A distributed auditory network mediated by pontine central gray underlies ultra-fast awakening in response to alerting sounds.

Sleeping animals can be woken up rapidly by external threat signals, which is an essential defense mechanism for survival. However, neuronal circuits underlying the fast transmission of sensory signals for this process remain unclear. Here, we report in mice that alerting sound can induce rapid awakening within hundreds of milliseconds and that glutamatergic neurons in the pontine central gray (PCG) play an important role in this process. These neurons exhibit higher sensitivity to auditory stimuli in sleep than wakefulness. Suppressing these neurons results in reduced sound-induced awakening and increased sleep in intrinsic sleep/wake cycles, whereas their activation induces ultra-fast awakening from sleep and accelerates awakening from anesthesia. Additionally, the sound-induced awakening can be attributed to the propagation of auditory signals from the PCG to multiple arousal-related regions, including the mediodorsal thalamus, lateral hypothalamus, and ventral tegmental area. Thus, the PCG serves as an essential distribution center to orchestrate a global auditory network to promote rapid awakening.

Hepatic arterial infusion of GEMOX plus systemic gemcitabine chemotherapy combined with lenvatinib and PD-1 inhibitor in large unresectable intrahepatic cholangiocarcinoma.

PURPOSE: To assess the efficacy and safety of hepatic arterial infusion chemotherapy (HAIC) of gemcitabine and oxaliplatin (GEMOX) plus systemic gemcitabine chemotherapy (GEM-SYS) in combination with lenvatinib and programmed cell death protein-1 (PD-1) inhibitor for patients with large unresectable intrahepatic cholangiocarcinoma (uICC). METHODS: From November 2019 to December 2022, 21 large uICC patients who underwent GEMOX-HAIC (Day 1) and GEM-SYS (Day 8) (3w/cycle) combined with lenvatinib and PD-1 inhibitor were retrospectively enrolled. Local tumor response, progression-free survival (PFS), overall survival (OS), and adverse events (AEs) were analyzed. Tumor response was assessed by the Response Evaluation Criteria in Solid Tumors (RECIST) version 1.1. AEs were evaluated by the common terminology criteria for adverse events (CTCAE) version 5.0. RESULTS: After a median follow-up duration of 16.0 months (range 5-43.5 months), 17 patients had died. The median OS was 19.5 months (range 9-43.5 months), and the median PFS was 6.0 months (range 2.5-38.5 months). The 1-, 2-, and 3-year OS rates were 71.4 %, 42.9 %, and 19.0 %, respectively. The 1-, 2-, and 3-year PFS rates were 33.3 %, 19.0 %, and 9.5 %, respectively. Complete response, partial response, stable disease, and progressive disease were observed in 0 (0 %), 11 (52.3 %), 5 (23.8 %), and 5 (23.8 %) patients, respectively. The disease control rate and objective response rate were 76.1 % and 52.3 %, respectively. None of the enrolled patients experienced grade 5 AEs. CONCLUSIONS: GEMOX-HAIC plus GEM-SYS in combination with lenvatinib and PD-1 inhibitor was effective and well tolerated for patients with large uICC.

Elucidating the regulatory role of long non-coding RNAs in drought stress response during seed germination in leaf mustard.

Leaf mustard (Brassica juncea L. Czern & Coss), an important vegetable crop, experiences pronounced adversity due to seasonal drought stress, particularly at the seed germination stage. Although there is partial comprehension of drought-responsive genes, the role of long non-coding RNAs (lncRNAs) in adjusting mustard's drought stress response is largely unexplored. In this study, we showed that the drought-tolerant cultivar 'Weiliang' manifested a markedly lower base water potential (-1.073 MPa vs -0.437 MPa) and higher germination percentage (41.2% vs 0%) than the drought-susceptible cultivar 'Shuidong' under drought conditions. High throughput RNA sequencing techniques revealed a significant repertoire of lncRNAs from both cultivars during germination under drought stress, resulting in the identification of 2,087 differentially expressed lncRNAs (DELs) and their correspondingly linked 12,433 target genes. It was noted that 84 genes targeted by DEL exhibited enrichment in the photosynthesis pathway. Gene network construction showed that MSTRG.150397, a regulatory lncRNA, was inferred to potentially modulate key photosynthetic genes (Psb27, PetC, PetH, and PsbW), whilst MSTRG.107159 was indicated as an inhibitory regulator of six drought-responsive PIP genes. Further, weighted gene co-expression network analysis (WGCNA) corroborated the involvement of light intensity and stress response genes targeted by the identified DELs. The precision and regulatory impact of lncRNA were verified through qPCR. This study extends our knowledge of the regulatory mechanisms governing drought stress responses in mustard, which will help strategies to augment drought tolerance in this crop.

