Extracellular vesicles derived from mesenchymal stem cells mediate extracellular matrix remodeling in osteoarthritis through the transport of microRNA-29a.

BACKGROUND: Knee osteoarthritis (KOA) is a common orthopedic condition with an uncertain etiology, possibly involving genetics and biomechanics. Factors like changes in chondrocyte microenvironment, oxidative stress, inflammation, and immune responses affect KOA development. Early-stage treatment options primarily target symptom relief. Mesenchymal stem cells (MSCs) show promise for treatment, despite challenges. Recent research highlights microRNAs (miRNAs) within MSC-released extracellular vesicles that can potentially promote cartilage regeneration and hinder KOA progression. This suggests exosomes (Exos) as a promising avenue for future treatment. While these findings emphasize the need for effective KOA progression management, further safety and efficacy validation for Exos is essential. AIM: To explore miR-29a's role in KOA, we'll create miR-29a-loaded vesicles, testing for early treatment in rat models. METHODS: Extraction of bone marrow MSC-derived extracellular vesicles, preparation of engineered vesicles loaded with miR-29a using ultrasonication, and identification using quantitative reverse transcription polymerase chain reaction; after establishing a rat model of KOA, rats were randomly divided into three groups: Blank control group injected with saline, normal extracellular vesicle group injected with normal extracellular vesicle suspension, and engineered extracellular vesicle group injected with engineered extracellular vesicle suspension. The three groups were subjected to general behavioral observation analysis, imaging evaluation, gross histological observation evaluation, histological detection, and immunohistochemical detection to compare and evaluate the progress of various forms of arthritis. RESULTS: General behavioral observation results showed that the extracellular vesicle group and engineered extracellular vesicle group had better performance in all four indicators of pain, gait, joint mobility, and swelling compared to the blank control group. Additionally, the engineered extracellular vesicle group had better pain relief at 4 wk and better knee joint mobility at 8 wk compared to the normal extracellular vesicle group. Imaging examination results showed that the blank control group had the fastest progression of arthritis, the normal extracellular vesicle group had a relatively slower progression, and the engineered extracellular vesicle group had the slowest progression. Gross histological observation results showed that the blank control group had the most obvious signs of arthritis, the normal extracellular vesicle group showed signs of arthritis, and the engineered extracellular vesicle group showed no significant signs of arthritis. Using the Pelletier gross score evaluation, the engineered extracellular vesicle group had the slowest progression of arthritis. Results from two types of staining showed that the articular cartilage of rats in the normal extracellular vesicle and engineered extracellular vesicle groups was significantly better than that of the blank control group, and the engineered extracellular vesicle group had the best cartilage cell and joint surface condition. Immunohistochemical detection of type II collagen and proteoglycan showed that the extracellular matrix of cartilage cells in the normal extracellular vesicle and engineered extracellular vesicle groups was better than that of the blank control group. Compared to the normal extracellular vesicle group, the engineered extracellular vesicle group had a better regulatory effect on the extracellular matrix of cartilage cells. CONCLUSION: Engineered Exos loaded with miR-29a can exert anti-inflammatory effects and maintain extracellular matrix stability, thereby protecting articular cartilage, and slowing the progression of KOA.

Humoral immune response to tumor-associated antigen Ubiquilin 1 (UBQLN1) and its tumor-promoting potential in lung cancer.

BACKGROUND: This study aims to investigate the expression of UBQLN1 in lung cancer (LC) tissue and the diagnostic capability of autoantibody to UBQLN1 (anti-UBQLN1) in the detection of LC and the discrimination of pulmonary nodules (PNs). METHODS: Sera from 798 participants were used to discover and validate the level of autoantibodies via HuProt microarray and Enzyme-linked immunosorbent assay (ELISA). Logistic regression analysis was applied to establish model. Receiver operating characteristic curve (ROC) analysis was performed to evaluate the diagnostic potential. Immunohistochemistry was performed to detect UBQLN1 expression in 88 LC tissues and 88 para-tumor tissues. qRT-PCR and western blotting were performed to detect the expression of UBQLN1 at the mRNA and protein levels, respectively. Trans-well assay and cell counting kit-8 (CCK-8) was used to investigate the function of UBQLN1. RESULTS: Anti-UBQLN1 was identified with the highest fold change by protein microarray. The level of anti-UBQLN1 in LC patients was obviously higher than that in NC or patients with benign lung disease of validation cohort 1 (P<0.05). The area under the curve (AUC) of anti-UBQLN1 was 0.610 (95%CI: 0.508-0.713) while reached at 0.822 (95%CI: 0.784-0.897) when combining anti-UBQLN1 with CEA, CYFRA21-1, CA125 and three CT indicators (vascular notch sign, lobulation sign and mediastinal lymph node enlargement) in the discrimination of PNs. UBQLN1 protein was overexpressed in lung adenocarcinoma (LUAD) tissues compared to para-tumor tissues. UBQLN1 knockdown remarkably inhibited the migration, invasion and proliferation of LUAD cell lines. CONCLUSIONS: Anti-UBQLN1 might be a potential biomarker for the diagnosis of LC and the discrimination of PNs.

