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Heat shock protein 70 (HSP70) is a class of important molecular chaperones that are involved in protein folding, stabilization, and maturation, and play a vital role in plant growth and response to environmental stress. Apple trees frequently suffer from different-degree salt stress, which seriously affects their growth, quality, and yield. However, whether HSP70 genes are involved in salt tolerance is unexplored in apple. In this study, 67 MdHSP70 genes were identified and unevenly distributed on 17 apple chromosomes. Gene structure and protein motif analysis revealed that MdHSP70 genes in the same subgroup have similar intron phase and motif organization, further supporting the phylogenetic results. RNA-seq analysis showed the expression level of nine of 67 MdHSP70 genes was induced by salt stress. Subsequent qRT-PCR analysis revealed that MdHSP70-38 was dramatically up-regulated in response to salt stress. The overexpression of MdHSP70-38 in transgenic tobacco and apple improved salt stress tolerance, which was associated with less electrolyte leakage and malondialdehyde (MDA), as well as diminished accumulation of hydrogen peroxide (H 2 O 2 ) and superoxide radicals (O 2 - ). Our findings demonstrated that MdHSP70-38 played a positive regulatory role in salt tolerance in tobacco and apple, and provided a promising candidate gene in genetic applications for improving salt tolerance., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Aging represents a significant risk factor for compromised tissue function and the development of chronic diseases in the human body. This process is intricately linked to oxidative stress, with HClO serving as a vital reactive oxygen species (ROS) within biological systems due to its strong oxidative properties. Hence, conducting a thorough examination of HClO in the context of aging is crucial for advancing the field of aging biology. In this work, we successfully developed a fluorescent probe, OPD , tailored specifically for detecting HClO in senescent cells and in vivo. Impressively, OPD exhibited a robust reaction with HClO, showcasing outstanding selectivity, sensitivity, and photostability. Notably, OPD effectively identified HClO in senescent cells for the first time, confirming that DOX- and ROS-induced senescent cells exhibited higher HClO levels compared to uninduced normal cells. Additionally, in vivo imaging of zebrafish demonstrated that d-galactose- and ROS-stimulated senescent zebrafish displayed elevated HClO levels compared to normal zebrafish. Furthermore, when applied to mouse tissues and organs, OPD revealed increased fluorescence in the organs of senescent mice compared to their nonsenescent counterparts. Our findings also illustrated the probe's potential for detecting changes in HClO content pre- and post-aging in living mice. Overall, this probe holds immense promise as a valuable tool for in vivo detection of HClO and for studying aging biology in live organisms.

Background: Anaplastic thyroid carcinoma (ATC) is a highly invasive malignant tumor with a poor prognosis. Traditional treatment methods have not been effective. However, advancements in targeted therapy and immunotherapy in recent years have shed new light on the management of ATC. The aim of this study was to examine the treatment plan and prognostic factors of ATC., Methods: This study conducted a retrospective analysis of ATC patients who received treatment at our institution between 2000 and 2023 to evaluate the clinical characteristics, treatment methods, and factors influencing survival. Survival analysis was performed by the Kaplan‒Meier method and log-rank test, and multivariate analysis was performed using the Cox proportional hazard model., Results: The 6-month and 1-year disease-specific OS rates were 49% and 29%, respectively. The presence of clinical symptoms and the timing of treatment significantly impacted patient prognosis (P < 0.05). Compared with surgery + radiotherapy/chemotherapy and only surgery, targeted therapy and targeted + immunotherapy represented an improved overall survival, The 6-month/1-year survival rates of which were 81%/61% and 91%/73% (P < 0.05), respectively. Multivariate analysis indicated that the symptoms at initial diagnosis, year of presentation, performance status and treatment plan were independent factors affecting the prognosis. The year of presentation (P = 0.048) and the treatment plan (P = 0.038) were significantly meaningful in predicting prognosis., Conclusion: Targeted therapy and targeted+immune therapy can effectively prolong the survival period of ATC patients. Symptoms at initial diagnosis and treatment plan have a significant impact on the prognosis., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

ONOO - , a bioactive molecule, plays a critical role in inflammation-related signaling pathways and pathological mechanisms. Numerous studies have established a direct correlation between elevated ONOO - levels and tumor progression. Therefore, investigating ONOO - levels in inflammation and tumors is of utmost importance. Fluorescence imaging presents a highly sensitive, non-invasive, easily operable, selective, and efficient method for ONOO - detection in situ . In this study, we designed and synthesized a rhodamine-based probe, NRho, which effectively identifies tumors, inflammatory cells, tissues, and organs by detecting ONOO - content. The synthesis process of NRho is simple, yielding a probe with favorable spectral characteristics and rapid response. Our cell imaging analysis has provided novel insights, revealing distinct ONOO - levels among different types of cancer cells, with hepatocellular carcinoma cells exhibiting higher ONOO - content than the others. This observation marks the proposal of such variations in ONOO - levels across cancer cell types. Furthermore, our study has showcased the practicality of our probe in live organ imaging, enabling the identification of tumors from living organs within a brief 5-minute incubation period. Additionally, our findings highlight the rapid detection capability of the probe NRho in various tissue samples, effectively identifying inflammation. This research holds important promise in advancing biomedical research and clinical diagnosis.

