Long-term prognosis of patients with gallbladder carcinoma after curative-intent resection based on changes in the ratio of carbohydrate antigen 19-9 to total bilirubin (CA19-9/TB): a multicenter retrospective cohort study.

BACKGROUND: The prognostic value of carbohydrate antigen 19-9 (CA19-9) is known to be affected by elevated bilirubin levels in patients with gallbladder carcinoma (GBC). The clinical significance of changes in the ratio of CA19-9 levels to total bilirubin (TB) levels in patients with GBC after curative-intent resection remains unknown. The aim of this study was to determine the prognostic value of changes in preoperative and postoperative CA19-9/TB ratio in these patients. METHODS: Prospectively colleced data on consecutive patients who underwent curative-intent resection for GBC between January 2015 and December 2020 stored in a multicenter database from 10 hospitals were analysed in this retrospective cohort study. Based on the adjusted CA19-9 defined as the ratio of CA19-9 to TB, and using 2×103 U/µmol as the upper normal value, patients were divided into a normal group (with normal preoperative and postoperative adjusted CA19-9), a normalization group (with abnormal preoperative but normal postoperative adjusted CA19-9), and a non-normalization group (with abnormal postoperative adjusted CA19-9). The primary outcomes were overall survival (OS) and recurrence-free survival (RFS). The log-rank test was used to compare OS and RFS among the groups. The Cox regression model was used to determine factors independently associated with OS and RFS. RESULTS: The normal group (n=179 patients) and the normalization group (n=73 patients) had better OS and RFS than the non-normalization group (n=65 patients) (the 3-year OS rates 72.0%, 58.4% and 24.2%, respectively; the RFS rates 54.5%, 25.5% and 11.8%, respectively; both P<0.001). There were no significant differences between the normal and the normalization groups in OS and RFS (OS, P=0.255; RFS, P=0.130). Cox regression analysis confirmed that the non-normalization group was independently associated with worse OS and RFS. Subgroup analysis revealed that the non-normalization group of patients who received adjuvant therapy had significantly improved OS and RFS as compared to those who did not receive adjuvant therapy (OS, P=0.025; RFS, P=0.003). CONCLUSIONS: Patients with GBC who underwent curative-intent surgical resection with postoperative abnormal levels of adjusted CA19-9 (the CA19-9/TB ratio) were associated with poorer long-term survival outcomes. Adjuvant therapy after surgery improved the long-term outcomes of these patients.

Development of a model based on the age-adjusted Charlson comorbidity index to predict survival for resected perihilar cholangiocarcinoma.

BACKGROUND: Perihilar cholangiocarcinoma (pCCA) has a poor prognosis and urgently needs a better predictive method. The predictive value of the age-adjusted Charlson comorbidity index (ACCI) for the long-term prognosis of patients with multiple malignancies was recently reported. However, pCCA is one of the most surgically difficult gastrointestinal tumors with the poorest prognosis, and the value of the ACCI for the prognosis of pCCA patients after curative resection is unclear. AIM: To evaluate the prognostic value of the ACCI and to design an online clinical model for pCCA patients. METHODS: Consecutive pCCA patients after curative resection between 2010 and 2019 were enrolled from a multicenter database. The patients were randomly assigned 3:1 to training and validation cohorts. In the training and validation cohorts, all patients were divided into low-, moderate-, and high-ACCI groups. Kaplan-Meier curves were used to determine the impact of the ACCI on overall survival (OS) for pCCA patients, and multivariate Cox regression analysis was used to determine the independent risk factors affecting OS. An online clinical model based on the ACCI was developed and validated. The concordance index (C-index), calibration curve, and receiver operating characteristic (ROC) curve were used to evaluate the predictive performance and fit of this model. RESULTS: A total of 325 patients were included. There were 244 patients in the training cohort and 81 patients in the validation cohort. In the training cohort, 116, 91 and 37 patients were classified into the low-, moderate- and high-ACCI groups. The Kaplan-Meier curves showed that patients in the moderate- and high-ACCI groups had worse survival rates than those in the low-ACCI group. Multivariable analysis revealed that moderate and high ACCI scores were independently associated with OS in pCCA patients after curative resection. In addition, an online clinical model was developed that had ideal C-indexes of 0.725 and 0.675 for predicting OS in the training and validation cohorts. The calibration curve and ROC curve indicated that the model had a good fit and prediction performance. CONCLUSION: A high ACCI score may predict poor long-term survival in pCCA patients after curative resection. High-risk patients screened by the ACCI-based model should be given more clinical attention in terms of the management of comorbidities and postoperative follow-up.

