A P450 superfamily member NtCYP82C4 promotes nicotine biosynthesis in Nicotiana tabacum.

In plants, cytochrome P450s are monooxygenase that play key roles in the synthesis and degradation of intracellular substances. In tobacco, the majority of studies examining the P450 superfamily have concentrated on the CYP82E subfamily, where multiple family members function as demethylases, facilitating the synthesis of nornicotine. In this study, NtCYP82C4, a tobacco P450 superfamily member, was identified from a gene-edited tobacco mutant that nicotine biosynthesis in tobacco leaves is evidently reduced. Compared to the wild-type plants, the knockout of NtCYP82C4 resulted in a significantly lower nicotine content and biomass in tobacco leaves. Transcriptome and metabolome analyses indicated that the knockout of NtCYP82C4 inhibites secondary metabolic processes in tobacco plants, leading to the accumulation of some important precursors in the nicotine synthesis process, including aspartic acid and nicotinic acid, and increases nitrogen partitioning associated with those processes such as amino acid synthesis and utilization. It is speculated that NtCYP82C4 may function as an important catalase downstream of the nicotine synthesis. Currently, most of the steps and enzymes involved in the nicotine biosynthesis process in tobacco have been elucidated. Here, our study deepens the current understanding of nicotine biosynthesis process and provides new enzyme targets for nicotine synthesis in tobacco plants.

Nomogram prediction of vessels encapsulating tumor clusters in small hepatocellular carcinoma ≤ 3 cm based on enhanced magnetic resonance imaging.

BACKGROUND: Vessels encapsulating tumor clusters (VETC) represent a recently discovered vascular pattern associated with novel metastasis mechanisms in hepatocellular carcinoma (HCC). However, it seems that no one have focused on predicting VETC status in small HCC (sHCC). This study aimed to develop a new nomogram for predicting VETC positivity using preoperative clinical data and image features in sHCC (≤ 3 cm) patients. AIM: To construct a nomogram that combines preoperative clinical parameters and image features to predict patterns of VETC and evaluate the prognosis of sHCC patients. METHODS: A total of 309 patients with sHCC, who underwent segmental resection and had their VETC status confirmed, were included in the study. These patients were recruited from three different hospitals: Hospital 1 contributed 177 patients for the training set, Hospital 2 provided 78 patients for the test set, and Hospital 3 provided 54 patients for the validation set. Independent predictors of VETC were identified through univariate and multivariate logistic analyses. These independent predictors were then used to construct a VETC prediction model for sHCC. The model's performance was evaluated using the area under the curve (AUC), calibration curve, and clinical decision curve. Additionally, Kaplan-Meier survival analysis was performed to confirm whether the predicted VETC status by the model is associated with early recurrence, just as it is with the actual VETC status and early recurrence. RESULTS: Alpha-fetoprotein_lg10, carbohydrate antigen 199, irregular shape, non-smooth margin, and arterial peritumoral enhancement were identified as independent predictors of VETC. The model incorporating these predictors demonstrated strong predictive performance. The AUC was 0.811 for the training set, 0.800 for the test set, and 0.791 for the validation set. The calibration curve indicated that the predicted probability was consistent with the actual VETC status in all three sets. Furthermore, the decision curve analysis demonstrated the clinical benefits of our model for patients with sHCC. Finally, early recurrence was more likely to occur in the VETC-positive group compared to the VETC-negative group, regardless of whether considering the actual or predicted VETC status. CONCLUSION: Our novel prediction model demonstrates strong performance in predicting VETC positivity in sHCC (≤ 3 cm) patients, and it holds potential for predicting early recurrence. This model equips clinicians with valuable information to make informed clinical treatment decisions.

Genome-wide identification of Shaker K+ channel family in Nicotiana tabacum and functional analysis of NtSKOR1B in response to salt stress.

Soil salinization poses a mounting global ecological and environmental threat. The identification of genes responsible for negative regulation of salt tolerance and their utilization in crop improvement through gene editing technologies emerges as a swift strategy for the effective utilization of saline-alkali lands. One efficient mechanism of plant salt tolerance is maintaining the proper intracellular K+/Na+ ratio. The Shaker K+ channels play a crucial role in potassium absorption, transport, and intracellular potassium homeostasis in plant cells. Here, the study presents the first genome-wide identification of Shaker K+ channels in Nicotiana tabacum L., along with a detailed bioinformatic analysis of the 20 identified members. Transcriptome analysis revealed a significant up-regulation of NtSKOR1B, an outwardly-rectifying member predominantly expressed in the root tissue of tobacco seedlings, in response to salt stress. This finding was then confirmed by GUS staining of ProNtSKOR1B::GUS transgenic lines and RT-qPCR analysis. Subsequently, NtSKOR1B knockout mutants (ntskor1) were then generated and subjected to salt conditions. It was found that ntskor1 mutants exhibit enhanced salt tolerance, characterized by increased biomass, higher K+ content and elevated K+/Na+ ratios in both leaf and root tissues, compared to wild-type plants. These results indicate that NtSKOR1B knockout inhibits K+ efflux in root and leaf tissues of tobacco seedlings under salt stress, thereby maintaining higher K+/Na+ ratios within the cells. Thus, our study identifies NtSKOR1B as a negative regulator of salt tolerance in tobacco seedlings.