Purpose: This study was aimed to examine the global research status and current research hotspots in the field of tendon stem cells. Methods: Bibliometric methods were employed to retrieve relevant data from the Web of Science Core Collection (WOSCC) database. Additionally, Citespace, Vosviewer, SCImago, and Graphad Prism were utilized to analyze the publication status in this field, identify the current research hotspots, and present a mini-review. Results: The most active countries in this field were China and the United States. Notable authors contributing significantly to this research included Lui Pauline Po Yee, Tang Kanglai, Zhang Jianying, Yin Zi, and Chen Xiao, predominantly affiliated with institutions such as the Hong Kong Hospital Authority, Third Military Medical University, University of Pittsburgh, and Zhejiang University. The most commonly published journals in this field were Stem Cells International, Journal of Orthopedic Research, and Stem Cell Research and Therapy. Moreover, the current research hotspots primarily revolved around scaffolds, molecular mechanisms, and inflammation regulation. Conclusion: Tendon stem cells hold significant potential as seed cells for tendon tissue engineering and offer promising avenues for further research Scaffolds, molecular mechanisms and inflammation regulation are currently research hotspots in this field., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Zhang, Shang, Gu, Gu, Wang, Hu, Wu, Zou, Ruan, He, Bao, Zhang, Li and Chen.)

In this paper, a series of alkaline-treated ZSM-22 zeolite samples were prepared by treating the parent ZSM-22 zeolite using NaOH aqueous solution with different concentrations. By investigating the effects of alkaline treatment on the parent ZSM-22 zeolite, we discovered that the alkaline treatment contributed to the reduction of Brønsted acid sites due to the coverage of extra-framework Al on its external surface. In addition, it was found that the alkaline-treated samples were favorable to the improvement of the isobutene yield and selectivity, while these features appeared to be low for the subsequent acid-washed counterparts in the skeletal isomerization reaction of 1-butene. These results indicate that the catalytic performance of ZSM-22 zeolite is related to reduced amounts of Brønsted acid sites in it. To further reveal the reasons for the promoted catalytic performances of the alkaline-treated ZSM-22 series zeolites, we studied the properties of coke deposited on the two series of samples using Raman spectroscopy and thermogravimetric analysis and mass spectrometry (TG/MS-TPO). It was shown that the carbon deposited on the alkaline-treated series samples was mainly distributed at the outer surface, while the coke was distributed to a relatively lesser extent at the exterior surface for the acid-washed series samples. Moreover, by partially passivating outer acid sites of the parent zeolite, the selected alkaline-treated zeolite, and acid-washed zeolite, their isobutene selectivities were all improved with the decrease in outer acid sites. These phenomena confirmed that the improved catalytic performances of the alkaline-treated samples are related to their decreased external Brønsted acid site density, which further demonstrated that the high isobutene yield and selectivity in the skeletal isomerization reaction of 1-butene is realized via the monomolecular reaction pathway of 1-butene., Competing Interests: The authors declare no competing financial interest., (© 2023 The Authors. Published by American Chemical Society.)

Background: Volatile components are important secondary metabolites essential to fruit aroma quality, thus, in the past decades many studies have been extensively performed in clarifying fruit aroma formation. However, aroma components and biosynthesis in the fruit of Binzi (Malus pumila × Malus asiatica), an old local species with attractive aroma remain unknown., Results: We investigated two Binzi cultivars, 'Xiangbinzi' (here named high-fragrant Binzi, 'HFBZ') and 'Hulabin' (here named low-fragrant Binzi, 'LFBZ') by monitoring the variation of volatiles and their precursors by Gas Chromatography-Mass Spectrometer (GC-MS), as well as their related genes by RNA-seq during post-harvest ripening. We firstly confirmed that 'HFBZ' and 'LFBZ' fruit showed respiratory climacteric by detecting respiratory rate and ethylene emission during post-harvest; found that esters were the major aroma components in 'HFBZ' fruit, and hexyl 2-methylbutyrate was responsible for the 'fruity' note and most potent aroma component, followed by ethyl acetate, ethyl butanoate, (E)-2-hexenal, and 1-hexanol. Regarding aroma synthesis, fatty acid metabolism seemed to be more important than amino acid metabolism for aroma synthesis in 'HFBZ' fruit. Based on RNA-seq and quantitative reverse transcription PCR (RT-qPCR), LOX2a, LOX5a, ADH1, and AAT1 genes are pointed to the LOX pathway, which may play a vital role in the aroma formation of 'HFBZ' fruit., Conclusion: Our study firstly investigated the aroma components and related genes of Binzi fruit, and provided an insight into the fragrant nature of Malus species., (© 2022. The Author(s).)

