PI3K/AKT/mTOR signaling regulates BCP ceramic-induced osteogenesis.

An increasing number of studies demonstrate that biphasic calcium phosphate (BCP) ceramics can induce bone regeneration. However, the underlying molecular mechanisms involved are still poorly understood. This work was proposed to investigate how PI3K/AKT/mTOR signaling influenced the osteogenesis mediated by BCP ceramics. The results showed that incubation with BCP ceramics promoted the proliferation of murine bone marrow-derived mesenchymal stem cells (BMSCs) in a time-dependent manner. The resulting cell proliferation was then suppressed by the selective inhibition of either PI3K, AKT, or mTOR signaling activation. Next, we confirmed that BCP ceramics up-regulated the phosphorylation levels of AKT and mTOR in BMSCs, suggesting the ability of BCP ceramics to drive the activation of PI3K/AKT/mTOR signaling in BMSCs. Furthermore, the blockade of PI3K/AKT/mTOR signaling prevented BCP ceramics-induced osteogenic differentiation and pro-angiogenesis of BMSCs by down-regulating the expression of genes encoding OPN, RUNX2 or VEGF. Moreover, the PI3K/AKT/mTOR signaling blockade suppressed stem cell infiltration and new bone formation in the implants following intra-muscular implantation of BCP ceramics in mice. Therefore, our results suggested that PI3K/AKT/mTOR signaling played a critical regulatory role in BCP ceramic-induced osteogenesis.

A two-center study of a combined nomogram based on mammography and MRI to predict ALN metastasis in breast cancer.

OBJECTIVES: To develop and validate a predictive method for axillary lymph node (ALN) metastasis of breast cancer by using radiomics based on mammography and MRI. MATERIALS AND METHODS: A retrospective analysis of 492 women from center 1 (The affiliated Hospital of Qingdao University) and center 2 (Yantai Yuhuangding Hospital) with primary breast cancer from August 2013 to May 2021 was carried out. The radscore was calculated using the features screened based on preoperative mammography and MRI from the training cohort of Center 1 (n = 231), then tested in the validation cohort (n = 99), an internal test cohort (n = 90) from Center 1, and an external test cohort (n = 72) from Center 2. Univariate and multivariate analyses were used to screen for the clinical and radiological characteristics most associated with ALN metastasis. A combined nomogram was established in combination with radscore that predicted the clinicopathological and radiological characteristics. Calibration curves were used to test the effectiveness of the combined nomogram. The receiver operating characteristic (ROC) curve was used to evaluate the performance of the combined nomogram and then compare with the clinical and radiomic models. The decision curve analysis (DCA) value was used to evaluate the combined nomogram for clinical applications. RESULTS: The constructed combined nomogram incorporating the radscore and MRI-reported ALN metastasis status exhibited good calibration and outperformed the radiomics signatures in predicting ALN metastasis (area under the curve [AUC]: 0.886 vs. 0.846 in the training cohort; 0.826 vs. 0.762 in the validation cohort; 0.925 vs. 0.899 in the internal test cohort; and 0.902 vs. 0.793 in the external test cohort). The combination nomogram achieved a higher AUC in the training cohort (0.886 vs. 0.786) and the internal test cohort (0.925 vs. 0.780) and similar AUCs in the validation (0.826 vs. 0.811) and external test (0.902 vs. 0.837) cohorts than the clinical model. CONCLUSION: A combined nomogram based on mammography and MRI can be used for preoperative prediction of ALN metastasis in primary breast cancer.

Broadband radar absorbing metamaterial based on Al @SiO2 conductive composite film.

Artificially designed metamaterial structures can manipulate electromagnetic waves, endowing them with exotic physical properties that are not found in natural materials, such as negative refractive index, superlens, and inverse Doppler effect. These characteristics are widely applied in various engineering and military applications. Due to increasingly complex application environments and innovation in radar detection technology, the combination of broadband absorption performance under thin thickness and efficient preparation methods at low cost is often the focus of research on new generation stealth materials. Here, we propose Al@SiO2 composite conductive film metamaterial (Al@SiO2 CCFM) to achieve wideband absorption of electromagnetic waves. This metamaterial structure combines two resonant units, resulting in three absorption bands in the absorption curve. The results show that the absorption rate of the metamaterial is above 90% in the frequency range of 10.6 GHz to 26.0 GHz. The resonance mechanism between multiple structures is a prerequisite for achieving wideband absorption. The materials Al and SiO2 used in Al@SiO2 CCFM are inexpensive and abundant, and the fabrication method is simple. Therefore, they hold great potential for large-scale applications in the multispectral stealth and electromagnetic shielding field.