Radiomics of Preoperative Multi-Sequence Magnetic Resonance Imaging Can Improve the Predictive Performance of Microvascular Invasion in Hepatocellular Carcinoma.

Background: The aim of the study is to demonstrate that radiomics of preoperative multi-sequence magnetic resonance imaging (MRI) can indeed improve the predictive performance of microvascular invasion (MVI) in hepatocellular carcinoma (HCC). Methods: A total of 206 patients with pathologically confirmed HCC who underwent preoperative enhanced MRI were retrospectively recruited. Univariate and multivariate logistic regression analysis identified the independent clinicoradiologic predictors of MVI present and constituted the clinicoradiologic model. Recursive feature elimination (RFE) was applied to select radiomics features (extracted from six sequence images) and constructed the radiomics model. Clinicoradiologic model plus radiomics model formed the clinicoradiomics model. Five-fold cross-validation was used to validate the three models. Discrimination, calibration, and clinical utility were used to evaluate the performance. Net reclassification improvement (NRI) and integrated discrimination improvement (IDI) were used to compare the prediction accuracy between models. Results: The clinicoradiologic model contained alpha-fetoprotein (AFP)_lg10, radiological capsule enhancement, enhancement pattern and arterial peritumoral enhancement, which were independent risk factors of MVI. There were 18 radiomics features related to MVI constructed the radiomics model. The mean area under the receiver operating curve (AUC) of clinicoradiologic, radiomics and clinicoradiomics model were 0.849, 0.925 and 0.950 in the training cohort and 0.846, 0.907 and 0.933 in the validation cohort, respectively. The three models' calibration curves fitted well, and decision curve analysis (DCA) confirmed the clinical usefulness. Compared with the clinicoradiologic model, the NRI of radiomics and clinicoradiomics model increased significantly by 0.575 and 0.825, respectively, and the IDI increased significantly by 0.280 and 0.398, respectively. Conclusions: Radiomics of preoperative multi-sequence MRI can improve the predictive performance of MVI in HCC.

Study on the correlation between the dominant microflora and the main flavor substances in the fermentation process of cigar tobacco leaves.

The flavor of cigar tobacco leaf determines the quality of finished cigar tobacco, and the enhancement of flavor generally relies on microbial fermentation. In this paper, the correlation between the dominant microorganisms and the main flavor substances of cigar tobacco leaves during fermentation and the correlation between the two were investigated to reveal the correlation between microorganisms and flavor and the metabolic pathways of microorganisms affecting the flavor substances. During the fermentation process, the main flavors of cigar tobacco leaves were sweet, light and grassy, with hexanal, 2,6-dimethylpyridine, nonanal, phenylacetaldehyde, naphthalene, and methyl benzoate as the main constituents, and the key microorganisms Haloferax mediterranei, Haloterrigena limicola, Candidatus Thorarchaeota archaeon SMTZ-45, the genera Methyloversatilis, Sphingomonas, Thauera, Pseudomonas, Penicillium, and Aspergillus. Correlation analysis revealed that fungi were negatively correlated with the main aroma and inhibited the main flavor substances, while bacteria were positively correlated with Benzoic acid, methyl ester in the main flavor substances, which was conducive to the accumulation of green aroma. Functional analysis revealed that the dominant bacterial population was producing aroma by metabolizing glycoside hydrolases and glycosyltransferases, performing amino acid metabolism, carbohydrate metabolism and film transport metabolism. The present study showed that the bacterial and fungal dominant microorganisms during the fermentation of cigar tobacco were influencing the production and degradation of the main flavor substances through the enzyme metabolism by the occurrence of the Merad reaction.

Clinical and DCE-CT signs in predicting microvascular invasion in cHCC-ICC.