Background: Walnuts are among the most important dry fruit crops worldwide, typically exhibiting green leaves and yellow-brown or gray-yellow seed coats. A specific walnut accession with red leaves and seed coats, 'RW-1', was selected for study because of its high anthocyanin and proanthocyanidin (PA) contents. Anthocyanins and PAs are important secondary metabolites and play key roles in plant responses to biotic and abiotic stresses. However, few studies have focused on the molecular mechanism of anthocyanin biosynthesis in walnuts., Methods: In this study, we determined the anthocyanin and PA components and their contents in different color leaves of 'RW-1' natural hybrid progenies at various developmental stages. Integrated transcriptome and metabolome analyses were used to identify the differentially expressed genes (DEGs) and differentially accumulated metabolites (DAMs). We also performed conjoint analyses on DEGs and DAMs to ascertain the degree pathways, and explore the regulation of anthocyanin and PA biosynthesis., Results: The results of widely targeted metabolome profiling and anthocyanin detection revealed 395 substances, including four PAs and 26 anthocyanins, in red (SR) and green leaves (SG) of 'RW-1' natural hybrid progenies. From the research, the contents of all anthocyanin components in SR were higher than that in SG. Among them, the contents of delphinidin 3-O-galactoside, cyanidin 3-O-galactoside, delphinidin 3-O-arabinoside and cyanidin 3-O-glucoside were significantly higher than others, and they were considered as the main types of anthocyanins. However, nine anthocyanins were detected only in SR. For PAs, the content of procyanidin C1 was higher in SR compared with SG, while procyanidin B1 and procyanidin B3 were higher in SR-1 and SR-3 but downregulated in SR-2 compared with the controls. Furthermore, transcriptome analysis revealed that the expressions of structural genes ( C4H , F3H , F3'5'H , UFGT , LAR and ANR ), three MYBs predicted as the activators of anthocyanin and PA biosynthesis, two MYBs predicted as the repressors of anthocyanin biosynthesis, and five WD40s in the anthocyanin and PA biosynthetic pathways were significantly higher in the SR walnuts. Gene-metabolite correlation analyses revealed a core set of 31 genes that were strongly correlated with four anthocyanins and one PA metabolites. The alteration of gene coding sequence altered the binding or regulation of regulatory factors to structural genes in different color leaves, resulting in the effective increase of anthocyanins and PAs accumulation in red walnut., Conclusions: This study provides valuable information on anthocyanin and PA metabolites and candidate genes for anthocyanin and PA biosynthesis, yielding new insights into anthocyanin and PA biosynthesis in walnuts., Competing Interests: The authors declare there are no competing interests., (©2022 Wang et al.)

With the increase in the number of patients and prolongation of their lives after esophagectomy for esophageal cancer, the quality of life after surgery has attracted more and more attention. Although anastomotic stenosis is a common complication, it seriously affects the quality of life and psychological state of patients or even threatens their lives. At present, the exact independent influencing factors of anastomotic stenosis after esophageal cancer surgery have not been determined, and relevant treatment options are still controversial. Here, we analyzed the independent risk factors leading to good postoperative anastomotic stenosis, in order to provide a basis for late prevention. At the same time, we deeply discussed the advantages and safety of stent implantation in the treatment of anastomotic stenosis., Competing Interests: The authors declare that they have no conflicts of interest.., (Copyright © 2022 Guoliang Wu et al.)

Background: Biomarkers in predicting the stages of nephropathy associated with type 2 diabetes mellitus are urgent, and adiponectin may be a promising biomarker. This meta-analysis examined the association of serum adiponectin level with the stages of type 2 diabetic nephropathy., Methods: Databases including PubMed, Cochrane Library, EMBASE, China National Knowledge Infrastructure (CNKI), and Wan Fang were searched for published studies on adiponectin and type 2 diabetic kidney disease. The Newcastle-Ottawa scale was used to assess the quality of the literature. STATA 14.0 was used to conduct the statistical analysis., Results: Thirty-four studies with 5254 patients were included in this meta-analysis. The results of this study show that there was no significant difference in serum adiponectin level between normoalbuminuria and the control group (mean difference = -0.42, 95% CI [-1.23, 0.40]), while serum adiponectin level was positively correlated with the severity of type 2 diabetic kidney disease. The serum adiponectin level in type 2 diabetic kidney disease patients ranks as macroalbuminuria > microalbuminuria > normoalbuminuria., Conclusions: Serum adiponectin level might be an important marker to predict the progression of type 2 diabetic kidney disease., Competing Interests: The authors declare that they have no conflict of interest., (Copyright © 2022 Li Li et al.)

Objective: Explore the factors affecting the QO of life after transcatheter aortic valve replacement (TAVR) and analyze and evaluate their surgical efficacy and postoperative survival status., Methods: Through correlation analysis and multiple regression analysis, we predict various clinical characteristics and postoperative quality and predict clinical changes in L postoperative quality., Results: The quality of life of patients with the disease has gradually improved and improved from 6 months after surgery. The differences in the three aspects of its physiological mechanism function, physiological function function, overall health, and vitality are statistically significant ( p < 0.05)., Conclusion: Compared with traditional open-thoracic aortic valve (AV) surgery, TAVR has the significant advantages of smaller surgical incision and less trauma to the patient, which has become one of the reasons why patients are willing to accept it., Competing Interests: There is no potential conflict of interest in our paper, and all authors have seen the manuscript and approved to submit to your journal. We confirm that the content of the manuscript has not been published or submitted for publication elsewhere., (Copyright © 2021 Guoliang Wu et al.)

