181a-5p knockdown or ZNF667 overexpression could alleviate the inhibitory effect of IRF8 down-regulation on osteocyte apoptosis in SONFH.

Conclusion: IRF8 was enriched in the miR-181a-5p promoter to inhibit miR-181a-5p, thus promoting ZNF667 levels and increasing osteocyte apoptosis in SONFH, which may be a new theoretical basis for the treatment of SONFH.