BACKGROUND: To predict the microvascular invasion (MVI) in patients with cHCC-ICC. METHODS: A retrospective analysis was conducted on 119 patients who underwent CT enhancement scanning (from September 2006 to August 2022). They were divided into MVI-positive and MVI-negative groups. RESULTS: The proportion of patients with CEA elevation was higher in the MVI-positive group than in the MVI-negative group, with a statistically significant difference (P = 0.02). The MVI-positive group had a higher rate of peritumoral enhancement in the arterial phase (P = 0.01) whereas the MVI-negative group had more oval and lobulated masses (P = 0.04). According to the multivariate analysis, the increase in CEA (OR = 10.15, 95% CI: 1.11, 92.48, p = 0.04), hepatic capsular withdrawal (OR = 4.55, 95% CI: 1.44, 14.34, p = 0.01) and peritumoral enhancement (OR = 6.34, 95% CI: 2.18, 18.40, p < 0.01) are independent risk factors for predicting MVI. When these three imaging signs are combined, the specificity of MVI prediction was 70.59% (series connection), and the sensitivity was 100% (parallel connection). CONCLUSIONS: Our multivariate analysis found that CEA elevation, liver capsule depression, and arterial phase peritumoral enhancement were independent risk factors for predicting MVI in cHCC-ICC.

The association of carbohydrate antigen 19-9 response with radiologic response and survival in intrahepatic cholangiocarcinoma: A prospective cohort study.

BACKGROUND: The role of carbohydrate antigen 19-9 (CA 19-9) in response assessment among patients with intrahepatic cholangiocarcinoma (iCCA) remains unknown. The authors studied the association of the CA 19-9 response (defined as a reduction >50% from baseline) with the radiologic response and the outcome in patients with unresectable iCCA. METHODS: A prospective cohort of 422 patients who were initially diagnosed with unresectable iCCA, had baseline CA 19-9 levels ≥100 U/mL, and received treatment with systemic therapies at the authors' institution between January 2017 and December 2021 were enrolled in this study. The radiologic response was assessed using the Response Evaluation Criteria in Solid Tumors version 1.1. A landmark assessment of the CA 19-9 response and the radiologic response was performed. The associations between CA 19-9 response and imaging response, progression-free survival (PFS), and overall survival (OS) were analyzed. RESULTS: Two hundred sixty-seven patients (63.3%) had a CA 19-9 response. A CA 19-9 response was observed in 123 of 132 (93.2%) radiologic responders and in 144 of 290 (49.7%) radiologic nonresponders (p < .001). CA 19-9 responders outperformed nonresponders in median PFS (10.6 vs. 3.6 months; hazard ratio [HR], 4.8 months; 95% confidence interval [CI], 3.8-6.0 months; p < .001) and OS (21.4 vs. 6.3 months; HR, 5.3 months; 95% CI, 4.2-6.7 months; p < .001). The common independent predictors of both OS and PFS included metastasis, CA 19-9 nonresponder status, and radiologic nonresponder status in multivariable analysis. CONCLUSIONS: CA 19-9 response is a valuable addition to assess tumor response and is associated with improved outcomes in patients with iCCA. Achieving a CA 19-9 response should be one of the therapeutic objectives of patients with iCCA after systemic therapies. PLAIN LANGUAGE SUMMARY: A decline in carbohydrate antigen 19-9 levels from elevated baseline levels should be one of the therapeutic aims of patients with intrahepatic cholangiocarcinoma who are managed with systemic therapies.

The antihyperuricemia activity of Astragali Radix through regulating the expression of uric acid transporters via PI3K/Akt signalling pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Astragali Radix (AR) is the dry root of the leguminous plants Astragalus membranaceus (Fisch) Beg. var. mongholicus (Beg) Hsiao, and Astragalus membranaceus (Fisch) Bge., being used as a medicinal and edible resource. AR is used in traditional Chinese medicine prescriptions to treat hyperuricemia, but this particular effect is rarely reported, and the associated mechanism of action is still need to be elucidated. AIM OF THE STUDY: To research the uric acid (UA)-lowering activity and mechanism of AR and the representative compounds through the constructed hyperuricemia mouse and cellular models. MATERIALS AND METHODS: In our study, the chemical profile of AR was analysed by UHPLC-QE-MS, as well as the mechanism of action of AR and the representative compounds on hyperuricemia was studied through the constructed hyperuricemia mouse and cellular models. RESULTS: The main compounds in AR were terpenoids, flavonoids and alkaloids. Mice group treated with the highest AR dosage showed significantly lower (p < 0.0001) serum uric acid (208 ± 9 µmol/L) than the control group (317 ± 11 µmol/L). Furthermore, UA increased in a dose-dependence manner in urine and faeces. Serum creatinine and blood urea nitrogen standards, as well as xanthine oxidase in mice liver, decreased (p < 0.05) in all cases, indicating that AR could relieve acute hyperuricemia. UA reabsorption protein (URAT1 and GLUT9) was down-regulated in AR administration groups, while the secretory protein (ABCG2) was up-regulated, indicating that AR could promote the excretion of UA by regulating UA transporters via PI3K/Akt signalling pathway. CONCLUSION: This study validated the activity, and revealed the mechanism of AR in reducing UA, which provided experimental and clinical basis for the treatment of hyperuricemia with it.