Identification and analysis of key genes in adipose tissue for human obesity based on bioinformatics.

BACKGROUND: Obesity is a complex condition that is affected by a variety of factors, including the environment, behavior, and genetics. However, the genetic mechanisms underlying obesity remains poorly elucidated. Therefore, our study aimed at identifying key genes for human obesity using bioinformatics analysis. METHODS: The microarray datasets of adipose tissue in humans were downloaded from the Gene Expression Omnibus (GEO) database. After the selection of differentially expressed genes (DEGs), we used Lasso regression and Support Vector Machine (SVM) algorithm to further identify the feature genes. Moreover, immune cell infiltration analysis, gene set variation analysis (GSVA), GeneCards database and transcriptional regulation analysis were conducted to study the potential mechanisms by which the feature genes may impact obesity. We utilized receiver operating characteristic (ROC) curve to analysis the diagnostic efficacy of feature genes. Finally, we verified the feature genes in cell experiments and animal experiments. The statistical analyses in validation experiments were conducted using SPSS version 28.0, and the graph were generated using GraphPad Prism 9.0 software. The bioinformatics analyses were conducted using R language (version 4.2.2), with a significance threshold of p < 0.05 used. RESULTS: 199 DEGs were selected using Limma package, and subsequently, 5 feature genes (EGR2, NPY1R, GREM1, BMP3 and COL8A1) were selected through Lasso regression and SVM algorithm. Through various bioinformatics analyses, we found some signaling pathways by which feature genes influence obesity and also revealed the crucial role of these genes in the immune microenvironment, as well as their strong correlations with obesity-related genes. Additionally, ROC curve showed that all the feature genes had good predictive and diagnostic efficiency in obesity. Finally, after validation through in vitro experiments, EGR2, NPY1R and GREM1 were identified as the key genes. CONCLUSIONS: This study identified EGR2, GREM1 and NPY1R as the potential key genes and potential diagnostic biomarkers for obesity in humans. Moreover, EGR2 was discovered as a key gene for obesity in human adipose tissue for the first time, which may provide novel targets for diagnosing and treating obesity.

Comparative effects of silicon and silicon nanoparticles on the antioxidant system and cadmium uptake in tomato under cadmium stress.

Cadmium (Cd) pollution is an important threat to agricultural production globally. Silicon (Si) and silicon nanoparticles (Si NPs) can mitigate Cd stress in plants. However, the mechanisms underlying the impacts of Si and Si NPs on Cd resistance, particularly in low-Si accumulators, remain inadequately understood. Accordingly, we conducted a comparative investigation into the roles of Si and Si NPs in regulating the antioxidant system (enzymes and antioxidants) and Cd uptake (influx rate, symplastic and apoplastic pathways) in tomato (a typical low-Si accumulator). The results revealed that Si and Si NPs improved tomato growth under Cd stress, and principal component analysis (PCA) demonstrated that Si NPs were more effective than Si. For oxidative damage, redundancy analysis (RDA) results showed that Si NPs ameliorated oxidative damage in both shoots and roots, whereas Si predominantly alleviated oxidative damage in roots. Simultaneously, Si and Si NPs regulated antioxidant enzymes and nonenzymatic antioxidants with distinct targets and strengths. Furthermore, Si and Si NPs decreased Cd concentration in tomato shoot, root, and xylem sap, while Si NPs induced a more significant decline in shoot and xylem sap Cd. Noninvasive microtest and quantitative estimation of trisodium-8-hydroxy-1,3,6-pyrenetrisulfonic (PTS, an apoplastic tracer) showed that Si and Si NPs reduced the Cd influx rate and apoplastic Cd uptake, while Si NPs induced a more significant reduction. Moreover, Si regulated the expression of genes responsible for Cd uptake (NRAMP2 and LCT1) and compartmentalization (HMA3), while Si NPs reduced the expression of NRAMP2. In conjunction with RDA, the results showed that Si and Si NPs decreased Cd uptake mainly by regulating the symplastic and apoplastic pathways, respectively. Overall, our results indicate that Si NPs is more effective in promoting tomato growth and alleviating oxidative damage than Si in tomato under Cd stress by modulating the antioxidant system and reducing apoplastic Cd uptake.