Exosomal miRNA-146a-5p Derived from Senescent Hepatocellular Carcinoma Cells Promotes Aging and Inhibits Aerobic Glycolysis in Liver Cells via Targeting IRF7.

Hepatocellular carcinoma (HCC) is a major global health challenge. Chemotherapy can cause HCC cells to become senescent. Senescent HCC cells play an important role in inhibiting or promoting cancer by producing extracellular vesicles with a senescence-associated secretory phenotype (EV-SASP). miRNA can be strongly upregulated in EV-SASP during the aging process and can substantially alter the phenotypic characteristics of cells. MiRNA microarray analysis revealed that miRNA-146a-5p was highly expressed in oxaliplatin- and H2O2-induced senescent Huh7 cells, and RT‒PCR confirmed its significant upregulation in exosomes. The transcriptome sequencing results of Huh7 cells overexpressing miRNA-146a-5p suggested that miRNA-146a-5p could regulate HCC cell glycolysis. Subsequently, a dual luciferase assay was used to verify whether miRNA-146a-5p can interact with IRF7 to promote aging. The key functions of miRNA-146a-5p and IRF7 in aerobic glycolysis in liver cancer cells were determined through experiments analyzing glucose uptake, lactate production, the oxygen consumption rate (OCR) and the proton efflux rate (PER). Subsequently, the regulatory effect of IRF7 on the key glycolytic gene PFKL was confirmed through luciferase reporter assays. The western blot experiment results showed that miR-146a-5p can activate CHK2 and p53 phosphorylated proteins by targeting IRF7, and upregulate p21 protein. Overexpression of miRNA-146a-5p effectively inhibited the aerobic glycolytic function of HCC cells. Moreover, silencing IRF7 effectively inhibited aerobic glycolysis. MiR-146a-5p. MiR-146a-5p can activate the phosphorylation of CHK2 phosphorylation protein and its downstream protein p53 by targeting IRF7, and the activated p53 upregulates the expression of p21. Our study revealed that exosomal miRNA-146a-5p produced by aging HCC cells, can inhibit HCC cell proliferation through inhibiting aerobic glycolysis and promote HCC cell aging by activating CHK2/p53/p21 signaling way by targeting IRF7.

An oncogenic role of lncRNA SNHG1 promotes ATG7 expression and autophagy involving tumor progression and sunitinib resistance of Renal Cell Carcinoma.

Renal cell carcinoma (RCC) is a malignant tumor with high incidence in adult kidney. Long non-coding RNAs (lncRNAs) have recently been recognized as important regulators in the development of RCC. However, whether lncRNA SNHG1 is associated with RCC progression remains to be elucidated. Here, the role of SNHG1 in RCC autophagy and sunitinib resistance was evaluated. Expression of SNHG1 in RCC tissues and cells was assessed using RT-qPCR. Western blot was utilized to measure the levels of autophagy-related molecules and ATG7. RNA pull-down and RIP assays were performed to confirm the molecular axis between SNHG1/PTBP1/ATG7. Cell proliferation, migration, invasion and apoptosis were analyzed by CCK-8, EdU, transwell and flow cytometry, respectively. The subcellular localization of SNHG1 was determined by an intracellular fractionation assay. The fluorescence intensity of GFP-LC3 autophagosome in RCC cells was detected. IHC staining was performed to test ATG7 expression in tumor tissues from nude mice. Here, a positive correlation of upregulated SNHG1 with poor prognosis of RCC patients was observed in RCC tissues and cells. SNHG1 knockdown suppressed tumor growth and reversed sunitinib resistance and autophagy of RCC cells. Additionally, SNHG1 was found to directly bind to PTBP1, thereby positively regulating ATG7 expression. Furthermore, we verified that SNHG1 mediated the malignant behavior of RCC cells through the PTBP1/ATG7 axis. To sum up, SNHG1 regulates RCC cell autophagy and sunitinib resistance through the PTBP1/ATG7 axis, which highlights a promising therapeutic target for RCC treatment.