Liver Metabolic Dysregulation Induced by Polypropylene Nano- and Microplastics in Nile Tilapia Using Internal Extractive Electrospray Ionization Mass Spectrometry.

Understanding the metabolic disorders induced by nano- and microplastics in aquatic organisms at the molecular level could help us understand the potential toxicity of nano- and microplastics more thoroughly and provide a fundamental scientific basis for regulating the usage and management of plastic products. In this research, the effect of polypropylene nanoplastics (PP-NPs) and microplastics (PP-MPs) on metabolites in the tilapia liver was comprehensively investigated by internal extractive electrospray ionization mass spectrometry (iEESI-MS). A partial least-squares discriminant analysis (PLS-DA) and a one-component analysis of variance (ANOVA) were used for selecting 46 differential metabolites, including phospholipids, amino acids, peptides, carbohydrates, alkaloids, purines, pyrimidines, and nucleosides. Pathway enrichment analysis showed significant effects on glycerophospholipid metabolism, arginine and proline metabolism, and aminoacyl-tRNA biosynthesis after tilapia were exposed to PP-N/MPs. Dysregulation of these metabolites is mainly reflected in the possible induction of hepatitis, oxidative stress, and other symptoms. The application of iEESI-MS technology without sample pretreatment to the study of metabolic disorders in aquatic organisms under the interference of nano- and microplastics provides a promising analytical method for environmental toxicology research.

Molecular mechanisms of the chemical constituents from anti-inflammatory and antioxidant active fractions of Ganoderma neo-japonicum Imazeki.

Ganoderma neo-japonicum Imazeki is a rare medicinal mushroom that has been reported to play a role in scavenging free radicals, protecting the liver, and inhibiting tumor cell activity. In this study, crude extracts were prepared, and 47 triterpenoids were identified by Ultra-high-performance liquid chromatography coupled with triple quadrupole time-of flight mass spectrometry (UHPLC-Triple TOF-MS/MS). Then, the crude extracts were subjected to column chromatography for the first time to obtain six fractions (Fr. (a), (b), (c), (d), (e) and (f)). Antioxidant and anti-inflammatory active tracking assays of all fractions found that Fr. (c) exhibited the strongest bioactivity. Subsequently, the chemical composition of Fr. (c) was clarified, and eight triterpenoids were determined in combination with the standard substances. In addition, this study demonstrated that Fr. (c) reduced the levels of inflammatory cytokines and reactive oxygen species (ROS) in LPS-stimulated RAW264.7 macrophages. Further studies showed that Fr. (c) could down-regulate the expression level of proteins associated of NF-κB signaling pathway, and upregulated Nrf2 and HO-1 protein level. In conclusion, our study showed that Fr. (c) inhibited LPS-mediated inflammatory response and oxidative stress by activating the Nrf2/HO-1 pathway and inactivating the NF-κB pathway. In the future, with the clearing of its composition and activity mechanism, Fr. (c) of G. neo-japonicum are expected to become a functional food for health and longevity.

Improved colonic inflammation by nervonic acid via inhibition of NF-κB signaling pathway of DSS-induced colitis mice.