Signature construction and molecular subtype identification based on liver-specific genes for prediction of prognosis, immune activity, and anti-cancer drug sensitivity in hepatocellular carcinoma.

BACKGROUND: Liver specific genes (LSGs) are crucial for hepatocyte differentiation and maintaining normal liver function. A deep understanding of LSGs and their heterogeneity in hepatocellular carcinoma (HCC) is necessary to provide clues for HCC diagnosis, prognosis, and treatment. METHODS: The bulk and single-cell RNA-seq data of HCC were downloaded from TCGA, ICGC, and GEO databases. Through unsupervised cluster analysis, LSGs-based HCC subtypes were identified in TCGA-HCC samples. The prognostic effects of the subtypes were investigated with survival analyses. With GSVA and Wilcoxon test, the LSGs score, stemness score, aging score, immune score and stromal score of the samples were estimated and compared. The HCC subtype-specific genes were identified. The subtypes and their differences were validated in ICGC-HCC samples. LASSO regression analysis was used for key gene selection and risk model construction for HCC overall survival. The model performance was estimated and validated. The key genes were validated for their heterogeneities in HCC cell lines with quantitative real-time PCR and at single-cell level. Their dysregulations were investigated at protein level. Their correlations with HCC response to anti-cancer drugs were estimated in HCC cell lines. RESULTS: We identified three LSGs-based HCC subtypes with different prognosis, tumor stemness, and aging level. The C1 subtype with low LSGs score and high immune score presented a poor survival, while the C2 subtype with high LSGs score and immune score indicated an enduring survival. Although no significant survival difference between C2 and C3 HCCs was shown, the C2 HCCs presented higher immune score and stroma score. The HCC subtypes and their differences were confirmed in ICGC-HCC dataset. A five-gene prognostic signature for HCC survival was constructed. Its good performance was shown in both the training and validation datasets. The five genes presented significant heterogeneities in different HCC cell lines and hepatocyte subclusters. Their dysregulations were confirmed at protein level. Furthermore, their significant associations with HCC sensitivities to anti-cancer drugs were shown. CONCLUSIONS: LSGs-based HCC subtype classification and the five-gene risk model might provide useful clues not only for HCC stratification and risk prediction, but also for the development of more personalized therapies for effective HCC treatment.

Blood FOLR3 methylation dysregulations and heterogeneity in non-small lung cancer highlight its strong associations with lung squamous carcinoma.

BACKGROUND: Non-small cell lung cancer (NSCLC) accounts for the vast majority of lung cancers. Early detection is crucial to reduce lung cancer-related mortality. Aberrant DNA methylation occurs early during carcinogenesis and can be detected in blood. It is essential to investigate the dysregulated blood methylation markers for early diagnosis of NSCLC. METHODS: NSCLC-associated methylation gene folate receptor gamma (FOLR3) was selected from an Illumina 850K array analysis of peripheral blood samples. Mass spectrometry was used for validation in two independent case-control studies (validation I: n = 2548; validation II: n = 3866). Patients with lung squamous carcinoma (LUSC) or lung adenocarcinoma (LUAD), normal controls (NCs) and benign pulmonary nodule (BPN) cases were included. FOLR3 methylations were compared among different populations. Their associations with NSCLC clinical features were investigated. Receiver operating characteristic analyses, Kruskal-Wallis test, Wilcoxon test, logistics regression analysis and nomogram analysis were performed. RESULTS: Two CpG sites (CpG_1 and CpG_2) of FOLR3 was significantly lower methylated in NSCLC patients than NCs in the discovery round. In the two validations, both LUSC and LUAD patients presented significant FOLR3 hypomethylations. LUSC patients were highlighted to have significantly lower methylation levels of CpG_1 and CpG_2 than BPN cases and LUAD patients. Both in the two validations, CpG_1 methylation and CpG_2 methylation could discriminate LUSC from NCs well, with areas under the curve (AUCs) of 0.818 and 0.832 in validation I, and 0.789 and 0.780 in validation II. They could also differentiate LUAD from NCs, but with lower efficiency. CpG_1 and CpG_2 methylations could also discriminate LUSC from BPNs well individually in the two validations. With the combined dataset of two validations, the independent associations of age, gender, and FOLR3 methylation with LUSC and LUAD risk were shown and the age-gender-CpG_1 signature could discriminate LUSC and LUAD from NCs and BPNs, with higher efficiency for LUSC. CONCLUSIONS: Blood-based FOLR3 hypomethylation was shown in LUSC and LUAD. FOLR3 methylation heterogeneity between LUSC and LUAD highlighted its stronger associations with LUSC. FOLR3 methylation and the age-gender-CpG_1 signature might be novel diagnostic markers for the early detection of NSCLC, especially for LUSC.