The early hematoma expansion of intracerebral hemorrhage (ICH) indicates a poor prognosis. This paper studies the relationship between cerebral blood flow (CBF) around the hematoma and hematoma expansion (HE) in the acute stage of intracerebral hemorrhage. A total of 50 patients with supratentorial cerebral hemorrhage were enrolled in this study. They underwent baseline whole-brain CTP within 6 h after intracerebral hemorrhage, and non-contrast CT within 24 h. Absolute hematoma growth and relative hematoma growth were calculated, respectively. A relative growth of Hematoma volume >33% was considered to be hematoma expansion. The Ipsilateral peri-edema CBF and Ipsilateral edema CBF were calculated by CTP maps in patients with and without hematoma expansion, respectively. In this study the incidence of hematoma expansion in the early stage of supratentorial cerebral hemorrhage was 32%; The CBF of the hematoma expansion group was higher than that of the patients without hematoma expansion (23.5 ± 12.5 vs. 15.1 ± 7.4, P = 0.004). After adjusting for age, gender, Symptom onset to NCCT and Baseline hematoma volume, ipsilateral peri-edema CBF was still an independent risk factor for early HE (or = 1.095, 95% CI = 1.01-1.19, P = 0.024). Here, we concluded that higher cerebral blood flow predicts early hematoma expansion in patients with intracerebral hemorrhage., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Wang, Jin, Feng, Wu, Wang, Jia, Ji, Wang and Zhao.)

Basic helix-loop-helix (bHLH) proteins are transcription factors (TFs) that have been shown to regulate anthocyanin biosynthesis in many plant species. However, the bHLH gene family in walnut ( Juglans regia L.) has not yet been reported. In this study, 102 bHLH genes were identified in the walnut genome and were classified into 15 subfamilies according to sequence similarity and phylogenetic relationships. The gene structure, conserved domains, and chromosome location of the genes were analyzed by bioinformatic methods. Gene duplication analyses revealed that 42 JrbHLHs were involved in the expansion of the walnut bHLH gene family. We also characterized cis -regulatory elements of these genes and performed Gene Ontology enrichment analysis of gene functions, and examined protein-protein interactions. Four candidate genes ( JrEGL1a , JrEGL1b , JrbHLHA1 , and JrbHLHA2 ) were found to have high homology to genes encoding bHLH TFs involved in anthocyanin biosynthesis in other plants. RNA sequencing revealed tissue- and developmental stage-specific expression profiles and distinct expression patterns of JrbHLHs according to phenotype (red vs. green leaves) and developmental stage in red walnut hybrid progeny, which were confirmed by quantitative real-time PCR analysis. All four of the candidate JrbHLH proteins localized to the nucleus, consistent with a TF function. These results provide a basis for the functional characterization of bHLH genes and investigations on the molecular mechanisms of anthocyanin biosynthesis in red walnut., Competing Interests: ZD was employed by the Shangluo Shengda Industrial Co., Ltd. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Zhao, Liu, Li, Meng, Yang, Dong, Wang and Wu.)

Ubiquitination is a critical post-translational modification machinery that governs a wide range of cellular functions by regulating protein homeostasis. Identification of ubiquitinated proteins and lysine residues can help researchers better understand the physiological roles of ubiquitin modification in different biological systems. In this study, we report the first comprehensive analysis of the peach ubiquitome by liquid chromatography-tandem mass spectrometry-based diglycine remnant affinity proteomics. Our systematic profiling revealed a total of 544 ubiquitination sites on a total of 352 protein substrates. Protein annotation and functional analysis suggested that ubiquitination is involved in modulating a variety of essential cellular and physiological processes in peach, including but not limited to carbon metabolism, histone assembly, translation and vesicular trafficking. Our results could facilitate future studies on how ubiquitination regulates the agricultural traits of different peach cultivars and other crop species.

Short interspersed nuclear elements (SINEs) are non-autonomous retrotransposons that are highly abundant, but not well annotated, in plant genomes. In this study, we identified 41,573 copies of SINEs in seven citrus genomes, including 11,275 full-length copies. The citrus SINEs were distributed among 12 families, with an average full-length rate of 0.27, and were dispersed throughout the chromosomes, preferentially in AT-rich areas. Approximately 18.4% of citrus SINEs were found in close proximity (≤1 kb upstream) to genes, indicating a significant enrichment of SINEs in promoter regions. Citrus SINEs promote gene and genome evolution by offering exons as well as splice sites and start and stop codons, creating novel genes and forming tandem and dispersed repeat structures. Comparative analysis of unique homologous SINE-containing loci (HSCLs) revealed chromosome rearrangements in sweet orange, pummelo, and mandarin, suggesting that unique HSCLs might be valuable for understanding chromosomal abnormalities. This study of SINEs provides us with new perspectives and new avenues by which to understand the evolution of citrus genes and genomes., (© The Author(s) 2020. Published by Oxford University Press on behalf of Kazusa DNA Research Institute.)

Three-dimensional (3D) reconstructed images have been increasingly applied for medical education. Although many studies have described the benefits of such applications, the best time to introduce 3D technology into surgical training has not been determined. Therefore, we conducted a randomized study to determine a suitable period for the introduction of this technology. Seventy-one surgical residents were randomized into 2 groups (two-dimensional computed tomography (CT) group and 3D image group), and they completed a test on anatomy and imaging as well as a questionnaire. Post-graduate year 1 (PGY1) residents in the 3D group performed significantly better than those in the CT group, although the third-year residents did not present significant differences in either the score or the time spent answering the questions. Although residents in different years of training held different attitudes toward the difficulty of anatomy and imaging learning, they all showed a high level of acceptance of the 3D training. This study revealed that 3D images improved the junior residents' performance in imaging reasoning. Thus, systematically introducing 3D images early in a surgical resident training program may help produce a better anatomy-imaging-surgery system.