Identification and Validation of Nicotinamide Metabolism-Related Gene Signatures as a Novel Prognostic Model for Hepatocellular Carcinoma.

Background: Nicotinamide (NAM+) regulates redox and metabolic activities in the mitochondria. The intention of the research was to identify key genes that relate to nicotinamide in hepatocellular carcinoma (HCC). Methods: Relevant clinical information were collected as well as RNA-seq data using the Cancer Genome Atlas (TCGA) database. Differential analysis was used to discover the genes that were differently expressed. On the key genes associated with NAM, functional enrichment analysis was carried out. Next, receiver operating characteristic (ROC) and prognosis Kaplan-Meier (K-M) curve analyses were used to evaluate the importance of important gene expression, respectively. The immune cell signatures were estimated using the CIBERSORT algorithm. Finally, the anticancer impact of NAM on HCC was experimentally confirmed, and important genes NADSYN1 and NT5C were validated at the protein level in clinical specimens. Results: Six prognostic key genes (NAXE, NADSYN1, NT5C, NT5C3A, PNP and NT5E) were identified. There is an association between the level of key gene expression and the clinical prognosis. Four key genes (NAXE, NADSYN1, NT5C and NT5C3A) have statistical significance of survival prognosis. Finally, the expression of NAM-related genes and the inhibitory effect of NAM on HCC were verified by experiments. Conclusion: The study first found some Nicotinamide metabolism-related differentially expressed genes (NMRDEGs) that are related to HCC can contribute to predicting survival and monitoring the treatment.

Toripalimab Plus Bevacizumab as First-line Treatment for Advanced Hepatocellular Carcinoma: A Prospective, Multicenter, Single-Arm, Phase II Trial.

PURPOSE: This phase II, multicenter, prospective, single-arm study aimed to evaluate the efficacy and safety of toripalimab plus bevacizumab for treating advanced hepatocellular carcinoma (HCC). PATIENTS AND METHODS: Treatment-naïve patients with advanced HCC received toripalimab 240 mg plus bevacizumab 15 mg/kg every 3 weeks. The primary endpoints included safety and tolerability and objective response rate (ORR) assessed by the investigator per Response Evaluation Criteria in Solid Tumors (RECIST) version 1.1. RESULTS: Fifty-four patients were enrolled between April 17, 2020, and December 11, 2020. As assessed by the investigator according to RECIST v1.1, the ORR was 31.5% [95% confidence interval (CI), 19.5-45.6] and the lower bound of the 95% CI was above the prespecified boundary of 10%. The independent review committee (IRC) assessed ORR according to the modified RECIST (mRECIST), which was 46.3% (95% CI, 32.6-60.4). The median progression-free survival was 8.5 (95% CI, 5.5-11.0) and 9.8 months (95% CI, 5.6 to not evaluable) as assessed by the investigator according to RECIST v1.1 and IRC according to mRECIST criteria, respectively. The median overall survival (OS) was not reached, and the 12- and 24-month OS rates were 77.3% and 63.5%, respectively. Grade 3 or higher treatment-emergent adverse events (TEAEs) occurred in 27 patients (50.0%). The most common TEAEs were proteinuria (59.3%), hypertension (38.9%), increased aspartate aminotransferase (33.3%), increased amylase (29.6%), decreased platelet count (27.8%), and increased bilirubin levels (27.8%). CONCLUSIONS: Toripalimab plus bevacizumab showed a favorable efficacy and safety profile, supporting further studies on this combination regimen as a first-line treatment for advanced HCC.