Textbook outcomes in liver surgery for gallbladder cancer patients treated with curative-intent resection: a multicenter observational study.

BACKGROUND: Cholecystectomy, hepatectomy, and lymphadenectomy are recommended as the curative treatment for resectable gallbladder cancer (GBC). Textbook outcomes in liver surgery (TOLS) is a novel composite measure that has been defined by expert consensus to represent the optimal postoperative course after hepatectomy. This study aimed to determine the incidence of TOLS and the independent predictors associated with TOLS after curative-intent resection in GBC patients. METHODS: All consecutive GBC patients who underwent curative-intent resection between 2014 and 2020 were enrolled from a multicenter database from 11 hospitals as the training and the internal testing cohorts, and Southwest Hospital as the external testing cohort. TOLS was defined as no intraoperative grade greater than or equal to 2 incidents, no grade B/C postoperative bile leaks, no postoperative grade B/C liver failure, no 90-day postoperative major morbidity, no 90-day readmission, no 90-day mortality after hospital discharge, and R0 resection. Independent predictors of TOLS were identified using logistic regression and were used to construct the nomogram. The predictive performance was assessed using the area under the curve and calibration curves. RESULTS: TOLS was achieved in 168 patients (54.4%) and 74 patients (57.8%) from the training and internal testing cohorts, and the external testing cohort, respectively. On multivariate analyses, age less than or equal to 70 years, absence of preoperative jaundice (total bilirubin≤3 mg/dl), T1 stage, N0 stage, wedge hepatectomy, and no neoadjuvant therapy were independently associated with TOLS. The nomogram that incorporated these predictors demonstrated excellent calibration and good performance in both the training and external testing cohorts (area under the curve: 0.741 and 0.726). CONCLUSIONS: TOLS was only achieved in approximately half of GBC patients treated with curative-intent resection, and the constructed nomogram predicted TOLS accurately.

Gastroduodenal artery disconnection during liver transplantation decreases non-anastomotic stricture incidence.

BACKGROUND: The hepatic artery is the only blood source nourishing the biliary duct and associated with biliary complication after liver transplantation (LT). Gastroduodenal artery (GDA) disconnection increased proper hepatic artery flow. Whether this procedure attenuates biliary non-anastomotic stricture (NAS) is not clear. METHODS: A total of 241 patients with LT were retrospectively analyzed. The patients were divided into the GDA disconnection (GDA-) and GDA preservation (GDA+) groups. Propensity score matching (PSM) was administrated to reduce bias. Logistic regression was conducted to analyze risk factors for biliary NAS before and after PSM. Postoperative complications were compared. Kaplan-Meier survival analysis and log-rank tests were performed to compare overall survival. RESULTS: In all, 99 patients (41.1%) underwent GDA disconnection, and 49 (20.3%) developed NAS. Multivariate logistic regression revealed that GDA preservation (OR = 2.24, 95% CI: 1.11-4.53; P = 0.025) and model for end-stage liver disease (MELD) score > 15 (OR = 2.14, 95% CI: 1.12-4.11; P = 0.022) were risk factors for biliary NAS. PSM provided 66 pairs using 1:2 matching method, including 66 GDA disconnection and 99 GDA preservation patients. Multivariate logistic regression after PSM also showed that GDA preservation (OR = 3.15, 95% CI: 1.26-7.89; P = 0.014) and MELD score > 15 (OR = 2.41, 95% CI: 1.08-5.36; P = 0.031) were risk factors for NAS. When comparing complications between the two groups, GDA preservation was associated with a higher incidence of biliary NAS before and after PSM (P = 0.031 and 0.017, respectively). In contrast, other complications including early allograft dysfunction (P = 0.620), small-for-size graft syndrome (P = 0.441), abdominal hemorrhage (P = 1.000), major complications (Clavien-Dindo grade ≥ 3, P = 0.318), and overall survival (P = 0.088) were not significantly different between the two groups. CONCLUSIONS: GDA disconnection during LT ameliorates biliary NAS incidence and may be recommended for application in clinical practice.