The rhizosphere microenvironment is the site of nutrient circulation and microbial community formation, and thus is an ongoing topic of research. Although research on this topic is extensive, studies into the rhizosphere microenvironment of fruit trees remain rare. To elucidate the mechanisms driving the fruit tree rhizosphere microenvironment, we assessed soil physicochemical properties, enzyme activities, the community-level physiological profile (CLPP) and microbial diversity in rhizospheric soils of eight common deciduous fruit trees in northern China. We found that the available minerals, pH, enzyme activities, microbial utilization of six types of carbon (C) substrates, and microbial diversity in the rhizosphere varied among tree species. Redundancy analysis (RDA) showed that rhizosphere microenvironmental parameters (ammonia nitrogen content, soil pH and invertase activity) were closely related to the soil microbial community. Further analysis revealed that the soil microbial utilization of six C sources, nitrate nitrogen content, and invertase activity were negatively correlated with Ambiguous species and Alternaria ; however, these groups were positively correlated with pH. The ammonia nitrogen content was positively correlated with C source utilization and negatively correlated with Ambiguous, Lysobacter, Nitrospira, Alternaria, Fusarium , and Colletotrichum . Interestingly, invertase was closely linked to the microbial community, especially fungal diversity, and was positively correlated with plant-beneficial microbes such as Mortierella, Geomyces, Lysobacter , and Chaetomium , but negatively correlated with pathogenic microbes such as Alternaria, Fusarium , and Colletotrichum . Hence, rhizosphere soil physicochemical properties, enzyme activities and microbial community were significantly affected by tree species. Additionally, a variety of environmental factors were closely related to the microbial community in the rhizospheric soils of eight species of deciduous fruit trees.

Regulatory T (Treg) cells are critical suppressors of inflammation and are thought to exert mainly deleterious effects in cancers. In colorectal cancer (CRC), Foxp3 + Treg accumulation in tumors was associated with poor prognosis. Hence, we examined the circulating Treg cells in CRC patients. Compared to controls, CRC patients presented mild upregulations in CD4 + CD25 +/hi T cells and in the more canonical CD4 + CD25 +/hi Foxp3 + Treg cells in peripheral blood mononuclear cells. Both of these Treg populations could be roughly divided into lymphocyte activation gene 3 negative T cell immunoglobulin and mucin-domain containing-3 negative (LAG3 - TIM3 - ) and LAG3 + TIM3 + subsets. In CRC patients, the LAG3 + TIM3 + subset represented approximately half of CD4 + CD25 +/hi T cells and greater than 60% of CD4 + CD25 +/hi Foxp3 + Treg cells, which was significantly more frequent than in healthy controls. Compared to the LAG3 - TIM3 - CD4 + CD25 +/hi T cells, the LAG3 + TIM3 + CD4 + CD25 +/hi T cells presented considerably higher transforming growth factor-β and slightly higher interleukin (IL)-10 secretion, together with higher cytotoxic T-lymphocyte associated protein 4 and Foxp3 expression levels. Notably, macrophages following incubation with LAG3 - TIM3 - CD4 + CD25 +/hi T cells and LAG3 + TIM3 + CD4 + CD25 +/hi T cells displayed different characteristics. Macrophages incubated with LAG3 + TIM3 + CD4 + CD25 +/hi T cells presented lower expression of major histocompatibility complex class II, CD80, CD86, and tumor necrosis factor-α but higher expression of IL-10, than macrophages incubated with LAG3 - TIM3 - CD4 + CD25 +/hi T cells. Together, our investigations demonstrated that CRC patients presented an enrichment of circulating Treg cells, in which the LAG3 + TIM3 + subset exhibited more potent expression of inhibitory molecules, and furthermore, the LAG3 + TIM3 + Treg cells could suppress the proinflammatory activation of macrophages more potently than the LAG3 - TIM3 - Treg cells., (© 2018 John Wiley & Sons Australia, Ltd.)

Although colorectal cancer (CRC) is a prevalent malignancy of the digestive system, the underlying mechanisms of CRC tumorigenesis are still elusive. Arrestin-related domain-containing protein-3 (ARRDC3) has been reported to promote lysosome-mediated protein degradation. In the present study, we find that the expression of ARRDC3 is downregulated in CRC specimens. Mechanistically, we reveal that ARRDC3 binds and decreases expression of the oncoprotein YAP, the cotranscription factor of the Hippo pathway. The regulation of the Hippo pathway by ARRDC3 is conserved from Drosophila to mammals. Furthermore, we demonstrate that ARRDC3 plays an anti-oncogenic role in CRC progression by promoting YAP degradation. Finally, we show that ARRDC3 increases the sensitivity of CRC cells toward chemotherapeutic drugs. Taken together, our findings point to ARRDC3 as a potential target for CRC treatment., (© 2018 Federation of European Biochemical Societies.)