Low-Dose Hexavalent Chromium Exposure Induces Endoplasmic Reticulum Stress-Mediated Apoptosis in Rat Liver.

This study investigated the toxic effects of low-dose hexavalent chromium (Cr(VI)) on rat liver. Male specific pathogen-free (SPF) Sprague-Dawley (SD) rats (4-5 weeks of age) were randomly divided into groups: saline, 0.05 mg/kg Cr(VI), and 0.25 mg/kg Cr(VI). The rats were subjected to intratracheal instillation of K2Cr2O7 suspensions or saline once weekly, for a total of five times. The results showed that the accumulation of Cr(VI) in the blood of the 0.25 mg/kg K2Cr2O7 group was significantly higher than that in the saline group. Transmission electron microscopy (TEM) showed that exposure to hexavalent chromium caused endoplasmic reticulum (ER) oedema and a disordered arrangement. The levels of endoplasmic reticulum stress (ERS)-related proteins (ATF6, P-PERK, P-IRE1, Grp78, and CHOP) in the 0.25 mg/kg K2Cr2O7 group were significantly higher than those in the saline group. The expression of apoptosis-inhibitory protein Bcl-2 was significantly lower in the 0.25 mg/kg K2Cr2O7 group than that in the saline group, and the expression of apoptosis protein Bax was significantly higher in the 0.25 mg/kg K2Cr2O7 group than that in the saline group, indicating that Cr(VI) increased apoptosis. These findings revealed that Cr(VI) may be involved in rat liver injury by initiating ERS-mediated apoptosis. The expression of ATF6, P-PERK, P-IRE1, and Bax in the 0.05 mg/kg K2Cr2O7 group was not significantly different from that in the saline group, and the different effects produced by the two different dose groups provide a possible experimental basis for further study of occupational exposure limits.

Hexavalent chromium caused DNA damage repair and apoptosis via the PI3K/AKT/FOXO1 pathway triggered by oxidative stress in the lung of rat.

Hexavalent chromium [Cr(VI)] is an occupational carcinogen that accumulates in the lungs and causes lung injury and even lung cancer. 36 SD male rats received inhalable intratracheal instillation of Cr(VI) (0.05, 0.25 mg Cr/kg) or the same volume (3 ml/kg) of normal saline weekly for 28 days (total 5 times). After 28 days of exposure, half of the rats in each group were sacrificed for investigation, and the rest stopped exposure and began to be self-repaired for two weeks. Histopathology analyses revealed that Cr(VI) induced slight dilatation and hemorrhage of perialveolar capillaries, pulmonary bronchodilation, and congestion with peripheral flaky-like necrosis accompanied by inflammatory cell infiltration, especially the 0.25 mg Cr/kg group. Cr(VI) exposure caused the increase of blood Cr, urinary Cr, MDA, urinary 8-hydroxy-2' -deoxyguanosine (8-OHdG), and the decrease of GSH and MDA, while two-week repair only reduced urinary Cr. Exposure to Cr(VI) significantly upregulated FOXO1 and downregulated p-AKT and p-FOXO1 for two weeks. PI3K in the 0.25 mg Cr/kg group was inhibited after two weeks of repair. Cr(VI) exposure mainly promoted GADD45a and CHK2 in the exposure group, promoted Bim, Bax/Bcl-2, and suppressed Bcl-2 and Bcl-xL in the repair group. These results demonstrate that Cr(VI) may induce DNA damage repair and apoptosis in the lung by activating the PI3K/AKT/FOXO1 pathway. Two-week repair may alleviate oxidative stress and DNA damage induced by Cr(VI) exposure but couldn't eliminate its effects. This study provides a new perspective for exploring the Cr(VI) induced lung cancer mechanism.

Dysregulations of metabolites and gut microbes and their associations in rats with noise induced hearing loss.

Background: Noise exposure could lead to hearing loss and disorders of various organs. Recent studies have reported the close relations of environmental noise exposure to the metabolomics dysregulations and gut microbiota disturbance in the exposers. However, the associations between gut microbial homeostasis and the body metabolism during noise-induced hearing loss (NIHL) were unclear. To get a full understanding of their synergy in noise-associated diseases, it is essential to uncover their impacts and associations under exposure conditions. Methods: With ten male rats with background noise exposure (≤ 40 dB) as controls (Ctr group), 20 age- and weight-matched male rats were exposed to 95 dB Sound pressure level (SPL) (LN group, n = 10) or 105 dB SPL noise (HN group, n = 10) for 30 days with 4 h/d. The auditory brainstem response (ABR) of the rats and their serum biochemical parameters were detected to investigate their hearing status and the potential effects of noise exposure on other organs. Metabolomics (UPLC/Q-TOF-MS) and microbiome (16S rDNA gene sequencing) analyses were performed on samples from the rats. Multivariate analyses and functional enrichments were applied to identify the dysregulated metabolites and gut microbes as well as their associated pathways. Pearson correlation analysis was performed to investigate the associations of the dysregulations of microbiota and the metabolites. Results: NIHL rat models were constructed. Many biochemical parameters were altered by noise exposure. The gut microbiota constitution and serum metabolic profiles of the noise-exposed rats were also dysregulated. Through metabolomics analysis, 34 and 36 differential metabolites as well as their associated pathways were identified in LN and HN groups, respectively. Comparing with the control rats, six and 14 florae were shown to be significantly dysregulated in the LN group and HN group, respectively. Further association analysis showed significant correlations between differential metabolites and differential microbiota. Conclusion: There were cochlea injuries and abnormalities of biochemical parameters in the rats with NIHL. Noise exposure could also disrupt the metabolic profiles and the homeostatic balance of gut microbes of the host as well as their correlations. The dysregulated metabolites and microbiota might provide new clues for prevention of noise-related disorders.