RPL9 acts as an oncogene by shuttling miRNAs through exosomes in human hepatocellular carcinoma cells.

The exosomal pathway is an essential mechanism that regulates the abnormal content of microRNAs (miRNAs) in hepatocellular carcinoma (HCC). The directional transport of miRNAs requires the assistance of RNA­binding proteins (RBPs). The present study found that RBPs participate in the regulation of miRNA content through the exosomal pathway in HCC cells. First, differential protein expression profiles in the serum exosomes of patients with HCC and benign liver disease were detected using mass spectrometry. The results revealed that ribosomal protein L9 (RPL9) was highly expressed in serum exosomes of patients with HCC. In addition, the downregulation of RPL9 markedly suppressed the proliferation, migration and invasion of HCC cells and reduced the biological activity of HCC­derived exosomes. In addition, using miRNA microarrays, the changes in exosomal miRNA profiles in HCC cells caused by RPL9 knockdown were examined. miR­24­3p and miR­185­5p were most differentially expressed, as verified by reverse transcription­quantitative PCR. Additionally, using RNA immunoprecipitation, it was found that RPL9 was directly bound to the two miRNAs and immunofluorescence assays confirmed that RPL9 was able to carry miRNAs into recipient cells via exosomes. Overexpression of miR­24­3p in cells increased the accumulation of miR­24­3p in exosomes and simultaneously upregulated RPL9. Excessive expression of miR­24­3p in exosomes also increased their bioactivity. Exosome­mediated miRNA regulation and transfer require the involvement of RBPs. RPL9 functions as an oncogene, can directly bind to specific miRNAs and can be co­transported to receptor cells through exosomes, thereby exerting its biological functions. These findings provide a novel approach for modulating miRNA profiles in HCC.

Cancer-Associated Fibroblasts Boost Tumorigenesis of Clear Cell Renal Cell Carcinoma via Exosome-Mediated Paracrine SNHG1.

Despite the dominant roles of cancer-associated fibroblasts (CAFs) have attached much attention in tumorigenesis, the CAFs-derived molecular determinants that regulate renal cell carcinoma (RCC) development remains elusive. Our previous study uncovered an oncogenic SNHG1 in the immune escape of RCC, whereas CAFs-derived exosomes could be a source accounting for increasing SNHG1 in RCC cells, this is still a mystery. The obtained CAFs and normal fibroblast (NFs) from fresh RCC and adjacent tissues were firstly identified using western blot and immunofluorescent staining. The enrichment of SNHG1 was validated by RT-qPCR. CAFs-derived exosomes were isolated from conditioned medium using ultracentrifugation method and ExoQuick-TC system. The internalization of exosomes, transfer of SNHG1, was measured by immunofluorescence. Regulation of conditioned medium or exosomal SNHG1 from CAFs on RCC biological functions was evaluated by CCK-8, EdU incorporation, colony formation, and transwell assays to assess the RCC cell proliferation, migration, and invasion. SNHG1 was significantly upregulated in CAFs isolated from RCC stroma. Exosomes derived from CAFs transferred SNHG1 to RCC cells and resulted in an increased SNHG1 expression in RCC cells. The exosomes excreted by CAFs promoted RCC cell proliferation, migration, and invasion, whereas the promotion effect of CAFs-exosomes on RCC progression was attenuated by SNHG1 knockdown. The present study revealed a new mechanism of exosomal SNHG1 extracted from CAFs enhanced RCC progression and may provide a potential target for the treatment of RCC.

Integrative analysis of lactylation-related genes and establishment of a novel prognostic signature for hepatocellular carcinoma.