Psychological Burden of Patients with Head and Neck Cancer Undergoing Radiotherapy and Their Family Caregivers: A Cross-Sectional Survey.

Background: The consequences of Head and neck cancer (HNC) affect both the person who receives the diagnosis and their family caregivers (FCs). Objective: To investigate the psychological status of patients and their FCs, and the burden of the FCs during radiotherapy. Methods: This cross-sectional study was conducted with a questionnaire survey by convenience sampling method. Patients with HNC and their caregivers (both N = 85) from the radiotherapy department of our hospital were recruited between March 2021 and March 2022. The Hospital Anxiety and Depression Scale (HADS), the Chinese version of the Connor and Davidson Resilience Scale (CD-RISC), and the Zarit Burden Interview (ZBI) were used to assess the symptoms of anxiety and depression, psychological resilience, and the impact of care work, emotions and social lives of participants. Pearson's correlation analysis and a Mann-Whitney test were used to analyse the association between the HADS and the CD-RISC scores of the patients. Results: About half of the patients (56.47%) and the caregivers (62.35%) have had anxiety. The average HADS-Anxiety scores, HADS-Depression scores, and CD-RISC scores of the patients with HNC were 7.4±1.9, 6.4±2.2, and 56.8±12.6. The "Strength" and "Resilience" scores of the patients were inversely related to their HADS anxiety scores (p < 0.05). The "Resilience" and "optimism" scores of them were inversely related to HADS depression scores (p < 0.05). The average ZBI score of the caregivers was 23.8±10.1; HADS anxiety scores and HADS depression scores of the caregivers were positively associated with total ZBI scores and individual burden scores (p < 0.05). Conclusion: More than half of patients with HNC undergoing radiotherapy have anxiety, and about a third have depression. The anxiety and depression status of the FCs of patients with HNC undergoing radiotherapy is related to caregiver burden, deserving the attention of clinical medical staff.

Hydroxyapatite nanoparticles promote TLR4 agonist-mediated anti-tumor immunity through synergically enhanced macrophage polarization.

Macrophages represent the most prevalent immune cells in the tumor micro-environment, making them an appealing target for tumor immunotherapy. One of our previous studies showed that hydroxyapatite nanoparticles (HANPs) enhanced Toll-like receptor 4 (TLR4) signal transduction in macrophages. This study was proposed to investigate how HANPs manipulated the phenotype and function of macrophage against 4T1 breast tumors in the presence or absence of MPLA, a low toxic Toll-like receptor 4 (TLR4) agonist. The results demonstrated that the addition of HANPs to MPLA significantly promoted cytokine secretion and macrophage polarization toward a tumoricidal M1 phenotype. Further, the resulting supernatant from HANPs/MPLA co-stimulated macrophages enhanced 4T1 tumor cells apoptosis compared to that from macrophages treated with a single component or PBS control. In particular, we found HANPs elicited immunogenic cell death (ICD) indicated by the increased expression of "danger signals", including HMGB1, CRT and ATP in 4T1 cells. Subsequently, the ICD derivatives-containing supernatant from HANPs-treated 4T1 cells activated macrophage and shifted the phenotype of the cells toward M1 type. Moreover, in a tumor-bearing mice model, HANPs and MPLA synergistically delayed tumor growth compared to PBS control, which was positively associated with the promoted macrophage polarization and ICD induction. Therefore, our findings demonstrated a potential platform to modulate the function of macrophages, and shed a new insight into the mechanism involving the immunomodulatory effect of HANPs for tumor therapy. STATEMENT OF SIGNIFICANCE: Polarizing macrophage toward tumoricidal phenotype by harnessing Toll-like receptor (TLR) agonists has been proven effective for tumor immunotherapy. However, the immunomodulatory potency of TLR agonists is limited due to immune suppression or tolerance associated with TLR activation in immune cells. Herein, we introduced hydroxyapatite nanoparticles (HANPs) to MPLA, a TLR4 agonist. The results demonstrated that the addition of HANPs to MPLA promoted macrophage shift toward tumoricidal M1 phenotype, supported a "hot" tumor transformation, and delayed 4T1 tumor growth. Moreover, we found that HANPs elicited immunogenic cell death that produced "danger" signals from cancer cells thereby further facilitated macrophage polarization. This work is significant to direct the rational design of HANPs coupled with or without TLR agonists for tumor immunotherapy.