Changes in soil fertility under partial organic substitution of chemical fertilizer: a 33-year trial.

BACKGROUND: This study examined the changes in soil fertility in a maize cropping area when chemical fertilizer was partially replaced with straw or livestock manure over a 33-year period. Four treatments were included: (i) CK (no fertilizer application); (ii) NPK (only chemical fertilizer application); (iii) NPKM (chemical fertilizer partly replaced with livestock manure); (iv) NPKS (chemical fertilizer partly replaced with straw). RESULTS: Soil organic carbon increased by 41.7% and 95.5% in the NPKS and NPKM treatments, respectively, over the 33-year trial compared with the initial concentration. However, soil organic carbon in NPK was significantly reduced by 9.8%. Soil total N, P and K increased in both NPKM and NPKS treatments compared to the original soil. Soil pH was significantly acidified from 7.6 to 5.97 in the NPK treatment during the experimental period. The NPKM and NPKS treatments buffered the acidification compared to NPK. Meta-analysis results showed that, compared with NPK, NPKM significantly raised soil bacteria and fungi populations by 38.7% and 58.6%; enhanced microbial biomass carbon and nitrogen by 66.3% and 63%, respectively; and increased sucrase, urease and catalase activities by 34.2%, 48.2% and 21.5%. NPKS significantly increased soil fungi and actinomycetes populations by 24.3% and 41.2%, respectively; enhanced microbial biomass carbon and nitrogen by 27.1% and 45%; and strengthened sucrase and urease activities by 36% and 20.3%, respectively. CONCLUSION: Long-term chemical fertilizer application led to the deterioration of soil fertility and environment. Partial replacement of chemical fertilizers with organic materials could significantly amend and buffer such negative effects. © 2023 Society of Chemical Industry.

Bone marrow mesenchymal stem cells paracrine TGF-ß1 to mediate the biological activity of osteoblasts in bone repair.

BACKGROUND: Bone marrow mesenchymal stem cells (BMSCs) are an important source of seed cells for regenerative medicine and tissue engineering therapy. BMSCs have multiple differentiation potentials and can release paracrine factors to facilitate tissue repair. Although the role of the osteogenic differentiation of BMSCs has been fully confirmed, the function and mechanism of BMSC paracrine factors in bone repair are still largely unclear. This study aimed to determine the roles of transforming growth factor beta-1 (TGF-ß1) produced by BMSCs in bone tissue repair. METHODS: To confirm our hypothesis, we used a Transwell system to coculture hBMSCs and human osteoblast-like cells without contact, which could not only avoid the interference of the osteogenic differentiation of hBMSCs but also establish the cell-cell relationship between hBMSCs and human osteoblast-like cells and provide stable paracrine substances. In the transwell coculture system, alkaline phosphatase activity, mineralized nodule formation, cell migration and chemotaxis analysis assays were conducted. RESULTS: Osteogenesis, migration and chemotaxis of osteoblast-like cells were regulated by BMSCs in a paracrine manner via the upregulation of osteogenic and migration-associated genes. A TGF-ß receptor I inhibitor (LY3200882) significantly antagonized BMSC-induced biological activity and related gene expression in osteoblast-like cells. Interestingly, coculture with osteoblast-like cells significantly increased the production of TGF-ß1 by BMSCs, and there was potential intercellular communication between BMSCs and osteoblast-like cells. CONCLUSIONS: Our findings provide evidence that the biological mechanism of BMSC-produced TGF-ß1 promotes bone regeneration and repair, providing a theoretical basis and new directions for the application of BMSC transplantation in the treatment of osteonecrosis and bone injury.

CD5L-associated gene analyses highlight the dysregulations, prognostic effects, immune associations, and drug-sensitivity predicative potentials of LCAT and CDC20 in hepatocellular carcinoma.