BACKGROUND: Nervonic acid (C24:1∆15, 24:1 ω-9, cis-tetracos-15-enoic acid; NA), a long-chain monounsaturated fatty acid, plays an essential role in prevention of metabolic diseases, and immune regulation, and has anti-inflammatory properties. As a chronic, immune-mediated inflammatory disease, ulcerative colitis (UC) can affect the large intestine. The influences of NA on UC are largely unknown. PURPOSE: The present study aimed to decipher the anti-UC effect of NA in the mouse colitis model. Specifically, we wanted to explore whether NA can regulate the levels of inflammatory factors in RAW264.7 cells and mouse colitis model. METHODS: To address the above issues, the RAW264.7 cell inflammation model was established by lipopolysaccharide (LPS), then the inflammatory factors tumor necrosis factor-α (TNF-α), Interleukin-6 (IL-6), Interleukin-1ß (IL-1ß), and Interleukin-10 (IL-10) were detected by Enzyme-linked immunosorbent assay (ELISA). The therapeutic effects of NA for UC were evaluated using C57BL/6 mice gavaged dextran sodium sulfate (DSS). Hematoxylin and eosin (H&E) staining, Myeloperoxidase (MPO) kit assay, ELISA, immunofluorescence assay, and LC-MS/MS were used to assess histological changes, MPO levels, inflammatory factors release, expression and distribution of intestinal tight junction (TJ) protein ZO-1, and metabolic pathways, respectively. The levels of proteins involved in the nuclear factor kappa-B (NF-κB) pathway in the UC were investigated by western blotting and RT-qPCR. RESULTS: In vitro experiments verified that NA could reduce inflammatory response and inhibit the activation of key signal pathways associated with inflammation in LPS-induced RAW264.7 cells. Further, results from the mouse colitis model suggested that NA could restore intestinal barrier function and suppress NF-κB signal pathways to ameliorate DSS-induced colitis. In addition, untargeted metabolomics analysis of NA protection against UC found that NA protected mice from colitis by regulating citrate cycle, amino acid metabolism, pyrimidine and purine metabolism. CONCLUSION: These results suggested that NA could ameliorate the secretion of inflammatory factors, suppress the NF-κB signaling pathway, and protect the integrity of colon tissue, thereby having a novel role in prevention or treatment therapy for UC. This work for the first time indicated that NA might be a potential functional food ingredient for preventing and treating inflammatory bowel disease (IBD).

Actual over 3-year survival after stereotactic body radiation therapy in patients with unresectable intrahepatic cholangiocarcinoma.

PURPOSE: As a non-invasive treatment, stereotactic body radiation therapy (SBRT) has been an emerging and effective option for patients with unresectable intrahepatic cholangiocarcinoma (ICC). The Cyber Knife has an SBRT system, which can realize real-time tracking of tumors during treatment. It can protect the surrounding normal liver tissue while the tumor gets the therapeutic dose. The purpose of this study was to evaluate the factors affecting the local control rate for patients after SBRT treatment, and to predict the factors affecting survival rates, then to report the 3-year actual survival rates after treatment and identify the influencing factors of 3-year survival rate. MATERIALS AND METHODS: We conducted a long-term follow-up of 43 patients with unresectable intrahepatic cholangiocarcinoma who underwent Cyber Knife in our hospital from January 2016 to December 2018. Regular medical check-ups were performed every 2-3 months after SBRT to evaluated the effect of treatment. RESULTS: The median follow-up time was 15 months (4-78 months), and the median progression-free survival (PFS) was 6 months (95% CI, 2.788-9.212) and the median overall survival (OS) was 12 months (95% CI, 3.434-20.566), respectively. Based on modified Response Evaluation and Criteria in Solid Tumor (mRECIST), response rate (RR) and disease control rate (DCR) of SBRT in unresectable ICC were 55.2% and 86%. The 1-, 2- and 3-years OS rate were 51.2%, 32.6% and 23.3%. Multivariate analysis based on competing risk survival analysis identified that patients with multiple nodules, large diameter, high level of CA199 and CEA, poor ECOG performance status had worse overall survival (p < 0.05). Patients who survived ≥3 years had significantly lower levels of CEA, CA199, smaller tumor diameters and lower number of lesions (p < 0.05). CONCLUSION: The SBRT might be a candidate option for patients who unable to perform surgery. The rate of 3-year survival after SBRT for unresectable ICC can be expected with 23.3%.

Reverse effects of Streptococcus mutans physiological states on neutrophil extracellular traps formation as a strategy to escape neutrophil killing.

Bacteria in nature are present in different lifestyles with distinct characteristics. Streptococcus mutans is the etiologic pathogen of dental caries and could easily gain access into the bloodstream after oral surgery and adopt a biofilm lifestyle, resulting in infective endocarditis. A growing amount of evidence have revealed that the large web-like structure composed of extracellular DNA and antimicrobial proteins released by neutrophils, named Neutrophil Extracellular Traps (NETs), play an active role in the defense against bacterial invasion. The present study demonstrated that NETs formation was discriminatively affected by S. mutans biofilm and its planktonic counterpart. The free-floating planktonic S. mutans exhibited an active NETs response, whereas the biofilm community exhibited a reverse negative NETs response. Besides, impaired biofilm killing correlated with the decrease in NETs production. Unlike planktonic cells, biofilm avoided the burst of reactive oxygen species (ROS) when co-culture with neutrophils, and the NADPH-oxidase pathway was partially involved. A mice infection model also supported the distinguishing response of neutrophils challenged by different lifestyles of S. mutans. In conclusion, different bacterial physiological states can affect the distinct response of the host-microbe interaction, thus contributing to the anti-pathogen immune response activation and immune surveillance survival.