Sly-miR398 Participates in Cadmium Stress Acclimation by Regulating Antioxidant System and Cadmium Transport in Tomato (Solanum lycopersicum).

Cadmium (Cd) pollution is one of the major threats in agricultural production, and can cause oxidative damage and growth limitation in plants. MicroRNA398 (miR398) is involved in plant resistance to different stresses, and the post-transcriptional regulation of miR398 on CSDs plays a key role. Here, we report that miR398 was down-regulated in tomato in response to Cd stress. Simultaneously, CSD1 and SOD were up-regulated, with CSD2 unchanged, suggesting CSD1 is involved in miR398-induced regulation under Cd stress. In addition, the role of miR398 in Cd tolerance in tomato was evaluated using a transgenic line overexpressing MIR398 (miR398#OE) in which the down-expression of miR398 was disrupted. The results showed that Cd stress induced more significant growth inhibition, oxidative damage, and antioxidant enzymes disorder in miR398#OE than that in wild type (WT). Moreover, higher Cd concentration in the shoot and xylem sap, and net Cd influx rate, were observed in miR398#OE, which could be due to the increased Cd uptake genes (IRT1, IRT2, and NRAMP2) and decreased Cd compartmentalization gene HMA3. Overall, our results indicate that down-regulated miR398 plays a protective role in tomato against Cd stress by modulating the activity of antioxidant enzymes and Cd uptake and translocation.

Regain flood adaptation in rice through a 14-3-3 protein OsGF14h.

Contemporary climatic stress seriously affects rice production. Unfortunately, long-term domestication and improvement modified the phytohormones network to achieve the production needs of cultivated rice, thus leading to a decrease in adaptation. Here, we identify a 14-3-3 protein-coding gene OsGF14h in weedy rice that confers anaerobic germination and anaerobic seedling development tolerance. OsGF14h acts as a signal switch to balance ABA signaling and GA biosynthesis by interacting with the transcription factors OsHOX3 and OsVP1, thereby boosting the seeding rate from 13.5% to 60.5% for anaerobic sensitive variety under flooded direct-seeded conditions. Meanwhile, OsGF14h co-inheritance with the Rc (red pericarp gene) promotes divergence between temperate japonica cultivated rice and temperate japonica weedy rice through artificial and natural selection. Our study retrieves a superior allele that has been lost during modern japonica rice improvement and provides a fine-tuning tool to improve flood adaptation for elite rice varieties.

A Selective Reduction of Osteosarcoma by Mitochondrial Apoptosis Using Hydroxyapatite Nanoparticles.

Background: In recent years, using hydroxyapatite nanoparticles (HANPs) for tumor therapy attracted increasing attention because HANPs were found to selectively suppress the growth of tumor cells but exhibit ignorable toxicity to normal cells. Purpose: This study aimed to investigate the capacities of HANPs with different morphologies and particle sizes against two kinds of osteosarcoma (OS) cells, human OS 143B cells and rat OS UMR106 cells. Methods: Six kinds of HANPs with different morphologies and particle sizes were prepared by wet chemical method. Then, the antitumor effect of these nanoparticles was characterized by means of in vitro cell experiments and in vivo tumor-bearing mice model. The underlying antitumor mechanism involving mitochondrial apoptosis was also investigated by analysis of intracellular calcium, expression of apoptosis-related genes, reactive oxygen species (ROS), and the endocytosis efficiency of the particles in tumor cells. Results: Both in vitro cell experiments and in vivo mice model evaluation revealed the anti-OS performance of HANPs depended on the concentration, morphology, and particle size of the nanoparticles, as well as the OS cell lines. Among the six HANPs, rod-like HANPs (R-HANPs) showed the best inhibitory activity on 143B cells, while needle-like HANPs (N-HANPs) inhibited the growth of UMR106 cells most efficiently. We further demonstrated that HANPs induced mitochondrial apoptosis by selectively raising intracellular Ca2+ and the gene expression levels of mitochondrial apoptosis-related molecules, and depolarizing mitochondrial membrane potential in tumor cells but not in MC3T3-E1, a mouse pre-osteoblast line. Additionally, the anti-OS activity of HANPs also linked with the endocytosis efficiency of the particles in the tumor cells, and their ability to drive oxidative damage and immunogenic cell death (ICD). Conclusion: The current study provides an effective strategy for OS therapy where the effectiveness was associated with the particle morphology and cell line.