BACKGROUND: Lactylation has been found to involve in regulating many types of biological process in cancers. However, research on lactylation-related genes in predicting the prognosis of hepatocellular carcinoma (HCC) remains limited. METHODS: The differential expression of lactylation-related genes (EP300 and HDAC1-3) in pan-cancer were examined in public databases. HCC patient tissues were obtained for mRNA expression and lactylation level detection by RT-qPCR and western blotting. Transwell migration assay, CCK-8 assay, EDU staining assay and RNA-seq were performed to verify the potential function and mechanisms in HCC cell lines after lactylation inhibitor apicidin treatment. lmmuCellAI, quantiSeq, xCell, TIMER and CIBERSOR were used to analyze the correlation between transcription levels of lactylation-related genes and immune cell infiltration in HCC. Risk model of lactylation-related genes was constructed by LASSO regression analysis, and prediction effect of the model was evaluated. RESULT: The mRNA levels of lactylation-related genes and lactylation levels were higher in HCC tissues than normal samples. The lactylation levels, cell migration, and proliferation ability of HCC cell lines were suppressed after apicidin treatment. The dysregulation of EP300 and HDAC1-3 was associated with proportion of immune cell infiltration, especially B cell. Upregulation of HDAC1 and HDAC2 was closely associated with poorer prognosis. Finally, a novel risk model, based on HDAC1 and HDAC2, was developed for prognosis prediction in HCC. CONCLUSION: HDAC1 and HDAC2 are expected to become new biomarkers for HCC. Risk scoring model based on HDAC1 and HDAC2 can be used to predict the prognosis of HCC patients.

Transcriptome profile analysis of Indian mustard (Brassica juncea L.) during seed germination reveals the drought stress-induced genes associated with energy, hormone, and phenylpropanoid pathways.

Indian mustard (Brassica juncea L. Czern and Coss) is an important oil and vegetable crop frequently affected by seasonal drought stress during seed germination, which retards plant growth and causes yield loss considerably. However, the gene networks regulating responses to drought stress in leafy Indian mustard remain elusive. Here, we elucidated the underlying gene networks and pathways of drought response in leafy Indian mustard using next-generation transcriptomic techniques. Phenotypic analysis showed that the drought-tolerant leafy Indian mustard cv. 'WeiLiang' (WL) had a higher germination rate, antioxidant capacity, and better growth performance than the drought-sensitive cv. 'ShuiDong' (SD). Transcriptome analysis identified differentially expressed genes (DEGs) in both cultivars under drought stress during four germination time points (i.e., 0, 12, 24, and 36 h); most of which were classified as drought-responsive, seed germination, and dormancy-related genes. In the Kyoto Encyclopedia of Genes and Genome (KEGG) analyses, three main pathways (i.e., starch and sucrose metabolism, phenylpropanoid biosynthesis, and plant hormone signal transduction) were unveiled involved in response to drought stress during seed germination. Furthermore, Weighted Gene Co-expression Network Analysis (WGCNA) identified several hub genes (novel.12726, novel.1856, BjuB027900, BjuA003402, BjuA021578, BjuA005565, BjuB006596, novel.12977, and BjuA033308) associated with seed germination and drought stress in leafy Indian mustard. Taken together, these findings deepen our understanding of the gene networks for drought responses during seed germination in leafy Indian mustard and provide potential target genes for the genetic improvement of drought tolerance in this crop.

Fluorescent Laparoscopic Central Hepatectomy for Liver Cancer Using Indocyanine Green Negative Staining.

Laparoscopic hepatectomy is an important treatment method for liver cancer. In the past, the resection boundary was usually determined by intraoperative ultrasound, important vascular structures, and surgeon experience. With the development of anatomical hepatectomy, visual surgery technology has gradually been applied to this type of surgery, particularly indocyanine green (ICG)-guided anatomical hepatectomy. As ICG can be specifically ingested by hepatocytes and used for fluorescence tracing, negative staining techniques have been applied according to different tumor positions. Under ICG fluorescent guidance, the surface boundary and deep resection plane can be more accurately displayed during liver resection. Thus, the tumor-bearing liver segment can be anatomically removed, which helps to avoid damage to important vessels and reduce ischemia or congestion of the remaining liver tissue. Finally, the incidence of postoperative biliary fistula and liver dysfunction is reduced; therefore, a better prognosis is obtained after the resection of liver cancer. Centrally located liver cancer is usually defined as a tumor located at segments 4, 5, or 8 that requires resection of the middle section of the liver. These are among the most difficult hepatectomies to perform because of the large surgical wounds and multiple vessel transections. Based on the specific tumor location, we formulated the required resection ranges by designing personalized fluorescent staining strategies. By completing anatomical resection based on the portal territory, this work aims to achieve the best therapeutic effect.