Associating liver partition and portal vein ligation for staged hepatectomy versus sequential transarterial chemoembolization and portal vein embolization in staged hepatectomy for HBV-related hepatocellular carcinoma: a randomized comparative study.

Background: Both portal vein embolization (PVE) and associating liver partition and portal vein ligation for staged hepatectomy (ALPPS) have merits and demerits when used in patients with unresectable liver cancers due to insufficient volumes in future liver remnant (FLR). Methods: This study was a single-center, prospective randomized comparative study. Patients with the diagnosis of hepatitis B related hepatocellular carcinoma (HCC) were randomly assigned in a 1:1 ratio to the 2 groups. The primary endpoints were tumor resection and three-year overall survival (OS) rates. Results: Between November 2014 to June 2016, 76 patients with unresectable HBV-related HCC due to inadequate volume of FLR were randomly assigned to ALPPS groups (n=38) and TACE + PVE groups (n=38). Thirty-seven patients (97.4%) in the ALPPS group compared with 25 patients (65.8%) in the TACE + PVE group were able to undergo staged hepatectomy (risk ratio 1.48, 95% CI: 1.17-1.87, P<0.001). The three-year OS rate of the ALPPS group (65.8%) (95% CI: 50.7-80.9) was significantly better than the TACE + PVE group (42.1%) (95% CI: 26.4-57.8) (HR 0.50, 95% CI: 0.26-0.98, two-sided P=0.036). However, no significant difference in the OS rates between patients who underwent tumor resection in the 2 groups of patients was found (HR 0.80, 95% CI: 0.35-1.83, two-sided P=0.595). Major postoperative complications rates after the stage-2 hepatectomy were 54.1% in the ALPPS group and 20.0% in the TACE + PVE group (risk ratio 2.70, 95% CI: 1.17-6.25, P=0.007). Conclusions: ALPPS resulted in significantly better intermediate-term OS outcomes, at the expenses of a significantly higher perioperative morbidity rate compared with TACE + PVE in patients who had initially unresectable HBV-related HCC.

Cabozantinib plus Nivolumab Phase I Expansion Study in Patients with Metastatic Urothelial Carcinoma Refractory to Immune Checkpoint Inhibitor Therapy.

PURPOSE: This study investigated the efficacy and tolerability of cabozantinib plus nivolumab (CaboNivo) in patients with metastatic urothelial carcinoma (mUC) that progressed on checkpoint inhibition (CPI). PATIENTS AND METHODS: A phase I expansion cohort of patients with mUC who received prior CPI was treated with cabozantinib 40 mg/day and nivolumab 3 mg/kg every 2 weeks until disease progression/unacceptable toxicity. The primary goal was objective response rate (ORR) per RECIST v.1.1. Secondary objectives included progression-free survival (PFS), duration of response (DoR), overall survival (OS), safety, and tolerability. RESULTS: Twenty-nine out of 30 patients enrolled were evaluable for efficacy. Median follow-up was 22.2 months. Most patients (86.7%) received prior chemotherapy and all patients received prior CPI (median seven cycles). ORR was 16.0%, with one complete response and three partial responses (PR). Among 4 responders, 2 were primary refractory, 1 had a PR, and 1 had stable disease on prior CPI. Median DoR was 33.5 months [95% confidence interval (CI), 3.7-33.5], median PFS was 3.6 months (95% CI, 2.1-5.5), and median OS was 10.4 months (95% CI, 5.8-19.5). CaboNivo decreased immunosuppressive subsets such as regulatory T cells (Tregs) and increased potential antitumor immune subsets such as nonclassical monocytes and effector T cells. A lower percentage of monocytic myeloid-derived suppressor cells (M-MDSC) and polymorphonuclear MDSCs, lower CTLA-4 and TIM-3 expression on Tregs, and higher effector CD4+ T cells at baseline were associated with better PFS and/or OS. CONCLUSIONS: CaboNivo was clinically active, well tolerated, and favorably modulated peripheral blood immune subsets in patients with mUC refractory to CPI.

MiR-6924-5p-rich exosomes derived from genetically modified Scleraxis-overexpressing PDGFRα(+) BMMSCs as novel nanotherapeutics for treating osteolysis during tendon-bone healing and improving healing strength.