Arctigenin impairs UBC12 enzyme activity and cullin neddylation to attenuate cancer cells.

Neddylation is a type of posttranslational protein modification that has been observed to be overactivated in various cancers. UBC12 is one of two key E2 enzymes in the neddylation pathway. Reports indicate that UBC12 deficiency may suppress lung cancer cells, such that UBC12 could play an important role in tumor progression. However, systematic studies regarding the expression profile of UBC12 in cancers and its relationship to cancer prognosis are lacking. In this study, we comprehensively analyzed UBC12 expression in diverse cancer types and found that UBC12 is markedly overexpressed in most cancers (17/21), a symptom that negatively correlates with the survival rates of cancer patients, including gastric cancer. These results demonstrate the suitability of UBC12 as a potential target for cancer treatment. Currently, no effective inhibitor targeting UBC12 has been discovered. We screened a natural product library and found, for the first time, that arctigenin has been shown to significantly inhibit UBC12 enzyme activity and cullin neddylation. The inhibition of UBC12 enzyme activity was newly found to contribute to the effects of arctigenin on suppressing the malignant phenotypes of cancer cells. Furthermore, we performed proteomics analysis and found that arctigenin intervened with cullin downstream signaling pathways and substrates, such as the tumor suppressor PDCD4. In summary, these results demonstrate the importance of UBC12 as a potential therapeutic target for cancer treatment, and, for the first time, the suitability of arctigenin as a potential compound targeting UBC12 enzyme activity. Thus, these findings provide a new strategy for inhibiting neddylation-overactivated cancers.

Impact of perioperative blood transfusion on long-term survival in patients with different stages of perihilar cholangiocarcinoma treated with curative resection: A multicentre propensity score matching study.

Background & aim: The association of perioperative blood transfusion (PBT) with long-term survival in perihilar cholangiocarcinoma (pCCA) patients after surgical resection with curative intent is controversial and may differ among different stages of the disease. This study aimed to investigate the impact of PBT on long-term survival of patients with different stages of pCCA. Methods: Consecutive pCCA patients from three hospitals treated with curative resection from 2012 to 2019 were enrolled and divided into the PBT and non-PBT groups. Propensity score matching (PSM) was used to balance differences in baseline characteristics between the PBT and non-PBT groups. Kaplan-Meier curves and log-rank test were used to compare overall survival (OS) and recurrence-free survival (RFS) between patients with all tumor stages, early stage (8th AJCC stage I), and non-early stage (8th AJCC stage II-IV) pCCA in the PBT and non-PBT groups. Cox regression analysis was used to determine the impact of PBT on OS and RFS of these patients. Results: 302 pCCA patients treated with curative resection were enrolled into this study. Before PSM, 68 patients (22 patients in the PBT group) were in the early stage and 234 patients (108 patients in the PBT group) were in the non-early stage. Patients with early stage pCCA in the PBT group had significantly lower OS and RFS rates than those in the non-PBT group. However, there were with no significant differences between the 2 groups with all tumor stages and non-early stage pCCA. After PSM, there were 18 matched pairs of patients with early stage and 72 matched pairs of patients with non-early stage. Similar results were obtained in the pre- and post-PSM cohorts: patients with early stage pCCA in the PBT group showed significantly lower OS and RFS rates than those in the non-PBT group, but there were no significant differences between the 2 groups for patients with all tumor stages and non-early stage pCCA. Cox regression analysis demonstrated that PBT was independently associated with worse OS and RFS for patients with early stage pCCA. Conclusions: PBT had a negative impact on long-term survival in patients with early stage pCCA after curative resection, but not in patients with non-early stage pCCA.