Spatial heterogeneity and Immune infiltration of cellular lysosomal pathways reveals a new blueprint for tumor heterogeneity in esophageal cancer.

Background: Esophageal squamous cell carcinoma (ESCC) is a common Malignant tumor of digestive tract which have a potential association with lysosomal pathway. The purpose of this study was to explore the correlation between lysosome pathway and immune infiltration of ESCC. Methods: The cell type annotation of ESCC patients and the distribution of their gene markers were analyzed by single cell data. They were also grouped according to the expression of lysosomal pathways. Gene set variation analysis (GSVA) enriched pathway scoring, Cellchat cell communication was performed to demonstrate the tumour-associated pathway scores and interactions of different cell populations. Relevant differential genes were screened, prognostic risk markers were constructed and direct associations of lysosomal pathway-related gene risk scores with immune infiltration and tumour treatment drug sensitivity were assessed by algorithms. In cellular experiments, qPCR and flow cytometry were used to assess the role of the lysosomal pathway gene-MT1X on tumour cell development. Results: ESCC single cell data were annotated into 7 Cluster clusters by t-sne downscaling analysis. Cellchat analysis revealed that the "MIF" cellular communication network is the main communication mode of the lysosomal pathway in ESCC cells. The lysosomal pathway genetic risk model was found to be significantly different from ESCC prognosis in both the training and validation groups. The lysosome pathway gene risk model was associated with treatment resistance in ESCC patients using oncopredict R package. The correlation between the expression of lysosomal-DEG and tumour immune infiltration and immune cell types by the MCPcounter method. Cellular assays showed that the lysosomal pathway gene MT1X was less expressed in oesophageal cancer cells than in normal oesophageal epithelial cells. Knockdown of MT1X significantly promoted the growth rate of oesophageal cancer cells. Conclusion: Based on the single cell sequencing technology and transcriptomic analysis, we confirmed that there is a close association between the lysosomal pathway and the immune infiltration and treatment sensitivity of ESCC, which may be a potential target for a new direction of ESCC therapy.

Glutamatergic and GABAergic neurons in pontine central gray mediate opposing valence-specific behaviors through a global network.

Extracting the valence of environmental cues is critical for animals' survival. How valence in sensory signals is encoded and transformed to produce distinct behavioral responses remains not well understood. Here, we report that the mouse pontine central gray (PCG) contributes to encoding both negative and positive valences. PCG glutamatergic neurons were activated selectively by aversive, but not reward, stimuli, whereas its GABAergic neurons were preferentially activated by reward signals. The optogenetic activation of these two populations resulted in avoidance and preference behavior, respectively, and was sufficient to induce conditioned place aversion/preference. Suppression of them reduced sensory-induced aversive and appetitive behaviors, respectively. These two functionally opponent populations, receiving a broad range of inputs from overlapping yet distinct sources, broadcast valence-specific information to a distributed brain network with distinguishable downstream effectors. Thus, PCG serves as a critical hub to process positive and negative valences of incoming sensory signals and drive valence-specific behaviors with distinct circuits.

Overexpression of soybean microRNA156b enhanced tolerance to phosphorus deficiency and seed yield in Arabidopsis.