Osteolysis at the tendon-bone interface can impair pullout strength during tendon-bone healing and lead to surgery failure, but the effects of clinical treatments are not satisfactory. Mesenchymal stem cell (MSC)-derived exosomes have been used as potent and feasible natural nanocarriers for drug delivery and have been proven to enhance tendon-bone healing strength, indicating that MSC-derived exosomes could be a promising therapeutic strategy. In this study, we explored Scleraxis (Scx) dynamically expressed in PDGFRα(+) bone marrow-derived mesenchymal stem cells (BMMSCs) during natural tendon-bone healing. Then, we investigated the role of PDGFRα(+) BMMSCs in tendon-bone healing after Scx overexpression as well as the underlying mechanisms. Our data demonstrated that Scx-overexpressing PDGFRα(+) BMMSCs (BMMSCScx) could efficiently inhibit peritunnel osteolysis and enhance tendon-bone healing strength by preventing osteoclastogenesis in an exosomes-dependent manner. Exosomal RNA-seq revealed that the abundance of a novel miRNA, miR-6924-5p, was highest among miRNAs. miR-6924-5p could directly inhibit osteoclast formation by binding to the 3'-untranslated regions (3'UTRs) of OCSTAMP and CXCL12. Inhibition of miR-6924-5p expression reversed the prevention of osteoclastogenic differentiation by BMMSCScx derived exosomes (BMMSCScx-exos). Local injection of BMMSCScx-exos or miR-6924-5p dramatically reduced osteoclast formation and improved tendon-bone healing strength. Furthermore, delivery of miR-6924-5p efficiently inhibited the osteoclastogenesis of human monocytes. In brief, our study demonstrates that BMMSCScx-exos or miR-6924-5p could serve as a potential therapy for the treatment of osteolysis during tendon-bone healing and improve the outcome.

Optimal Lymph Node Staging System in Evaluating Prognosis of Gallbladder Carcinoma: A Multi-institutional Study.

BACKGROUND: Lymph node (LN) involvement is a critical prognostic factor in patients with gallbladder carcinoma (GBC). Controversy exists regarding optimal categorization of nodal metastasis status, including anatomical location of positive nodes (AJCC 7th N staging), number of metastatic lymph nodes (NMLN), log odds of metastatic LNs (LODDS), and lymph node ratio (LNR). METHODS: Patients who underwent curative-intent resection for GBC from six Chinese tertiary hospitals between 2008 and 2013 were analyzed retrospectively. The relative discriminative abilities of the different LN staging systems were assessed by different models including the tree-augmented naïve Bayesian (TAN) model, Cox proportional hazards regression model, and binary logistic regression model. RESULTS: A total of 226 patients were involved in this cohort. Based on the TAN model and composite importance measures, the most important factor affecting the prognosis in the different LN staging systems was NMLN. Among the four TAN models which were built with 4 metastatic LN markers and baseline variables, the accuracy of the NMLN-based prognostic model was 88.15%, higher than 7th N staging (86.44%), LNR (87.34%), and LODDS (85.19%). The Cox model based on NMLN (C-index: 0.763, AIC: 1371.62) had a higher fitness than the others (7th N staging C-index: 0.756, AIC: 1375.51; LNR C-index: 0.759, AIC: 1378.82; LODDS C-index 0.748, AIC: 1390.99). The AUCs of different staging binary logistic regression models were NMLN (0.872), LNR (0.872), 7th N staging (0.869) and LODDS (0.856), respectively. CONCLUSIONS: NMLN was the optimal LN staging system in evaluating prognosis of GBC.

Impact of Concomitant Impairments of the Left and Right Ventricular Myocardial Strain on the Prognoses of Patients With ST-Elevation Myocardial Infarction.