Development and Validation of a Prognostic Model to Predict Recurrence-Free Survival After Curative Resection for Perihilar Cholangiocarcinoma: A Multicenter Study.

Background: Recurrence is the main cause of death in perihilar cholangiocarcinoma (pCCA) patients after surgery. Identifying patients with a high risk of recurrence is important for decision-making regarding neoadjuvant therapy to improve long-term outcomes. Aim: The objective of this study was to develop and validate a prognostic model to predict recurrence-free survival (RFS) after curative resection of pCCA. Methods: Patients following curative resection for pCCA from January 2008 to January 2016 were identified from a multicenter database. Using random assignment, 70% of patients were assigned to the training cohort, and the remaining 30% were assigned to the validation cohort. Independent predictors of RFS after curative resection for pCCA were identified and used to construct a prognostic model. The predictive performance of the model was assessed using calibration curves and the C-index. Results: A total of 341 patients were included. The median overall survival (OS) was 22 months, and the median RFS was 14 months. Independent predictors associated with RFS included lymph node involvement, macrovascular invasion, microvascular invasion, maximum tumor size, tumor differentiation, and carbohydrate antigen 19-9. The model incorporating these factors to predict 1-year RFS demonstrated better calibration and better performance than the 8th American Joint Committee on Cancer (AJCC) staging system in both the training and validation cohorts (C-indexes: 0.723 vs. 0.641; 0.743 vs. 0.607). Conclusions: The prognostic model could identify patients at high risk of recurrence for pCCA to inform patients and surgeons, help guide decision-making for postoperative adjuvant therapy, and improve survival.

Postoperative morbidity adversely impacts oncological prognosis after curative resection for hilar cholangiocarcinoma.

BACKGROUND: Postoperative morbidity after curative resection for hilar cholangiocarcinoma (HCCA) is common; however, whether it has an impact on oncological prognosis is unknown. AIM: To evaluate the influence of postoperative morbidity on tumor recurrence and mortality after curative resection for HCCA. METHODS: Patients with recently diagnosed HCCA who had undergone curative resection between January 2010 and December 2017 at The First Affiliated Hospital of Army Medical University in China were enrolled. The independent risk factors for morbidity in the 30 d after surgery were investigated, and links between postoperative morbidity and patient characteristics and outcomes were assessed. Postoperative morbidities were divided into five grades based on the Clavien-Dindo classification, and major morbidities were defined as Clavien-Dindo ≥ 3. Univariate and multivariate Cox regression analyses were used to evaluate the risk factors for recurrence-free survival (RFS) and overall survival (OS). RESULTS: Postoperative morbidity occurred in 146 out of 239 patients (61.1%). Multivariate logistic regression revealed that cirrhosis, intraoperative blood loss > 500 mL, diabetes mellitus, and obesity were independent risk factors. Postoperative morbidity was associated with decreased OS and RFS (OS: 18.0 mo vs 31.0 mo, respectively, P = 0.003; RFS: 16.0 mo vs 26.0 mo, respectively, P = 0.002). Multivariate Cox regression analysis indicated that postoperative morbidity was independently associated with decreased OS [hazard ratios (HR): 1.557, 95% confidence interval (CI): 1.119-2.167, P = 0.009] and RFS (HR: 1.535, 95%CI: 1.117-2.108, P = 0.008). Moreover, major morbidity was independently associated with decreased OS (HR: 2.175; 95%CI: 1.470-3.216, P < 0.001) and RFS (HR: 2.054; 95%CI: 1.400-3.014, P < 0.001) after curative resection for HCCA. CONCLUSION: Postoperative morbidity (especially major morbidity) may be an independent risk factor for unfavorable prognosis in HCCA patients following curative resection.

Evaluation of Four Lymph Node Classifications for the Prediction of Survival in Hilar Cholangiocarcinoma.