Fruit color is an important economic trait. The color of red walnut cultivars is mainly attributed to anthocyanins. The aim of this study was to explore the differences in the molecular mechanism of leaf and peel color change between red and green walnut. A reference transcriptome of walnut was sequenced and annotated to identify genes related to fruit color at the ripening stage. More than 290 million high-quality reads were assembled into 39,411 genes using a combined assembly strategy. Using Illumina digital gene expression profiling, we identified 4568 differentially expressed genes (DEGs) between red and green walnut leaf and 3038 DEGs between red and green walnut peel at the ripening stage. We also identified some transcription factor families ( MYB , bHLH , and WD40 ) involved in the control of anthocyanin biosynthesis. The trends in the expression levels of several genes encoding anthocyanin biosynthetic enzymes and transcription factors in the leaf and peel of red and green walnut were verified by quantitative real-time PCR. Together, our results identified the genes involved in anthocyanin accumulation in red walnut. These data provide a valuable resource for understanding the coloration of red walnut., Competing Interests: The authors declare no conflict of interest.

Liquid-liquid phase separation (LLPS) has long been observed during the physical stability investigation of therapeutic protein formulations. The buffer conditions and the presence of various excipients are thought to play important roles in the formulation development of monoclonal antibodies (mAbs). In this study, the effects of several small-molecule excipients (histidine, alanine, glycine, sodium phosphate, sodium chloride, sorbitol and sucrose) with diverse physical-chemical properties on LLPS of a model IgG1 (JM2) solutions were investigated by multiple techniques, including UV-vis spectroscopy, circular dichroism, differential scanning calorimetry/fluorimetry, size exclusion chromatography and dynamic light scattering. The LLPS of JM2 was confirmed to be a thermodynamic equilibrium process with no structural changes or irreversible aggregation of proteins. Phase diagrams of various JM2 formulations were constructed, suggesting that the phase behavior of JM2 was dependent on the solution pH, ionic strength and the presence of other excipients such as glycine, alanine, sorbitol and sucrose. Furthermore, we demonstrated that for this mAb, the interaction parameter (k D ) determined at low protein concentration appeared to be a good predictor for the occurrence of LLPS at high concentration., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Recently, increasing research efforts have been devoted into developing high-concentration protein drugs for subcutaneous injection, especially for those with short half-lives and high-dose requirement. Proteins at high concentrations normally present increased colloidal and structural instability, such as aggregation, fibrillation and gelation, which significantly challenges the high-concentration formulation development of protein drugs. Here we used endostatin, a 20kD recombinant protein, as a model drug for high-concentration formulation optimization. The colloidal and conformational stability of endostatin at high concentration of 30mg/mL were investigated in formulations containing various excipients, including saccharides (mannitol, sorbitol and sucrose), salts (ArgHCl and NaCl), and surfactants (tween 20 and 80). Protein fibrillation was characterized and semi-quantified by optical polarized light microscopy and transmission electron microscopy, and the amount of fiber formation at elevated temperature of 40°C was determined. The soluble protein aggregates were characterized by dynamic and static light scattering before and after dilution. The conformational stability were characterized by polyacrylamide gel electrophoresis, fluorescence, circular dichroism, and differential scanning calorimetry. We observed that the soluble aggregation, fibrillation and gelation, induced by conformational and colloidal instabilities of the protein solution, could be substantially optimized by using suitable stabilizers such as combinations of saccharides and surfactants; while formation of gel and soluble aggregates at high protein concentration (e.g., 30mg/mL) and elevated temperature (40°C) could be prevented by avoiding the usage of salts. It's worth emphasizing that some stabilizers, such as salts and surfactants, could show opposite contributions in conformational and colloidal stabilities of endostatin. Therefore, cautions are needed when one attempts to correlate the colloidal stability of high-concentration proteins with their conformational stability, and the colloidal and conformational protein stabilities must be harmonized by a balanced selection of various types of excipients., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Surfactants are widely used as stabilizers in the biopharmaceutical formulations to minimize protein aggregation. Under a fixed stress condition, the protecting and destabilizing effects of surfactants are hypothesized to be highly dependent on the species and concentrations of surfactants and mAb. Therefore, we here studied the aggregation-prevention and structure-perturbation effects of eight commonly used surfactants (Tw20, Tw80, Brij35, Chaps, TrX-100, SDS, Pluronic F68 and F127) on two IgG1 solution formulations under agitation, using analytical methodologies including visual inspection, OD 350 measurement, HPLC-SEC, circular dicroism, fluorescence spectroscopy and differential scanning calorimetry. We found that: (1) With concentrations range from 0.02 to 2mg/mL, nonionic surfactants were found to offer efficient aggregation-prevention effect, which is superior than the ionic surfactants; and higher surfactant concentration prevented mAb aggregation better especially under prolonged stability test under stress conditions. (2) The surfactant induced structure-perturbation emerged when even higher surfactant concentration (≥2mg/mL) was used, and such effect was surfactant-property dependent; and (3) the two IgG1 demonstrated different aggregation mechanisms and surfactant dependency, especially at high mAb concentrations. In conclusion, surfactants usage in mAb formulations, including the types and concentrations, should strike an optimal balance between the desirable aggregation-prevention and the detrimental structure-perturbation effects, while the consideration of mAb aggregation mechanism and concentration is also required for surfactant assessment., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Oxymatrine (OMT), one of the main active components of extracts from the dry roots of Sophora flavescens, has been reported to possess many pharmacological properties including cancer-preventive and anti-cancer effects. The aim of the present study is to explore the efficiency of combination therapy with OMT and oxaliplatin (OXA) and identify the in vitro and in vivo cytotoxicity on colon cancer lines (HT29 and SW480) and mice model. Cells were treated with OMT and/or OXA and subjected to cell viability, colony formation, apoptosis, cell cycle, western blotting, xenograft tumorigenicity assay and immunohistochemistry. The results demonstrated that OMT and OXA inhibited the proliferation of colon cancer cells, and combination therapy of OMT and OXA resulted in a combination index < 1, indicating a synergistic effect. Co-treatment with OMT and OXA caused G0/G1 phase arrest by upregulating P21, P27 and downregulating cyclin D, and induced apoptosis through decreasing the expression of p-PI3K, p-AKT, p-mTOR, p-p70S6K. In addition, pretreatment with a specific PI3K/AKT activator (IGF-1) significantly neutralized the pro-apoptotic activity of OXA + OMT, demonstrating the important role of PI3K/AKT in this process. Moreover, in nude mice model, co-treatment displayed more efficient inhibition of tumor weight and volume on SW480 xenograft mouse model than single-agent treatment with OXA or OMT. Immunohistochemistry analysis suggests the combinations greatly suppressed tumor proliferation, which consistent with our in vitro results. In conclusion, our findings highlight that the combination therapy with OMT and OXA exerted synergistic antitumor effects in colon cancer cells through PI3K/AKT/mTOR pathway and combination treatment with OMT and OXA would be a promising therapeutic strategy for colon carcinoma treatment.