Background: The impact of concomitant impairments of left and right ventricular (LV and RV) strain on the long-term prognosis of acute ST-elevation myocardial infarction (STEMI) is not clear. Methods: We analyzed CMR images and followed up 420 first STEMI patients from the EARLY Assessment of MYOcardial Tissue Characteristics by CMR in STEMI (EARLY-MYO-CMR) registry (NCT03768453). These patients received timely primary percutaneous coronary intervention (PCI) within 12 h and CMR examination within 1 week (median, 5 days; range, 2-7 days) after infarction. Global longitudinal strain (GLS), global radial strain (GRS), and global circumferential strain (GCS) of both ventricles were measured based on CMR cine images. Conventional CMR indexes were also assessed. Primary clinical outcome was composite major adverse cardiac and cerebrovascular events (MACCEs) including cardiovascular death, re-infarction, re-hospitalization for heart failure and stroke. In addition, CMR data from 40 people without apparent heart disease were used as control group. Results: Compared to controls, both LV and RV strains were remarkably reduced in STEMI patients. During follow-up (median: 52 months, interquartile range: 29-68 months), 80 patients experienced major adverse cardiac and cerebrovascular events (MACCEs) including cardiovascular death, re-infarction, heart failure, and stroke. LV-GCS > -11.20% was an independent predictor of MACCEs (P < 0.001). RV-GRS was the only RV strain index that could effectively predict the risk of MACCEs (AUC = 0.604, 95% CI [0.533, 0.674], P = 0.004). Patient with RV-GRS ≤ 38.79% experienced more MACCEs than those with preserved RV-GRS (log rank P < 0.001). Moreover, patients with the concomitant decrease of LV-GCS and RV-GRS were more likely to experience MACCEs than patients with decreased LV-GCS alone (log rank P = 0.010). RV-GRS was incremental to LV-GCS for the predictive power of MACCEs (continuous NRI: 0.327; 95% CI: 0.095-0.558; P = 0.006). Finally, tobacco use (P = 0.003), right coronary artery involvement (P = 0.002), and LV-GCS > -11.20% (P = 0.012) was correlated with lower RV-GRS. Conclusions: The concomitant decrease of LV and RV strain is associated with a worse long-term prognosis than impaired LV strain alone. Combination assessment of both LV and RV strain indexes could improve risk stratification of patients with STEMI. Trial Registration: ClinicalTrials.gov, NCT03768453. Registered 7 December 2018 - Retrospectively registered, https://clinicaltrials.gov/ct2/show/NCT03768453.

Critical biodegradation process of a widely used surfactant in the water environment: dodecyl benzene sulfonate (DBS).

Sodium dodecyl benzene sulfonate (DBS) is a widely used surfactant that is now found extensively in water bodies because of anthropogenic emissions. Since the degradation of DBS in the environment mainly relies on microorganisms, it is essential to study the mechanism by which DBS is biodegraded. In this study, Chlorella vulgaris was used to research the biodegradation process of DBS. The C. vulgaris suspension was centrifuged to remove the supernatant, then secondary deionized water and DBS were added to the C. vulgaris. And the intermediate products were detected in real time by electrospray ionization mass spectrometry (ESI-MS). Some novel intermediate products, including 4-sodium sulfophenyldodecanoate acid and its homologs, were detected that had not been mentioned in previous studies. In this work, the process of DBS degradation was indicated, which consisted of three main steps: chain-shorting oxidation, ring-opening oxidation of benzene rings, and degradation of small molecules. By investigating the process of DBS degradation by C. vulgaris, we were able to propose a preliminary mechanism of DBS biodegradation, which is of great significance for research on the migration and conversion of surfactants in the environment.

Identification of a Hypoxia-Related Gene Model for Predicting the Prognosis and Formulating the Treatment Strategies in Kidney Renal Clear Cell Carcinoma.

PURPOSE: The present study aimed to establish a hypoxia related genes model to predict the prognosis of kidney clear cell carcinoma (KIRC) patients using data accessed from The Cancer Genome Atlas (TCGA) database and International Cancer Genome Consortium (ICGC) database. METHODS: Patients' data were downloaded from the TCGA and ICGC databases, and hypoxia related genes were accessed from the Molecular Signatures Database. The differentially expressed genes were evaluated and then the differential expressions hypoxia genes were screened. The TCGA cohort was randomly divided into a discovery TCGA cohort and a validation TCGA cohort. The discovery TCGA cohort was used for constructing the hypoxia genes risk model through Lasso regression, univariate and multivariate Cox regression analysis. Receiver operating characteristic (ROC) curves were used to assess the reliability and sensitivity of our model. Then, we established a nomogram to predict the probable one-, three-, and five-year overall survival rates. Lastly, the Tumor Immune Dysfunction and Exclusion (TIDE) score of patients was calculated. RESULTS: We established a six hypoxia-related gene prognostic model of KIRC patients in the TCGA database and validated in the ICGC database. The patients with high riskscore present poorer prognosis than those with low riskscore in the three TCGA cohorts and ICGC cohort. ROC curves show our six-gene model with a robust predictive capability in these four cohorts. In addition, we constructed a nomogram for KIRC patients in the TCGA database. Finally, the high risk-group had a high TIDE score than the patients with low riskscore. CONCLUSIONS: We established a six hypoxia-related gene risk model for independent prediction of the prognosis of KIRC patients was established and constructed a robust nomogram. The different riskscores might be a biomarker for immunotherapy strategy.