A preoperative risk model for early recurrence after radical resection may facilitate initial treatment decisions concerning the use of neoadjuvant therapy for patients with pancreatic ductal adenocarcinoma.

BACKGROUND: Neoadjuvant chemotherapy may benefit patients with pancreatic ductal adenocarcinoma with resectable and borderline disease. Inappropriate use of neoadjuvant therapy, however, may lead to the loss of therapeutic opportunities. Until an effective prediction model of individual drug sensitivity is established, no accurate model exists to help surgeons decide on the appropriate use of neoadjuvant chemotherapy. We hypothesized that early recurrence in patients undergoing upfront, early resection may be an indication for neoadjuvant chemotherapy. Therefore, we aimed to use preoperative clinical parameters to establish a model of early recurrence to select patients at high risk for neoadjuvant chemotherapy. METHODS: Patients who underwent resection for pancreatic ductal adenocarcinoma between January 2014 and November 2017 were analyzed retrospectively. After the minimum P-value approach, the patients were divided into three groups: early recurrence, middle recurrence, and late/non-recurrence. Preoperative clinicopathologic factors that could predict early recurrence were included in a Cox proportional hazards regression model for univariate and multivariate analyses. The factors related to early recurrence were included to establish nomogram and decision tree models, which were then validated in 68 patients. RESULTS: We found that 235 (72.5%) of 324 patients had recurrence with a median recurrence-free survival of 210 days. The early recurrence, middle recurrence, and late/non-recurrence groups differed in preoperative carbohydrate antigen 19-9 and carcinoembryonic antigen levels, "resectability" on cross-sectional imaging, resection requiring a vascular resection, T stage, tumor size, and adjuvant chemotherapy. The best cutoff value of early recurrence was the first 162 days postoperatively. Univariate and multivariate analyses showed that selected preoperative chief complaints, lymph node enlargement and resectability on cross-sectional imaging, preoperative carbohydrate antigen 19-9 levels >210 kU/L, and a neutrophil/lymphocyte ratio >4.2 were independent predictors for early recurrence. CONCLUSION: We have successfully built a prediction model of early recurrence of patients with pancreatic ductal adenocarcinoma with the optimal cutoff early-recurrence value of 162 days. Our nomogram and decision tree models may be used to select those at high risk for early recurrence to guide preoperative decision-making concerning the use of neoadjuvant therapy in those patients who have "resectable" disease and not only the more classic criteria of borderline resectability.

Vancomycin intermediate-resistant Staphylococcus aureus (VISA) isolated from a patient who never received vancomycin treatment.

BACKGROUND: With the abuse of antibiotics, the methicillin-resistant Staphylococcus aureus (MRSA) strain became prevalent. Furthermore, Staphylococcus aureus with a character of vancomycin intermediate-resistance (VISA) has been found globally since the first report in Japan. The main objectives of this study were to report a case of VISA isolated from a Chinese patient who had never undergone Vancomycin treatment, and to determine its molecular character. METHODS: A total of 9 strains were recovered from a patient during the therapeutic process. Antimicrobial susceptibility testing was performed to determine their antibiotic susceptibility patterns. To detect the VISA strain's molecular epidemiological features, growth and morphological characters, we used multilocus sequence typing, autolysis assay and transmission electric microscope tests. Pulsed-field gel electrophoresis (PFGE) was performed to characterize the heterogeneities of all isolates. RESULTS: One isolate was found to exhibit vancomycin intermediated-resistant with MIC of 8 µg/ml. It was ST239-T030-agr-1, had thickened cell wall, and displayed a slower growth rate and reduced susceptibility to Triton X-100-induced autolysis than other strains. All 9 strains exhibited the same PFGE pattern. CONCLUSION: This is the first report of VISA found in central China from a patient who had never received vancomycin treatment.

Construction of mTNFR1shRNA plasmid and its biological effects on MHV-3 induced fulminant hepatitis in BALB/cJ mice.