BACKGROUND: The dysregulation of CD5L has been reported in hepatocellular carcinoma (HCC). However, its functions in HCC were controversial. In this study, we aimed to identify CD5L-associated pathways and markers and explore their values in HCC diagnosis, prognosis and treatment. METHODS: HCC datasets with gene expression profiles and clinical data in TCGA and ICGC were downloaded. The immune/stroma cell infiltrations were estimated with xCell. CD5L-associated pathways and CD5L-associated genes (CD5L-AGs) were identified with gene expression comparisons and gene set enrichment analysis (GSEA). Cox regression, Kaplan-Meier survival analysis, and least absolute shrinkage and selection operator (LASSO) regression analysis were performed. The correlations of the key genes with immune/stroma infiltrations, immunoregulators, and anti-cancer drug sensitivities in HCC were investigated. At protein level, the key genes dysregulations, their correlations and prognostic values were validated in clinical proteomic tumor analysis consortium (CPTAC) database. Serum CD5L and LCAT activity in 50 HCC and 30 normal samples were evaluated and compared. The correlations of serum LCAT activity with alpha-fetoprotein (AFP), albumin (ALB) and high-density lipoprotein (HDL) in HCC were also investigated. RESULTS: Through systemic analyses, 14 CD5L-associated biological pathways, 256 CD5L-AGs and 28 CD5L-associated prognostic and diagnostic genes (CD5L-APDGs) were identified. A risk model consisting of LCAT and CDC20 was constructed for HCC overall survival (OS), which could discriminate HCC OS status effectively in both the training and the validation sets. CD5L, LCAT and CDC20 were shown to be significantly correlated with immune/stroma cell infiltrations, immunoregulators and 31 anti-cancer drug sensitivities in HCC. At protein level, the dysregulations of CD5L, LCAT and CDC20 were confirmed. LCAT and CDC20 were shown to be significantly correlated with proliferation marker MKI67. In serum, no significance of CD5L was shown. However, the lower activity of LCAT in HCC serum was obvious, as well as its significant positive correlations ALB and HDL concentrations. CONCLUSIONS: CD5L, LCAT and CDC20 were dysregulated in HCC both at mRNA and protein levels. The LCAT-CDC20 signature might be new predicator for HCC OS. The associations of the three genes with HCC microenvironment and anti-cancer drug sensitivities would provide new clues for HCC immunotherapy and chemotherapy.

CapG promoted nasopharyngeal carcinoma cell motility involving Rho motility pathway independent of ROCK.

BACKGROUND: Gelsolin-like capping actin protein (CapG) modulates actin dynamics and actin-based motility with a debatable role in tumorigenic progression. The motility-associated functions and potential molecular mechanisms of CapG in nasopharyngeal carcinoma (NPC) remain unclear. METHODS: CapG expression was detected by immunohistochemistry in a cohort of NPC tissue specimens and by Western blotting assay in a variety of NPC cell lines. Loss of function and gain of function of CapG in scratch wound-healing and transwell assays were performed. Inactivation of Rac1 and ROCK with the specific small molecular inhibitors was applied to evaluate CapG's role in NPC cell motility. GTP-bound Rac1 and phosphorylated-myosin light chain 2 (p-MLC2) were measured in the ectopic CapG overexpressing cells. Finally, CapG-related gene set enrichment analysis was conducted to figure out the significant CapG-associated pathways in NPC. RESULTS: CapG disclosed increased level in the poorly differentiated NPC tissues and highly metastatic cells. Knockdown of CapG reduced NPC cell migration and invasion in vitro, while ectopic CapG overexpression showed the opposite effect. Ectopic overexpression of CapG compensated for the cell motility loss caused by simultaneous inactivation of ROCK and Rac1 or inactivation of ROCK alone. GTP-bound Rac1 weakened, and p-MLC2 increased in the CapG overexpressing cells. Bioinformatics analysis validated a positive correlation of CapG with Rho motility signaling, while Rac1 motility pathway showed no significant relationship. CONCLUSIONS: The present findings highlight the contribution of CapG to NPC cell motility independent of ROCK and Rac1. CapG promotes NPC cell motility at least partly through MLC2 phosphorylation and contradicts with Rac1 activation.

Effects of angiotensin II combined with asparaginase and dexamethasone on the femoral head in mice: A model of steroid-induced femoral head osteonecrosis.