BACKGROUND: An important prognostic indicator of hilar cholangiocarcinoma (HCCA) in patients after surgery is metastasis of lymph nodes (LN). However, there are many types of LN staging systems to the issue of a better determination of the prognosis of patients through the lymphatic staging system which needs research. Based on the above, we tried to re-evaluate the staging system of HCCA LNs. We compared the American Joint Committee on Cancer (AJCC), number of metastatic LNs (MLN), ratio of LN (LNR), and log odds of MLNs (LODDS) in individuals undergoing curative resection to determine the best LN staging system. METHODS: In the current study, we retrospectively analyzed 229 patients undergoing curative resection. We evaluated the impact of the stage of AJCC pN, LNR, LODDS, and MLN on OS (overall survival) and RFS (recurrence-free survival). According to the curve of receiver operating characteristic (ROC), we compared the predictive capacity of different staging systems of LN for survival and recurrence. RESULTS: Multivariate analysis results revealed that LODDS > - 0.45 (95% CI = 1.115-2.709, P = 0.015; 95% CI = 1.187-2.780, P = 0.006) are independent risk factors affecting OS and RFS, respectively. Compared with LN status, AJCC pN stage, MLN, and LNR, the variable having the highest area under the ROC curve (AUC) was LODDS when predicting 1-year, 3-year, and 5-year OS and RFS. CONCLUSION: This study shows that metastasis of LNs is a key indicator for predicting patient death and recurrence. Among them, LODDS is the best LN staging system for the prognostic evaluation of HCCA patients after surgery. Clinicians can incorporate LODDS into HCCA patient lymphatic staging system for a more accurate prognosis of HCCA patients post-surgery.

Assessment of the prognostic value of the neutrophil-to-lymphocyte ratio and platelet-to-lymphocyte ratio in perihilar cholangiocarcinoma patients following curative resection: A multicenter study of 333 patients.

Background & Aims: Tumor-associated chronic inflammation has been determined to play a crucial role in tumor progression, angiogenesis and immunosuppression. The objective of this study was to assess the prognostic value of the neutrophil-to-lymphocyte ratio (NLR) and platelet-to-lymphocyte ratio (PLR) in perihilar cholangiocarcinoma (pCCA) patients following curative resection. Methods: Consecutive pCCA patients following curative resection at 3 Chinese hospitals between 2014 and 2018 were included. The NLR was defined as the ratio of neutrophil count to lymphocyte count. PLR was defined as the ratio of platelet count to lymphocyte count. The optimal cutoff values of preoperative NLR and PLR were determined according to receiver operating characteristic (ROC) curves for the prediction of 1-year overall survival (OS), and all patients were divided into high- and low-risk groups. Kaplan-Meier curves and Cox regression models were used to investigate the relationship between values of NLR and PLR and values of OS and recurrence-free survival (RFS) in pCCA patients. The usefulness of NLR and PLR in predicting OS and RFS was evaluated by time-dependent ROC curves. Results: A total of 333 patients were included. According to the ROC curve for the prediction of 1-year OS, the optimal cutoff values of preoperative NLR and PLR were 1.68 and 113.1, respectively, and all patients were divided into high- and low-risk groups. The 5-year survival rates in the low-NLR (<1.68) and low-PLR groups (<113.1) were 30.1% and 29.4%, respectively, which were significantly higher than the rates of 14.9% and 3.3% in the high-NLR group (≥1.68) and high-PLR group (≥113.1), respectively. In multivariate analysis, high NLR and high PLR were independently associated with poor OS and RFS for pCCA patients. The time-dependent ROC curve revealed that both NLR and PLR were ideally useful in predicting OS and RFS for pCCA patients. Conclusions: This study found that both NLR and PLR could be used to effectively predict long-term survival in patients with pCCA who underwent curative resection.

Study on a Novel Cold-Active and Halotolerant Monoacylglycerol Lipase Widespread in Marine Bacteria Reveals a New Group of Bacterial Monoacylglycerol Lipases Containing Unusual C(A/S)HSMG Catalytic Motifs.