Hydroxyapatite nanoparticles drive the potency of Toll-like receptor 9 agonist for amplified innate and adaptive immune response.

The potency of Toll-like receptor 9 (TLR9) agonist to drive innate immune response was limited due to immune suppression or tolerance during TLR9 signaling activation in immune cells. Herein we addressed this problem by introducing hydroxyapatite nanoparticles (HANPs) to CpG ODN (CpG), a TLR9 agonist. The study revealed that HANPs concentration and duration-dependently reprogramed the immune response by enhancing the secretion of immunostimulatory cytokines (tumor necrosis factor α (TNFα) or IL-6) while reducing the production of immunosuppressive cytokine (IL-10) in macrophages in response to CpG. Next, the enhanced immune response benefited from increased intracellular Ca2+ in macrophage by the addition of HANPs. Further, we found exposure to HANPs impacted the mitochondrial function of macrophages in support of the synthesis of adenosine triphosphate (ATP), the production of nicotinamide adenine dinucleotide (NAD), and reactive oxygen species (ROS) in the presence or absence of CpG. In vaccinated mice model, only one vaccination with a mixture of CpG, HANPs, and OVA, a model antigen, allowed the development of a long-lasting balanced humoral immunity in mice without any histopathological change in the local injection site. Therefore, this study revealed that HANPs could modulate the intracellular calcium level, mitochondrial function, and immune response in immune cells, and suggested a potential combination adjuvant of HANPs and TLR9 agonist for vaccine development. Electronic Supplementary Material: Supplementary material (TEM image, LDH activity, the Ca2+ release in PBS, qRT-PCR analysis, H&E staining, and IL-6 level in the injection site and serum) is available in the online version of this article at 10.1007/s12274-022-4683-x.

Maresin 1 promotes nerve regeneration and alleviates neuropathic pain after nerve injury.

BACKGROUND: Peripheral nerve injury (PNI) is a public health concern that results in sensory and motor disorders as well as neuropathic pain and secondary lesions. Currently, effective treatments for PNI are still limited. For example, while nerve growth factor (NGF) is widely used in the treatment of PNI to promote nerve regeneration, it also induces pain. Maresin 1 (MaR1) is an anti-inflammatory and proresolving mediator that has the potential to regenerate tissue. We determined whether MaR1 is able to promote nerve regeneration as well as alleviating neuropathic pain, and to be considered as a putative therapeutic agent for treating PNI. METHODS: PNI models were constructed with 8-week-old adult male ICR mice and treated with NGF, MaR1 or saline by local application, intrathecal injection or intraplantar injection. Behavioral analysis and muscle atrophy test were assessed after treatment. Immunofluorescence assay was performed to examine the expression of ATF-3, GFAP, IBA1, and NF200. The expression transcript levels of inflammatory factors IL1ß, IL-6, and TNF-α were detected by quantitative real-time RT-PCR. AKT, ERK, mTOR, PI3K, phosphorylated AKT, phosphorylated ERK, phosphorylated mTOR, and phosphorylated PI3K levels were examined by western blot analysis. Whole-cell patch-clamp recordings were executed to detect transient receptor potential vanilloid 1 (TRPV1) currents. RESULTS: MaR1 demonstrated a more robust ability to promote sensory and motor function recovery in mice after sciatic nerve crush injury than NGF. Immunohistochemistry analyses showed that the administration of MaR1 to mice with nerve crush injury reduced the number of damaged DRG neurons, promoted injured nerve regeneration and inhibited gastrocnemius muscle atrophy. Western blot analysis of ND7/23 cells cultured with MaR1 or DRG neurons collected from MaR1 treated mice revealed that MaR1 regulated neurite outgrowth through the PI3K-AKT-mTOR signaling pathway. Moreover, MaR1 dose-dependently attenuated the mechanical allodynia and thermal hyperalgesia induced by nerve injury. Consistent with the analgesic effect, MaR1 inhibited capsaicin-elicited TRPV1 currents, repressed the nerve injury-induced activation of spinal microglia and astrocytes and reduced the production of proinflammatory cytokines in the spinal cord dorsal horn in PNI mice. CONCLUSIONS: Application of MaR1 to PNI mice significantly promoted nerve regeneration and alleviated neuropathic pain, suggesting that MaR1 is a promising therapeutic agent for PNI.