Background: To study the pathogenesis of steroid-induced femoral head osteonecrosis, an ideal animal model is very important. As experimental animals, mice are beneficial for studying the pathogenesis of disease. However, there are currently few mouse models of steroid-induced femoral head osteonecrosis, and there are many questions that require further exploration and research. Purposes: The purpose of this study was to establish a new model of osteonecrosis in mice using angiotensin II (Ang II) combined with asparaginase (ASP) and dexamethasone (DEX) and to study the effects of this drug combination on femoral head osteonecrosis in mice. Methods: Male BALB/c mice (n = 60) were randomly divided into three groups. Group A (normal control, NC) was treated with physiological saline and given a normal diet. Group B (DEX + ASP, DA) was given free access to food and water (containing 2 mg/L DEX) and subjected to intraperitoneal injection of ASP (1200 IU/kg twice/week for 8 weeks). Group C (DEX + ASP + Ang II, DAA) was treated the same as group B, it was also given free access to food and water (containing 2 mg/L DEX) and subjected to intraperitoneal injection of ASP (1200 IU/kg twice/week for 8 weeks), but in the 4th and 8th weeks, subcutaneous implantation of a capsule osmotic pump (0.28 mg/kg/day Ang II) was performed. The mice were sacrificed in the 4th and 8th weeks, and the model success rate, mouse mortality rate, body weight, blood lipids, coagulation factors, histopathology, and number of local vessels in the femoral head were evaluated. Results: DAA increased the model success rate [4th week, 30% (DA) vs. 40% (DAA) vs. 0% (NC); 8th week, 40% (DA) vs. 70% (DAA) vs. 0% (NC)]. There was no significant difference in mortality rate between the groups [4th week, 0% (DA) vs. 0% (DAA) vs. 0% (NC); 8th week, 5% (DA) vs. 10% (DAA) vs. 0% (NC)]. DAA affected mouse body weight and significantly affected blood lipids and blood coagulation factors. DAA reduces the number of blood vessels in the femoral head and destroys the local blood supply. Conclusion: Angiotensin II combined with asparaginase and dexamethasone can obviously promote the necrosis of femoral head and provide a new idea for the model and treatment of osteonecrosis.

Modulation of Osteogenesis and Angiogenesis Activities Based on Ionic Release from Zn-Mg Alloys.

The enhancement of osteogenesis and angiogenesis remains a great challenge for the successful regeneration of engineered tissue. Biodegradable Mg and Zn alloys have received increasing interest as potential biodegradable metallic materials, partially due to the biological functions of Mg2+ and Zn2+ with regard to osteogenesis and angiogenesis, respectively. In the present study, novel biodegradable Zn-xMg (x = 0.2, 0.5, 1.0 wt.%) alloys were designed and fabricated, and the effects of adding different amounts of Mg to the Zn matrix were investigated. The osteogenesis and angiogenesis beneficial effects of Zn2+ and Mg2+ release during the biodegradation were characterized, demonstrating coordination with the bone regeneration process in a dose-dependent manner. The results show that increased Mg content leads to a higher amount of released Mg2+ while decreasing the Zn2+ concentration in the extract. The osteogenesis of pre-osteoblasts was promoted in Zn-0.5Mg and Zn-1Mg due to the higher concentration of Mg2+. Moreover, pure Zn extract presented the highest activity in angiogenesis, owing to the highest concentration of Zn2+ release (6.415 µg/mL); the proliferation of osteoblast cells was, however, inhibited under such a high Zn2+ concentration. Although the concentration of Zn ion was decreased in Zn-0.5Mg and Zn-1Mg compared with pure Zn, the angiogenesis was not influenced when the concentration of Mg in the extract was sufficiently increased. Hence, Mg2+ and Zn2+ in Zn-Mg alloys show a dual modulation effect. The Zn-0.5Mg alloy was indicated to be a promising implant candidate due to demonstrating the appropriate activity in regulating osteogenesis and angiogenesis. The present work evaluates the effect of the Mg content in Zn-based alloys on biological activities, and the results provide guidance regarding the Zn-Mg composition in designs for orthopedic application.

Plasma metabolomics analyses highlight the multifaceted effects of noise exposure and the diagnostic power of dysregulated metabolites for noise-induced hearing loss in steel workers.

Noise exposure can lead to various kinds of disorders. Noise-induced hearing loss (NIHL) is one of the leading disorders confusing the noise-exposed workers. It is essential to identify NIHL markers for its early diagnosis and new therapeutic targets for its treatment. In this study, a total of 90 plasma samples from 60 noise-exposed steel factory male workers (the noise group) with (NIHL group, n = 30) and without NIHL (non-NIHL group, n = 30) and 30 male controls without noise exposure (control group) were collected. Untargeted human plasma metabolomic profiles were determined with HPLC-MS/MS. The levels of the metabolites in the samples were normalized to total peak intensity, and the processed data were subjected to multivariate data analysis. The Wilcoxon test and orthogonal partial least square-discriminant analysis (OPLS-DA) were performed. With the threshold of p < 0.05 and the variable importance of projection (VIP) value >1, 469 differential plasma metabolites associated with noise exposure (DMs-NE) were identified, and their associated 58 KEGG pathways were indicated. In total, 33 differential metabolites associated with NIHL (DMs-NIHL) and their associated 12 KEGG pathways were identified. There were six common pathways associated with both noise exposure and NIHL. Through multiple comparisons, seven metabolites were shown to be dysregulated in the NIHL group compared with the other two groups. Through LASSO regression analysis, two risk models were constructed for NIHL status predication which could discriminate NIHL from non-NIHL workers with the area under the curve (AUC) values of 0.840 and 0.872, respectively, indicating their efficiency in NIHL diagnosis. To validate the results of the metabolomics, cochlear gene expression comparisons between susceptible and resistant mice in the GSE8342 dataset from Gene Expression Omnibus (GEO) were performed. The immune response and cell death-related processes were highlighted for their close relations with noise exposure, indicating their critical roles in noise-induced disorders. We concluded that there was a significant difference between the metabolite's profiles between NIHL cases and non-NIHL individuals. Noise exposure could lead to dysregulations of a variety of biological pathways, especially immune response and cell death-related processes. Our results might provide new clues for noise exposure studies and NIHL diagnosis.