Quercetin (Q), a flavonoid compound, which is obtained in variety of fruits, seeds, and vegetables, has been reported to possess many pharmacological properties including cancer-preventive and anticancer effects. However, studies on the anticancer effects and underlying mechanisms of Q in human hepatocellular carcinoma (HCC) are still limited. The present study is conducted to investigate the anticancer efficacy and adjuvant chemotherapy action of Q in HCC. HCC cell lines HepG2 and SMCC-7721 were treated with different concentrations of Q. The antiproliferative effects of Q were measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), and the apoptosis and cell cycle dynamics were assessed by flow cytometry; the expression of apoptosis-associated proteins were evaluated by Western blot and immunohistochemistry staining; the tumor growth in vivo was evaluated in a xenograft mouse model. Our results showed that Q effectively inhibited human HCC cell proliferation and induced apoptosis by upregulating the expression of Bad and Bax and downregulating the expression of Bcl-2 and Survivin in vitro. Furthermore, Q obviously inhibited the tumor growth and enhanced the 5-fluorouracil (5-FU) therapeutic efficacy in vitro and in vivo. Taken together, our findings highlight that Q effectively inhibited the growth of tumor and enhanced the sensitivity to thermotherapy, indicating Q is a potential treatment option for HCC.

Sorafenib is a clinically important oral tyrosine kinase inhibitor for the treatment of various cancers. However, the oral bioavailability of sorafenib tablet (Nexavar) is merely 38-49% relative to the oral solution, due to the low aqueous solubility of sorafenib and its relatively high daily dose. It is desirable to improve the oral bioavailability of sorafenib to expand the therapeutic window, reduce the drug resistance, and enhance patient compliance. In this study, we observed that the solubility of sorafenib could be increased ∼50-fold in the coexistence of poly(vinylpyrrolidone-vinyl acetate) (PVP-VA) and sodium lauryl sulfate (SLS), due to the formation of PVP-VA/SLS complexes at a lower critical aggregation concentration. The enhanced solubility provided a faster initial sorafenib dissolution rate, analogous to a forceful "spring" to release drug into solution, from tablets containing both PVP-VA and SLS. However, SLS appears to impair the ability of PVP-VA to act as an efficient "parachute" to keep the drug in solution and maintain drug supersaturation. Using 2D (1)H NMR, (13)C NMR, and FT-IR analysis, we concluded that the solubility enhancement and supersaturation of sorafenib were achieved by PVP-VA/SLS complexes and PVP-VA/sorafenib interaction, respectively, both through molecular interactions hinged on the PVP-VA VA groups. Therefore, a balance between "spring" and "parachute" must be carefully considered in formulation design. To confirm the in vivo relevance of these molecular interaction mechanisms, we prepared three tablet formulations containing PVP-VA alone, SLS alone, and PVP-VA/SLS in combination. The USP II in vitro dissolution and dog pharmacokinetic in vivo evaluation showed clear differentiation between these three formulations, and also good in vitro-in vivo correlation. The formulation containing PVP-VA alone demonstrated the best bioavailability with 1.85-fold and 1.79-fold increases in Cmax and AUC, respectively, compared with the formulation containing SLS only, the poorest performing one. Despite its forceful "spring", the formulation containing both PVP-VA and SLS showed a moderate bioavailability enhancement, due to the lack of an efficient "parachute".