Monoacylglycerol lipases (MGLs) are present in all domains of life. However, reports on bacterial MGLs are still limited. Until now, reported bacterial MGLs are all thermophilic/mesophilic enzymes from warm terrestrial environments or deep-sea hydrothermal vent, and none of them originates from marine environments vastly subject to low temperature, high salts, and oligotrophy. Here, we characterized a novel MGL, GnMgl, from the marine cold-adapted and halophilic bacterium Glaciecola nitratireducens FR1064T. GnMgl shares quite low sequence similarities with characterized MGLs (lower than 31%). GnMgl and most of its bacterial homologs harbor a catalytic Ser residue located in the conserved C(A/S)HSMG motif rather than in the typical GxSxG motif reported on other MGLs, suggesting that GnMgl-like enzymes might be different from reported MGLs in catalysis. Phylogenetic analysis suggested that GnMgl and its bacterial homologs are clustered as a separate group in the monoglyceridelipase_lysophospholipase family of the Hydrolase_4 superfamily. Recombinant GnMgl has no lysophospholipase activity but could hydrolyze saturated (C12:0-C16:0) and unsaturated (C18:1 and C18:2) MGs and short-chain triacylglycerols, displaying distinct substrate selectivity from those of reported bacterial MGLs. The substrate preference of GnMgl, predicted to be a membrane protein, correlates to the most abundant fatty acids within the strain FR1064T, suggesting the role of GnMgl in the lipid catabolism in this marine bacterium. In addition, different from known bacterial MGLs that are all thermostable enzymes, GnMgl is a cold-adapted enzyme, with the maximum activity at 30°C and retaining 30% activity at 0°C. GnMgl is also a halotolerant enzyme with full activity in 3.5M NaCl. The cold-adapted and salt-tolerant characteristics of GnMgl may help its source strain FR1064T adapt to the cold and saline marine environment. Moreover, homologs to GnMgl are found to be abundant in various marine bacteria, implying their important physiological role in these marine bacteria. Our results on GnMgl shed light on marine MGLs.

Author Correction: Invasion of white matter tracts by glioma stem cells is regulated by a NOTCH1-SOX2 positive-feedback loop.

The original and corrected figures are shown in the accompanying Author Correction.

Association between socio-demographic factors, coping style, illness perceptions and preference for disclosure/nondisclosure of diagnosis in Chinese patients with hepatocellular carcinoma.

We explored the association between socio-demographic factors, coping style, illness perceptions and preferences for disclosure/nondisclosure of cancer diagnosis in 384 Chinese patients with hepatocellular carcinoma. We found that (1) 69.3 percent of the patients preferred disclosure and (2) multivariate analysis showed that four variables were significantly positively associated with preference for disclosure, including active emotional-focused coping style, illness perceptions of personal control, chronic infection of hepatitis B virus, and educational level, whereas perceived emotional impact of illness and objective social support (mainly family support) were significantly associated with preference for nondisclosure. The findings provide useful information for understanding patients' preferences for disclosure/nondisclosure of cancer diagnosis from a psychosocial perspective.

KLF4 functions as an oncogene in promoting cancer stem cell-like characteristics in osteosarcoma cells.

Despite more effective chemotherapy combined with limb-salvage surgery for the osteosarcoma treatment, survival rates for osteosarcoma patients have stagnated over the past three decades due to the poor prognosis. Osteosarcoma cancer stem cells (OSCs) are responsible for the growth and metastasis of osteosarcoma. The existence of OSCs offers a theoretical explanation for therapeutic failures including tumor recurrence, metastasis, and drug resistance. Understanding the pathways that regulate properties of OSCs may shed light on mechanisms that lead to osteosarcoma and suggest better modes of treatment. In this study, we showed that the expression level of Kruppel-like factor 4 (KLF4) is highly associated with human osteosarcoma cancer stemness. KLF4-overexpressed osteosarcoma cells displayed characteristics of OSCs: increased sphere-forming potential, enhanced levels of stemness-associated genes, great chemoresistance to adriamycin and CDDP, as well as more metastasis potential. Inversely, KLF4 knockdown could reduce colony formation in vitro and inhibit tumorigenesis in vivo, supporting an oncogenic role for KLF4 in osteosarcoma pathogenesis. Furthermore, KLF4 was shown to activate the p38 MAPK signaling pathway to promote cancer stemness. Altogether, our studies uncover an essential role for KLF4 in regulation of OSCs and identify KLF4-p38 MAPK axis as a potential therapeutic target for osteosarcoma treatment.