Ginkgo biloba L. extract prevents steroid-induced necrosis of the femoral head by rescuing apoptosis and dysfunction in vascular endothelial cells via the PI3K/AKT/eNOS pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Ginkgo biloba L. extract (EGb) is one of the world's most extensively used herbal medicines. Due to the diverse pharmacological properties of EGb, it has been used in the treatment of neurological illnesses, as well as cardiovascular and cerebrovascular ailments. However, the effect and pharmacological mechanism of EGb on steroid-induced necrosis of the femoral head (SINFH) are still unclear. AIM OF THE STUDY: SINFH remains a challenging problem in orthopedics. Previous investigations have shown that EGb has the potential to reduce the occurrence of SINFH. The goal was to determine the effect and mechanism of EGb in preventing SINFH by inhibiting apoptosis and improving vascular endothelial cells (VECs) functions. MATERIALS AND METHODS: CCK-8, nitric oxide (NO) production and flow cytometry were used to determine the cell apoptosis and function. The scratch and angiogenesis tests assessed migration and tube formation. Western blot analysis detected the expressions of apoptosis-related proteins and PI3K/AKT/eNOS pathway-related proteins. Apoptosis and angiogenesis were also detected treated with the inhibitors. A mouse model of SINFH was established. Paraffin section was used to determine the necrotic pathology and apoptosis. Vessels in the femoral heads were assessed by immunofluorescence staining. RESULTS: When stimulated by methylprednisolone (MPS), cell viability, NO generation and tube formation were decreased, the apoptotic rate increased. Simultaneously, MPS decreased the expression levels of p-PI3K, p-AKT, and p-eNOS. EGb increased the expression levels of these proteins, restrained apoptosis, and restored cell functions. The addition of the inhibitors decreased anti-apoptotic effect and angiogenesis. In addition, when compared to the model mice, there were fewer empty lacunae and normal trabecular arrangement after taking different doses of EGb. The protective effect was also confirmed by the vascular quantitative analysis in vivo. CONCLUSION: This study established that EGb increased endothelial cell activity and inhibited apoptosis and function loss induced by MPS, elucidating the effect and molecular mechanism of EGb on early SINFH.

Hsa_circ_0001495 contributes to cervical cancer progression by targeting miR-526b-3p/TMBIM6/mTOR axis.

Cervical cancer (CC) is a common gynecological malignant tumor, causing poor survival rate. Circular RNAs (circRNAs) are abundantly expressed in CC with their stable loop structure. However, the underlying mechanism and biological function of circRNAs remained unclear. Using quantitative real-time polymerase chain reaction (qRT-PCR) or western blot assay, we measured the expression of hsa_circ_0001495, miR-526b-3p, and transmembrane Bax inhibitor motif containing 6 (TMBIM6) in CC tissues and cells. The relationship between miR-526b-3p and hsa_circ_0001495 or TMBIM6 was investigated by bioinformatics analysis, dual-luciferase and RIP analysis. Enzyme linked immunosorbent assay (ELISA) was conducted to evaluate glucose consumption and lactate production. 5-ethynyl-2'-deoxyuridine (EDU) assay were used to test cell proliferation. Cell apoptosis was analyzed by using flow cytometry assay. Transwell and wound-healing assays were used to measure cell invasion and migration. The expression of proteins was examined by western blot. Xenograft assay was applied to detect the effect of hsa_circ_0001495 in vivo. Our finding showed that hsa_circ_0001495 and TMBIM6 expression were upregulated, while miR-526b-3p was downregulated in CC tissues and cell lines. Hsa_circ_0001495 knockdown or TMBIM6 knockdown suppressed cell proliferation, migration, glycolysis, while promoted cell apoptosis in vitro, and hsa_circ_0001495 silence curbed tumor growth in vivo. Beside, hsa_circ_0001495 exerted its function in CC by positively regulating TMBIM6. Furthermore, hsa_circ_0001495 acted as a sponge for miR-526b-3p to regulate TMBIM6 expression. Hsa_circ_0001495/miR-526b-3p/TMBIM6 axis also regulated the phosphorylation of mammalian target of rapamycin (mTOR) in CC cells. In summary, hsa_circ_0001495 regulated the progression of CC by regulating miR-526b-3p/TMBIM6/mTOR pathway.

Skimmianine as a novel therapeutic agent suppresses proliferation and migration of human esophageal squamous cell carcinoma via blocking the activation of ERK1/2.