Previous study on TNFR1-mediated hepatocyte apoptosis has been implicated in the development of fulminant viral hepatitis. To interfere with the potentially effective target, plasmid named p-mTNFR1shRNA complimentary to the sequence responsible for mTNFR1 was also constructed and further confirmed by sequence analysis. To investigate the effect of mTNFR1shRNA plasmid on mTNFR1 expression in vivo and the disease progress in MHV-3 induced fulminant hepatitis mice model. By hydrodynamic injection of mTNFR1shRNA plasmid, the survival rate of mice, hepatic pathological change were examined and compared between mice with/without mTNFR1shRNA plasmid intervention. The expression of mTNFR1 was detected by Real-time PCR, immunohistochemistry staining. The mTNFR1shRNA plasmid significantly reduced mTNFR1 expression in vivo, markedly ameliorates inflammatory infiltration, prolonged the survival time period and elevated the survival rate from 0 up to 13.3% in Balb/cJ mice with MHV-3 induced fulminant hepatitis. This study was designed to explore the opportunity of RNA interference technique in inhibiting TNFR1 expression, which has been reported to be involved in the development of a variety of diseases including fulminant viral hepatitis and severe chronic hepatitis B.

The FGL2/fibroleukin prothrombinase is involved in alveolar macrophage activation in COPD through the MAPK pathway.

Fibrinogen-like protein 2 (FGL2)/fibroleukin has been reported to play a vital role in the pathogenesis of some critical inflammatory diseases by possessing immunomodulatory activity through the mediation of "immune coagulation" and the regulation of maturation and proliferation of immune cells. We observed upregulated FGL2 expression in alveolar macrophages from peripheral lungs of chronic obstructive pulmonary disease (COPD) patients and found a correlation between FGL2 expression and increased macrophage activation markers (CD11b and CD14). The role of FGL2 in the activation of macrophages was confirmed by the detection of significantly decreased macrophage activation marker (CD11b, CD11c, and CD71) expression as well as the inhibition of cell migration and inflammatory cytokine (IL-8 and MMP-9) production in an LPS-induced FGL2 knockdown human monocytic leukemia cell line (THP-1). Increased FGL2 expression co-localized with upregulated phosphorylated p38 mitogen-activated protein kinase (p38-MAPK) in the lung tissues from COPD patients. Moreover, FGL2 knockdown in THP-1 cells significantly downregulated LPS-induced phosphorylation of p38-MAPK while upregulating phosphorylation of c-Jun N-terminal kinase (JNK). Thus, we demonstrate that FGL2 plays an important role in macrophage activation in the lungs of COPD patients through MAPK pathway modulation.

Dual interference with novel genes mfgl2 and mTNFR1 ameliorates murine hepatitis virus type 3-induced fulminant hepatitis in BALB/cJ mice.

Our studies and those of many others have implicated hepatocyte necrosis and apoptosis mediated by fibrinogen-like protein-2 (fgl2) prothrombinase and tumor necrosis factor receptor (TNFR) in the development of fulminant viral hepatitis, a disease with a mortality rate greater than 80% in cases lacking immediate organ transplantation. This study was designed to explore the efficacy of dual short hairpin RNA (shRNA) interference with fgl2 and TNFR1 in the treatment of murine hepatitis virus strain 3 (MHV-3)-induced fulminant hepatitis in mice. Plasmids p-mfgl2shRNA and p-mTNFR1shRNA, complementary to the sequences for mfgl2 and mTNFR1, were constructed. Plasmids pEGFP-mfgl2 and pEGFP-mTNFR1 expressing mfgl2-EGFP (enhanced green fluorescent protein) and mTNFR1-EGFP fusion proteins were also constructed to screen the inhibitory effect of p-mfgl2shRNA and p-mTNFR1shRNA on mfgl2 and mTNFR1 expression. Cotransfection of individual shRNA plasmids and pcDNA3.0-mfgl2 and pcDNA3.0-mTNFR1 expression constructs into Chinese hamster ovary (CHO) cells significantly inhibited mfgl2 and mTNFR1 gene expression, as evidenced by fluorescence microscopy, reverse transcription-polymerase chain reaction, and Western blotting. In vivo hydrodynamic delivery of dual-interference shRNA plasmids for mfgl2 and mTNFR1 significantly decreased mfgl2 and mTNFR1 expression; markedly ameliorated fibrin deposition, hepatocyte necrosis, and apoptosis; and prolonged survival against fulminant viral hepatitis induced by MHV-3 in BALB/cJ mice compared with mfgl2 or TNFR1 single-gene interference. These results indicate that in vivo interference with genes for more than one key target provides superior treatment efficacy compared with single-gene interference.