Invasion of white matter tracts by glioma stem cells is regulated by a NOTCH1-SOX2 positive-feedback loop.

Early invasive growth along specific anatomical structures, especially the white matter tract, is regarded as one of the main causes of poor therapeutic outcome of people with gliomas. We show that some glioma stem cells (GSCs) are preferentially located along white matter tracts, which exhibit a demyelinated phenotype, at the invasive frontier of glioma tissues. These GSCs are CD133+Notch1+, whereas the nerve fibers express the Notch ligand Jagged1. The Notch-induced transcription factor Sox9 promotes the transcription of SOX2 and the methylation level of the NOTCH1 promoter is attenuated by the upregulation of SOX2 to reinforce NOTCH1 expression in GSCs. This positive-feedback loop in a cohort of glioma subjects is correlated with a poor prognosis. Inhibition of Notch signaling attenuates the white-matter-tract tropism of GSCs. These findings provide evidence indicating that the NOTCH1-SOX2 positive-feedback loop controls GSC invasion along white matter tracts.

The impact of insulin pump therapy to oxidative stress in patients with diabetic nephropathy.

BACKGROUND: The oxidative stress resulting from increased production of ROS plays a crucial role in the development of diabetic complications. We aim to explore the relationships between oxidative stress, diabetic nephropathy (DN) and short-term insulin pump intensive therapy (insulin therapy). METHODS: Levels of 8-hydroxy-deoxyguanosine (8-OHdG), 3-nitrotyrosine (3-NT), glutathione (GSH), superoxide dismutase (SOD) and Interleukin-6 (IL-6) were estimated before and after 2 weeks of insulin therapy in normal group (NC) and type 2 diabetic (DM) with normal albuminuria (NA), microalbuminuria (MA) and clinical albuminuria (CA). RESULTS: In DM group, levels of 8-OHdG and 3-NT were higher than those in NC group (P < 0.05); GSH and SOD were lower (P < 0.05). And their levels changed with urine albumin-creatinine ratio (P < 0.05). After insulin therapy, these derangements were significantly ameliorated and the changes in NA and MA groups were more significant than CA group (P < 0.05). Correlation analysis showed glycated hemoglobin, the course of disease, the HOME-IR and fasting plasma glucose were positively correlated with 8-OHdG and 3-NT, but negatively correlated with GSH and SOD. CONCLUSIONS: The oxidative stress gradually increased with the magnitude of DN, and insulin pump intensive therapy can significantly ameliorate the derangements in the early stage of DN. Trial registration NCT03174821.

Effect of Group Density on the Physiology and Aggressive Behavior of Male Brandt's Voles (Lasiopodomys brandtii).

Xin Dai, Ling-Yu Zhou, Jie-Xia Cao, Yan-Qi Zhang, Feng-Ping Yang, Ai-Qin Wang, Wan-Hong Wei, and Sheng-Mei Yang (2018) Population density is well known to influence animal physiology and behavior. How population density affects the aggressive behavior of the Brandt's vole (Lasiopodomys brandtii) is, however, little known. The aim of this study was to investigate the effect of group density on physiologic responses and aggressive behavior of male Brandt's voles and their potential underlying neuro-mechanism. The results show that increasing group density led to elevated serum corticosterone levels and increased spleen weight; it also induced more male-male aggressive behavior. By contrast, it had a negative effect on body growth and the weight of testis and epididymis. Aging also increased male-male aggressive behavior. Higher density reduced mRNA levels of tryptophan hydroxylase 2 (TPH2), 5-hydroxytryptamine receptor 1A (5HT1A), and 5-hydroxytryptamine receptor 1B (5HT1B) in the amygdala and the dorsal raphe nucleus (DRN). Our results demonstrate that higher population density can intensify stress reactions and male-male aggressive behavior in Brandt's voles at the price of inhibiting body growth and reproduction. Serotonergic systems in the amygdala and the DRN may take part in the control of aggressive behavior among male voles. Our results provide novel insights into the neuro-mechanism underlying the influence of population density on aggressive behavior in Brandt's vole, and imply that aggressive behavior may play an important role in the population fluctuation of the animal.