The cytostatic drug from fruits and other plant derived products have acted as a chemotherapeutic agent used in treatment of a wide variety of cancers. Lupeol, a dietary triterpene, present in many fruits and medicinal plants, has been shown to possess many pharmacological properties including anti-cancer effect in both in vitro and in vivo assay systems. However, the cancer proliferative and invasive inhibitory effects and molecular mechanisms on gallbladder carcinoma GBC-SD cells have not been studied. In the present study, GBC-SD cells were treated by lupeol and subjected to methyl thiazolyl tetrazolium analysis, Hoechst 33342 staining, annexin V/propidium iodide double-staining, transwell chamber assay and Western blot analysis. In addition, GBC-SD xenograft tumors were established in male nude BALB/c mice, and lupeol was intravenously administered to evaluate the anti-cancer capacity in vivo. Our results showed that lupeol inhibited the proliferation, migration, invasion and induced apoptosis of GBC-SD cells in a dose-dependent manner in vitro. Furthermore, the expression of p-EGFR, p-AKT and MMP-9 levels were significantly down-regulated. These protein interactions may play a pivotal role in the regulation of apoptosis and invasion. More importantly, our in vivo studies showed that administration of lupeol decreased tumor growth in a dose-dependent manner. Immunohistochemistry analysis demonstrated the down-regulation of p-EGFR and MMP-9 in tumor tissues following lupeol treatment, consistent with the in vitro results. Taken together, our findings indicated that lupeol can induce apoptotic cell death and inhibit the migration as well as invasion of GBC-SD cells. The mechanism may be associated with the suppression of EGFR/MMP-9 signaling. These results might offer a therapeutic potential advantage for human gallbladder carcinoma chemoprevention or chemotherapy.

By lyophilization of rifampicin (RIF) solution in TBA/water with various solvent compositions, uniform and amorphous rifampicin (RIF) microspheres were produced. Using 55% TBA solution, the obtained RIF microspheres have a mono-dispersive size distribution with diameters range from 1 to 3 μm. The RIF microspheres are found to be amorphous by X-ray diffraction, and are expected to dissolve much faster than the crystalline RIF upon inhalation. Mechanistic investigation revealed that the amorphous RIF microspheres were formed due to liquid-liquid phase separation (LLPS) occurred during the freezing of the TBA/water solution. We also observed that the RIF microspheres can be readily phagocytized by activated THP-1 cells within 15 min. The suitable size distribution, high solubility, and readiness for phagocytosis by macrophages, all suggest that the lyophilized amorphous RIF microspheres could be potentially used as an anti-tuberculosis inhalation therapy. In addition, similar process was used to lyophilize TBA/water solutions of several other drugs, including rifaximin, rifapentine, paclitaxel, and isoniazid. We found that for drugs with appropriate physiochemical properties, such as paclitaxel and rifaximin, mono-dispersive microspheres could be obtained as well, which demonstrated that freezing induced LLPS could be utilized as a novel particle engineering methodology to produce drug microspheres by lyophilization., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Lupeol, a dietary triterpene, present in many fruits and medicinal plants, has been reported to possess many pharmacological properties including anti-cancer activities both in vitro and in vivo. However, the precise mechanism involved remains largely unknown. The present study is conducted to investigate the anti-cancer activity and the underlying mechanisms of lupeol on human pancreatic cancer proliferating cell nuclear antigen 1 (PCNA-1) cells in vitro and in vivo. Lupeol significantly inhibited the proliferation of the cells in dose- and time-dependent manners and induced apoptosis as well as cell cycle arrest in G0/G1 phase by upregulating P21 and P27 and downregulating cyclin D1. The expression of apoptosis-related proteins in cells was evaluated by western blot analysis, and we found that lupeol induced cell apoptosis by decreasing the levels of p-AKT and p-ERK. In addition, pretreatment with a specific PI3K/AKT activator (IGF-1) significantly neutralized the pro-apoptotic activity of lupeol in PCNA-1 cells, demonstrating the important role of AKT in this process. More importantly, our in vivo studies showed that administration of lupeol decreased tumor growth in a dose-dependent manner. Immunohistochemistry analysis demonstrated the downregulation of p-AKT and p-ERK in tumor tissues following lupeol treatment, consistent with the in vitro results. Therefore, these findings indicate that lupeol can inhibit cell proliferation and induce apoptosis as well as cell cycle arrest of PCNA-1 cells and might offer a therapeutic potential advantage for human pancreatic cancer chemoprevention or chemotherapy.

Background/aims: We used bioinformatics technology to analyze gene expression profiles involved in colorectal cancer tissue samples and healthy controls., Methodology: In this paper, we downloaded the gene expression profile GSE4107 from Gene Expression Omnibus (GEO) database, in which a total of 22 chips were available, including normal colonic mucosa tissue from normal healthy donors (n=10), colorectal cancer tissue samples from colorectal patients (n=33). To further understand the biological functions of the screened DGEs, the KEGG pathway enrichment analysis were conducted. Then we built a transcriptome network to study differentially co-expressed links., Results: A total of 3151 DEGs of CRC were selected. Besides, total 164 DCGs (Differentially Coexpressed Gene, DCG) and 29279 DCLs (Differentially Co-expressed Link, DCL) were obtained. Furthermore, the significantly enriched KEGG pathways were Endocytosis, Calcium signaling pathway, Vascular smooth muscle contraction, Linoleic acid metabolism, Arginine and proline metabolism, Inositol phosphate metabolism and MAPK signaling pathway., Conclusion: Our results show that the generation of CRC involves multiple genes, TFs and pathways. Several signal and immune pathways are linked to CRC and give us more clues in the process of CRC. Hence, our work would pave ways for novel diagnosis of CRC, and provided theoretical guidance into cancer therapy.