[Construction of p-hfgl2shRNA and its effect on hfgl2 expression in vitro].

OBJECTIVE: Our previous studies have shown that an anti-sense plasmid to mouse fibrinogen like protein 2 (mfgl2) significantly reduced mfgl2 expression in vivo, markedly ameliorated inflammatory infiltration, fibrin deposition and hepatocyte necrosis, prolonged the survival time period and elevated the survival rate in Balb/cJ mice with murine hepatitis virus type 3 (MHV-3) induced fulminant hepatitis. This study was designed to explore the opportunity of RNA interference technique in the inhibitory application of hfgl2 expression, which has been reported to be involved in a variety of disease developments including fulminant viral hepatitis, acute rejection of allo/xeno transplantation and fetal loss syndrome. METHODS: A plasmid named p-hfgl2shRNA complimentary to the sequence responsible for hfgl2 was constructed; meanwhile irrelevant shRNA plasmid with a random combination of the p-hfgl2shRNA sequence was used as a control. A plasmid named pEGFP-hfgl2 expressing hfgl2-EGFP fusion protein was also constructed for the screening of the effect of p-hfgl2shRNA on the hfgl2 expression. By cotransfection of the constructed p-hfgl2shRNA and pEGFP-hfgl2 or pcDNA3.1-hfgl2 expression plasmids into CHO cells, the inhibition of hfgl2 expression by hfgl2shRNA was analyzed by direct observation through fluorescent microscopy, FACS, real time PCR and immunohistochemistry staining. RESULTS: The experiments showed a significant inhibitory effect of p-hfgl2shRNA on hfgl2 expression at 48 h post-cotransfection by observation of green fluorescent cells, FACS, real time PCR and immunohistochemistry staining, with the inhibitory efficiency reaching as high as 85.5%. CONCLUSION: The study demonstrated that p-hfgl2shRNA successfully interfered with hfgl2 expression in vitro. This provides a foundation to further explore its application in vivo.

Hepatitis B virus-induced hFGL2 transcription is dependent on c-Ets-2 and MAPK signal pathway.

Fibrinogen-like protein 2 (FGL2)/fibroleukin plays a pivotal role in the pathogenesis of experimental and human fulminant and chronic viral hepatitis. To define the transcription factor(s) and upstream signal transduction pathways involved in the transcription of human FGL2 (hFGL2) in response to hepatitis B (HB) virus, hepatitis B core (HBc), hepatitis B virus S protein (HBs), or hepatitis B virus X protein (HBx) protein, expression plasmids were cotransfected with an hFGL2 promoter luciferase reporter construct into Chinese hamster ovary and HepG2 cells, respectively. HBc and HBx proteins, but not HBs protein, enhanced hFGL2 transcription in both cell lines. A strong regulatory region from -712 to -568 (relative to the transcriptional starting site) was shown to be responsible for hFGL2 gene transcription in response to both HBc and HBx proteins. c-Ets-2 was shown to be translocated to the nucleus in association with hFGL2 expression in response to both HBc and HBx proteins. Short hairpin RNA (shRNA) interference of c-Ets-2 expression inhibited hFGL2 gene transcription by 64.8 and 60.0% in response to HBc and HBx, respectively. c-Ets-2 protein was highly expressed in peripheral blood mononuclear cells from patients with severe chronic hepatitis B (CHB) in contrast to patients with mild CHB. Increased phosphorylation of ERK and JNK was detected in peripheral blood mononuclear cells from patients with severe CHB. ERK inhibitor PD098059 or ERK shRNA abolished the nuclear c-Ets-2 DNA binding activity and hFGL2 induction in response to HBc, whereas JNK inhibitor SP600125 or JNK shRNA abolished the nuclear c-Ets-2 DNA binding activity and hFGL2 induction in response to HBx. In conclusion, HBc and HBx proteins enhance transcription of hFGL2 through c-Ets-2 dependent on MAPK signal pathways.