Structural and Mechanistic Insights into the Improvement of the Halotolerance of a Marine Microbial Esterase by Increasing Intra- and Interdomain Hydrophobic Interactions.

Halotolerant enzymes are beneficial for industrial processes requiring high salt concentrations and low water activity. Most halophilic proteins are evolved to have reduced hydrophobic interactions on the surface and in the hydrophobic cores for their haloadaptation. However, in this study, we improved the halotolerance of a thermolabile esterase, E40, by increasing intraprotein hydrophobic interactions. E40 was quite unstable in buffers containing more than 0.3 M NaCl, and its kcat and substrate affinity were both significantly reduced in 0.5 M NaCl. By introducing hydrophobic residues in loop 1 of the CAP domain and/or α7 of the catalytic domain in E40, we obtained several mutants with improved halotolerance, and the M3 S202W I203F mutant was the most halotolerant. ("M3" represents a mutation in loop 1 of the CAP domain in which residues R22-K23-T24 of E40 are replaced by residues Y22-K23-H24-L25-S26 of Est2.) Then we solved the crystal structures of the S202W I203F and M3 S202W I203F mutants to reveal the structural basis for their improved halotolerance. Structural analysis revealed that the introduction of hydrophobic residues W202 and F203 in α7 significantly improved E40 halotolerance by strengthening intradomain hydrophobic interactions of F203 with W202 and other residues in the catalytic domain. By further introducing hydrophobic residues in loop 1, the M3 S202W I203F mutant became more rigid and halotolerant due to the formation of additional interdomain hydrophobic interactions between the introduced Y22 in loop 1 and W204 in α7. These results indicate that increasing intraprotein hydrophobic interactions is also a way to improve the halotolerance of enzymes with industrial potential under high-salt conditions.IMPORTANCE Esterases and lipases for industrial application are often subjected to harsh conditions such as high salt concentrations, low water activity, and the presence of organic solvents. However, reports on halotolerant esterases and lipases are limited, and the underlying mechanism for their halotolerance is still unclear due to the lack of structures. In this study, we focused on the improvement of the halotolerance of a salt-sensitive esterase, E40, and the underlying mechanism. The halotolerance of E40 was significantly improved by introducing hydrophobic residues. Comparative structural analysis of E40 and its halotolerant mutants revealed that increased intraprotein hydrophobic interactions make these mutants more rigid and more stable than the wild type against high concentrations of salts. This study shows a new way to improve enzyme halotolerance, which is helpful for protein engineering of salt-sensitive enzymes.

Erythrobacter xanthus sp. nov., isolated from surface seawater of the South China Sea.

A Gram-reaction-negative, aerobic, oxidase- and catalase-positive, yellow-pigmented, non-flagellated, rod-shaped bacterium, designed strain SM1501T, was isolated from surface seawater of the South China Sea. SM1501T grew at 7-42 °C and with 0-11 % (w/v) NaCl. Phylogenetic analyses based on 16S rRNA gene sequences revealed that SM1501T represented a member of the genus Erythrobacter, sharing the highest 16S rRNA gene sequence similarity (97.4 %) with Erythrobacter luteus and 94.2-96.5 % 16S rRNA gene sequence similarities to other species of the genus Erythrobacterwith validly published names. The average nucleotide identity (ANI) value and in silico DNA-DNA hybridization value between SM1501T and E. luteus were only 74.6 and 20.0 %, respectively. The predominant cellular fatty acids of SM1501T were C17 : 1ω6c, C18 : 1ω7c and summed feature 3 (C16 : 1ω7c and/or iso-C15 : 0 2-OH). The major polar lipids of the strain were phosphatidylethanolamine, phosphatidylglycerol, sphingoglycolipid, diphosphatidylglycerol and phosphatidylcholine and the main respiratory quinone of was Q-10. Polyphasic data presented in this paper support the notion that SM1501T represents a novel species in the genus Erythrobacter, for which the name Erythrobacter xanthus sp. nov. is proposed. The type strain of Erythrobacterxanthus is SM1501T (=KCTC 42669T=CCTCC AB 2015396T).