microRNAs (miRNAs) are endogenous small RNAs that are key regulatory factors participating in various biological activities such as the signaling of phosphorus deficiency in the plant. Previous studies have shown that miR156 expression was modulated by phosphorus starvation in Arabidopsis and soybean. However, it is not clear whether the over-expression of soybean miR156b (GmmiR156b) can improve a plant's tolerance to phosphorus deficiency and affect yield component traits. In this study, we generated Arabidopsis transgenic lines overexpressing GmmiR156b and investigated the plant's response to phosphorus deficiency. Compared with the wild type, the transgenic Arabidopsis seedlings had longer primary roots and higher phosphorus contents in roots under phosphorus-deficit conditions, but lower fresh weight root/shoot ratios under either phosphorus-deficient or sufficient conditions. Moreover, the GmmiR156b overexpression transgenic lines had higher phosphorus content in shoots of adult plants and grew better than the wide type under phosphorus-deficient conditions, and exhibited increased seed yields as well as strong pleiotropic developmental morphology such as dwarfness, prolonged growth period, bushy shoot/branching, and shorter silique length, suggesting that the transgenic lines were more tolerant to phosphorus deficiency. In addition, the expression level of four SQUAMOSA PROMOTER BINDING PROTEIN LIKE (SPL) genes (i.e., AtSPL4/5/6/15) were markedly suppressed in transgenic plants, indicating that they were the main targets negatively regulated by GmmiR156b (especially AtSPL15) and that the enhanced tolerance to phosphorus deficiency and seed yield is conferred mainly by the miR156-mediated downregulation of AtSPL15.

p62 Promotes Malignancy of Hepatocellular Carcinoma by Regulating the Secretion of Exosomes and the Localization of ß-Catenin.

BACKGROUND: p62 is a multi-domain protein and participates in a variety of cellular biological activities. p62 is also related to tumor malignancy. However, the underlying molecular mechanism of p62 regulating the progression of hepatocellular carcinoma (HCC) remains unclear. METHODS: The expression levels of p62 in HCC tissues and adjacent non-tumor tissues were confirmed using the TCGA dataset and immunohistochemistry. Stable p62-overexpressing HepG2 cells and p62-knockdown MHCC97H cells were established with lentiviral vectors. Cell proliferation, migration, and invasion assays were carried out to investigate the role of p62 in HCC cells and HCC-derived exosomes. The relationship between p62 and ß-catenin was investigated by immunofluorescence and co-immunoprecipitation assays. Male nude mice (BALB/c-nu/nu) were used to establish the xenograft tumors. RESULTS: We found that p62 was significantly upregulated in HCC, and a high level of p62 indicated the promotion of malignancy including cell proliferation, migration, and invasion. Exosomes derived from p62-overexpressing HepG2 also demonstrated the ability to promote tumor malignancy. Immunofluorescence and co-immunoprecipitation assays indicated that p62 interacts with ß-catenin and regulates the localization of ß-catenin to affect the intercellular junction. p62 also promotes tumor growth of HCC and down-regulates the expression of ß-catenin in vivo. CONCLUSIONS: The results of this study concluded that p62 promotes the malignancy of HCC by regulating the secretion of exosomes and the localization of ß-catenin. These findings may provide new ideas for the diagnosis and treatment of HCC.

Osteoinductive activity of bisdemethoxycurcumin and its synergistic protective effect with human amniotic mesenchymal stem cells against ovariectomy-induced osteoporosis mouse model.

Osteoporosis is a common disease characterized by skeletal fragility and microarchitectural deterioration. However, existing conventional drugs exhibit limited efficacy and can elicit severe adverse effects; moreover, and novel stem cell-based therapies have not exhibited sufficient therapeutic efficacy. Our hypothesis is that an appropriate osteogenic inducer may improve their therapeutic efficacy. In this study, we found that bisdemethoxycurcumin (BDMC) stimulates the differentiation of human amniotic mesenchymal stem cells (hAMSCs) into osteoblasts without inducing cytotoxicity. Here BDMC enhances calcium deposition in hAMSCs, while promoting the expression of early and late markers of osteoblast differentiation, including ALP, runt-related transcription factor 2, osterix, COL1-α1, osteocalcin, and osteopontin at the transcriptional and translational levels. Mechanistically, BDMC was found to activate the JAK2/STAT3 pathway; whereas AG490 (JAK2/STAT3 pathway inhibitor) inhibited BDMC functioning. Subsequently, we found that the combinatorial therapy of BDMC and hAMSC had a positive synergistic effect on osteoporotic mouse model induced by bilateral ovariectomy, including inhibiting bone loss and bone resorption and improving bone micro-architecture. Moreover, BDMC inhibited production of the bone resorption markers C-terminal telopeptide of type I collagen, and tartrate resistant acid phosphatase, while promoting serum levels of bone formation markers OCN, and procollagen I N-terminal propeptide. BDMC also improved liver and kidney function in osteoporotic mouse model. Collectively, BDMC improved osteoporosis by enhancing hAMSC osteogenesis and exhibited a protective effect on liver and kidney function in an osteoporotic mouse model. Hence, BDMC may serve as an effective adjuvant, and combined therapy with hAMSCs is a promising new approach toward osteoporosis treatment.