Exposure to hydroxyapatite nanoparticles enhances Toll-like receptor 4 signal transduction and overcomes endotoxin tolerance in vitro and in vivo.

Emerging studies indicate hydroxyapatite nanoparticles (HANPs) exhibit modest immunogenicity to elicit innate immune response which might involve Toll-like receptor 4 (TLR4) activation. This study was proposed to elucidate how HANPs direct over TLR4 signal activity in macrophage in response to TLR4 ligand, lipopolysaccharide (LPS). The present study for the first time reveals that HANPs themselves can induce TLR4 endocytosis and activate pathways both of nuclear factor-kappa B (NF-κB) and interferon regulatory factor 3 (IRF3), which potentially trigger the production of inflammatory cytokine by macrophage. Further, HANPs dose-dependently reprogram over LPS driven TLR4 signaling transduction in macrophage, leading to synergistically augmented innate immune response. In particular, HANPs synergize with LPS to promote macrophage polarization toward M1 phenotype. Moreover, HANPs abrogate the endotoxin tolerance in macrophages by restoring the production of inflammatory cytokines from macrophage in response to secondary LPS stimulation, and enhance the responsiveness of the body to LPS re-challenge in the endotoxin tolerance mice model. Therefore, this study sheds a new light on the mechanism by which HANPs drive the innate immune response, and offers a powerful strategy to potentiate LPS mediated TLR4 signaling activation in macrophage. STATEMENT OF SIGNIFICANCE: In recent years, increasing attention has been given to hydroxyapatite nanoparticles (HANPs) on how they interact with immune cells for achieving appropriate biological effect such as bone tissue repair, soft tissue filler, tumor treatment, vaccine delivery, et al. This study indicated HANPs can induce TLR4 signaling activation. In the further, HANPs dose-dependently synergize with LPS to program over LPS induced TLR4 signaling transduction in macrophage, to favor macrophage polarizing toward M1 phenotype, as well as to abrogate immune tolerance in macrophage in response to repeated LPS stimulation. This work opens a window for the intrinsic mechanism of HANPs to drive immune response and facilitate to direct the rational use or design of HANPs for their better biomedical application.

Weedy Rice as a Novel Gene Resource: A Genome-Wide Association Study of Anthocyanin Biosynthesis and an Evaluation of Nutritional Quality.

The pericarp color of rice grains is an important agronomic trait affected by domestication, and the color pigment, anthocyanin, is one of the key determinants of rice nutritional quality. Weedy rice, also called red rice because its pericarp is often red, may be a novel gene resource for the development of new rice. However, the genetic basis and nutritional quality of anthocyanin are poorly known. In this study, we used a genome-wide association study (GWAS) to find novel and specific QTLs of red pericarp in weedy rice. The known key gene site of red pericarp Rc was detected as the common genetic basis of both weedy and cultivated rice, and another 13 associated signals of pericarp color that were identified may contribute specifically to weedy rice pericarp color. We then nominated three pericarp color genes that may contribute to weedy rice divergence from cultivated rice based on selection sweep analysis. After clarifying the distribution and growth dynamics of pigment in weedy rice caryopsis, we compared its nutritional quality with cultivated rice. We found that sampled weedy rice pericarps had much greater quantities of anthocyanin, beneficial trace elements, free amino acids, and unsaturated fatty acids than the cultivated rice. In conclusion, the gene resources and novel genetic systems of rice anthocyanin biosynthesis explored in this study are of great value for the development of nutritious, high anthocyanin content rice.