Esophageal squamous cell carcinoma (ESCC), one of the main histopathological subtypes of esophageal cancer (EC), is characterized by high morbidity and mortality. Clinical treatment for ESCC lacks specific molecular targets and effective therapeutic drugs. Skimmianine (SK), one of the natural fluroquinolone alkaloids, is widely present in Rutaceae family plants. Here, we mainly used CCK-8 assay, clone formation, flow cytometry analysis, wound-healing assay, Transwell assay, western blot, quantitative real-time PCR (qRT-PCR), molecular docking analysis, tumor xenograft assay, and immunohistochemistry (IHC) staining to investigate the potential anti-tumor effect of SK on ESCC. We demonstrated that SK inhibited the proliferation of TE-1 and Eca109 cells via inducing the G0/G1 phase cell cycle arrest, prevented the migration and invasion of tumor cells via regulating epithelial-mesenchymal transition (EMT) in vitro. In addition, SK obviously suppressed the growth of xenografted Eca109 tumors in nude mice. The anti-tumor mechanism of SK could be blocking the activation of extracellular signal-regulated kinases 1/2 (ERK1/2) in the mitogen-activated protein kinase (MAPK)/ERK signaling pathway. Our basic research suggests that SK can be a potential therapeutic agent for ESCC.

The m6 A reader MhYTP2 regulates MdMLO19 mRNA stability and antioxidant genes translation efficiency conferring powdery mildew resistance in apple.

N6 -methyladenosine (m6 A) reader protein plays an important role in trichome morphology, developmental timing and morphogenesis in Arabidopsis. However, the function of m6 A readers in plant-microbe interaction remains unclear. Here, a Malus YTH-domain family protein MhYTP2 was initially characterized as an m6 A reader. MhYTP2 overexpression increased mRNA m6 A modification level and translation efficiency. The m6 A in the exon regions appeared to destabilize the mRNAs, whereas m6 A in the untranslated regions positively correlated with the associated mRNA abundance. MhYTP2 overexpression enhanced apple powdery mildew resistance, possibly by rapidly degrading the bound mRNAs of MdMLO19 and MdMLO19-X1 and improving the translation efficiency of the antioxidant genes. To conclude, the results shed light on the apple m6 A profile, the effect of MhYTP2 on m6 A profile, and the m6 A roles in MdMLO19 and MdMLO19-X1 mRNAs stability and glutamate dehydrogenase 1-like MdGDH1L mRNA translation efficiency.

Activation of the ABA Signal Pathway Mediated by GABA Improves the Drought Resistance of Apple Seedlings.

Drought seriously affects the yield and quality of apples. γ-aminobutyric acid (GABA) plays an important role in the responses of plants to various stresses. However, the role and possible mechanism of GABA in the drought response of apple seedlings remain unknown. To explore the effect of GABA on apple seedlings under drought stress, seedlings of Malus hupehensis were treated with seven concentrations of GABA, and the response of seedlings under 15-day drought stress was observed. The results showed that 0.5 mM GABA was the most effective at relieving drought stress. Treatment with GABA reduced the relative electrical conductivity and MDA content of leaves induced by drought stress and significantly increased the relative water content of leaves. Exogenous GABA significantly decreased the stomatal conductance and intercellular carbon dioxide concentration and transpiration rate, and it significantly increased the photosynthetic rate under drought. GABA also reduced the accumulation of superoxide anions and hydrogen peroxide in leaf tissues under drought and increased the activities of POD, SOD, and CAT and the content of GABA. Exogenous treatment with GABA acted through the accumulation of abscisic acid (ABA) in the leaves to significantly decrease stomatal conductance and increase the stomatal closure rate, and the levels of expression of ABA-related genes PYL4, ABI1, ABI2, HAB1, ABF3, and OST1 changed in response to drought. Taken together, exogenous GABA can enhance the drought tolerance of apple seedlings.

Protein deubiquitylase USP3 stabilizes Aurora A to promote proliferation and metastasis of esophageal squamous cell carcinoma.

Aurora A kinase is a cell cycle regulator that is dysregulated in several different malignancies. Nevertheless, its regulatory mechanisms are still not fully understood. Here, we report that ubiquitin specific peptidase 3 (USP3) promotes proliferation and metastasis of esophageal squamous cell carcinoma (ESCC) cells by mediating deubiquitination of Aurora A. Analysis of human clinical samples indicated that USP3 and Aurora A are highly expressed in ESCC. Cellular experiments confirmed that high expression of USP3 and Aurora A in ESCC cells promoted malignant cell proliferation and invasion. In this mechanism, USP3 leads to suppression of Aurora A ubiquitination, resulting less proteasome degradation. We constructed the deubiquitinated mimetic K143R of Aurora A and found that K143R significantly promoted the proliferation and invasion of ESCC cells and was not regulated by the deubiquitination of USP3. Moreover, Aurora A K143R potentiated the kinase activity of Aurora A in ESCC cells. Thus, our findings demonstrate that the tumorigenic feature of ESCC is in part mediated by USP3-facilitated deubiquitination of Aurora A.