Case Report: Hepatic Artery Infusion Chemotherapy After Stage I ALPPS in a Patient With Huge HCC.

Associating liver partition and portal vein ligation for staged hepatectomy (ALPPS) can induce rapid hypertrophy of the liver remnant. However, with a background of liver cirrhosis or other chronic liver diseases, patients with a huge hepatocellular carcinoma (HCC) may sometimes face insufficiency of hepatocellular regeneration after associating liver partition and portal vein ligation for staged hepatectomy (ALPPS). Herein, we report a 56-year-old male with a vast HCC (13.3 × 8.5 × 13 cm) whose ratio of the future liver remnant (FLR)/standard liver volume (SLV) was 28.7% when the disease was first diagnosed. Inadequate hypertrophy of FLR was shown in postoperative volumetric assessment a month after stage I ALPPS. After multidisciplinary team discussion (MDT), the patient was decided to follow three courses of hepatic arterial infusion chemotherapy (HAIC) with oxaliplatin, fluorouracil, and leucovorin (FOLFOX4). The last HAIC was performed together with transhepatic arterial embolization (TAE). Finally, ratio of the FLR/SLV increased from 28.7% to 40% during three-month intervals, meeting the requirements of the surgery. Stage II ALPPS, right trisectionectomy, was then successfully performed. There was no recurrence at half years of follow-up. In our case, HAIC seems to be more potent than transcatheter arterial chemoembolization (TACE) in maintaining the hyperplasia of the liver remnant, reducing tumor load, and preventing tumor progression in patients with a large HCC during ALPPS procedure. HAIC, following the first step of ALPPS, a pioneering treatment modality aiming for inadequate hypertrophy of FLR induced by ALPPS, could be an alternative procedure for patients with a vast HCC in clinical practice.

LncRNA SNHG1 regulates immune escape of renal cell carcinoma by targeting miR-129-3p to activate STAT3 and PD-L1.

Immune escape of renal cell carcinoma (RCC) impacts patient survival. However, the molecular mechanism of long noncoding RNA (lncRNA) small nucleolar RNA host gene 1 (SNHG1) in RCC immune escape remains unclear. Quantitative real-time PCR and western blotting results revealed that the expression of lncRNA SNHG1 and STAT3 were upregulated in RCC tissues and cells and that the expression of miR-129-3p was downregulated. Enzyme-linked immunosorbent assay results revealed the increased levels of immune-related factors (interferon-γ, tumour necrosis factor α, and interleukin-2) in RCC tissues. SNHG1 knockdown or miR-129-3p overexpression inhibited the proliferation and invasion of A498 and 786-O cells, while the proliferation and cytotoxicity of CD8+ T cells increased, which promoted the secretion of immune-related factors. STAT3 overexpression decreased the protective effect of miR-129-3p overexpression on RCC cell immune escape. In addition, miR-129-3p knockdown and STAT3 overexpression decreased the protective effect of lncRNA SNHG1 knockdown on RCC cell immune escape. In addition, PD-L1 expression was downregulated after lncRNA SNHG1 knockdown but upregulated after miR-129-3p knockdown and STAT3 overexpression. Dual-luciferase assays showed that lncRNA SNHG1 targets miR-129-3p, and miR-129-3p targets STAT3. RNA pull-down and RNA immunoprecipitation assays verified the regulatory relationship between SNHG1 and STAT3. In vivo, shSNHG1 prolonged the overall survival of RCC tumour model mice and inhibited RCC tumour growth and immune escape but increased CD8+ T cell infiltration in mice. Our findings provide an experimental basis for elucidating the molecular mechanisms of immune escape by RCC and reveal a novel target to treat this disease.