A study on expression of GRP78 and CHOP in neutrophil endoplasmic reticulum and their relationship with neutrophil apoptosis in the development of sepsis.

We investigated the relationship between neutrophil apoptosis and endoplasmic reticulum stress (ERS) in sepsis and its mechanism. A prospective cohort study was conducted by recruiting a total of 58 patients with sepsis. Peripheral blood samples were collected on 1, 3, 5 and 7 days after admission to the ICU. The expressions of endoplasmic reticulum specific glucose regulatory protein 78 (GRP78), C/EBP homologous protein (CHOP), apoptosis signal-regulating kinase 1 (ASK1), Bcl-2-like 11 (BIM), death receptor 5 (DR5), c-Jun N-terminal kinases (JNK) and p38 were detected by Western blot and PCR. The subcellular location of CHOP and GRP78 was observed by immunofluorescence analysis. Spearman correlation was used to analyze the correlation between the expression of chop protein and the apoptosis rate of peripheral blood neutrophils. Healthy volunteers in the same period were selected as the healthy control group. The expression of GRP78 protein was significantly elevated on the first day of ICU admission and showed a decreasing trend on the third, fifth and seventh day, but was significantly higher than the corresponding healthy control group. The expression of CHOP protein reached the highest level on the third day. The expression of chop protein in each group was significantly higher than that in the corresponding healthy control group. Immunofluorescence staining clearly showed that the CHOP protein accumulated in the nucleus, with an elevation in the intensity of GRP78. The neutrophil apoptosis rate of sepsis patients on the 1st, 3rd, 5th and 7th day of ICU stay was significantly higher than that of the healthy control group, with the highest apoptosis rate on the 3rd day, and then decreased gradually. CHOP protein expression level was significantly positively correlated with neutrophil apoptosis rate in sepsis patients. Endoplasmic reticulum stress occurs in neutrophils during the development of sepsis. GRP78 protein and CHOP protein may be involved in the pathological process of neutrophil apoptosis in sepsis.

Regional Character, Restaurant Size, and Food Safety Risk: Evidence from Food Safety Violation Data in Gansu Province, China.

ABSTRACT: Restaurants are a place where food is prepared and cooked directly for customers. Food safety in restaurants is a public health concern and a multidisciplinary issue that needs to be explored. To protect the health of consumers and identify external factors that may affect food safety risk, this study explores how economic development and population density at the local level relate to food safety inspection outcomes in restaurants of different sizes. Using food safety violation data from 2017 and 2018, we categorized restaurants in Gansu Province, China, into small and large ventures to examine the relationships among regional character, restaurant size, and food safety risk. Data were analyzed using Mann-Whitney U tests and negative binomial regression models. Our results show that large restaurants have a higher food safety risk than small restaurants. Moreover, the region with the lowest level of economic development had the highest food safety risk, while the region with the lowest population density had insufficient local inspections. By providing insight into which establishments demonstrate the highest food safety risks, our findings contribute to the development of processes that seek to effectively identifying food safety risks.

ERG rearrangement as a novel marker for predicting the extra-prostatic extension of clinically localised prostate cancer.

Currently, there are no well-established preoperative clinicopathological parameters for predicting extra-prostatic extension (EPE) in patients with clinically localised prostate cancer (PCa). The transmembrane protease serine 2 (TMPRSS2)-ETS-related gene (ERG) fusion gene is a specific biomarker of PCa and is considered a prognostic predictor. The aim of the present study was to assess the value of this marker for predicting EPE in patients with clinically localised PCa. In total, 306 PCa patients with clinically localised disease, including 220 patients (71.9%) with organ-confined disease and 86 EPE cases (28.1%), were included in the study. Receiver operating characteristic curves and logistic regression were employed to establish the optimal cut-off value and to investigate whether ERG rearrangement was an independent predictor for the EPE of clinically localised PCa. A leave-one-out cross-validation (LOOCV) model was implemented to validate the predictive power of ERG rearrangement. An increase in ERG rearrangements was identified to be associate'd with EPE, and the optimal cut-off for predicting EPE was determined to be 2.25%, with a sensitivity of 70.24% [95% confidence interval (CI), 62.6-78.9%], a specificity of 80.43% (95% CI, 75.4-85.1%), and an area under the curve (AUC) of 0.781 (95% CI, 0.730-0.826). In the LOOCV model, ERG rearrangement also demonstrated good performance for predicting EPE (sensitivity, 76.923%; specificity, 71.429%; 95% CI for AUC, 0.724-0.958). In addition, a high Gleason score (≥7) and a cT2c classification upon biopsy were independent factors for EPE.

Overexpression of ribosomal L1 domain containing 1 is associated with an aggressive phenotype and a poor prognosis in patients with prostate cancer.

The aim of the present study was to investigate the overexpression and significance of ribosomal L1 domain containing 1 (RSL1D1) in prostate cancer (PCA). The present study performed immunohistochemical analysis on the tissues of 138 patients with pathologically confirmed PCA. The patients were followed up for a median of 87 months. In addition, 50 patients with benign prostatic hyperplasia (BPH) were enrolled in the present study as a control group. Of the 138 PCA tissue samples, 124 (89.9%) expressed RSL1D1, while 4 out of the 50 (8.0%) BPH tissues expressed RSL1D1. The present study defined a high RSL1D1 expression level as the relative gene expression that was equal to or higher than the median, and low expression as the gene expression lower than the median. The pathological stage of patients with PCA (≥pT3a vs. pT2c) and the Gleason scores of patients (≥7 vs. <7) were associated with RSL1D1 expression (χ2=4.809 and 14.703; P=0.028 and P<0.0001, respectively) and a high expression of RSL1D1 (χ2=10.294 and 17.520; P=0.001 and P<0.0001, respectively). Kaplan-Meier curve analysis demonstrated that the biochemical recurrence (BCR)-free survival rate of the patients was increased in patients without RSL1D1 expression (P=0.0046), in those with low RSL1D1 expression (P<0.0001) and in those without RSL1D1 expression in the mesenchyme (P=0.006) compared with those patients with no expression, low expression and no mesenchymal expression, respectively. A high expression level of RSL1D1 was demonstrated to be an independent prognostic factor of BCR in patients with PCA using Cox regression analysis. Overall, the present study demonstrated that RSL1D1 expression was associated with PCA, and that it may aid in the improvement of diagnosis, prognosis and risk stratification of patients with PCA.

Overexpression of cofilin 1 in prostate cancer and the corresponding clinical implications.

Cofilin 1 (CFL1) is a cytoskeletal protein and overexpression of the protein has been associated with aggressiveness in certain types of malignancies. The aim of the present study was to investigate the clinical implications of CFL1 expression in prostate cancer (PCa). Immunohistochemical analysis was performed using formalin-fixed paraffin-embedded tissue sections obtained from 111 patients with PCa and 47 patients with benign prostatic hyperplasia (BPH). In total, 78 (70.3%) out of 111 PCa tissues were found to express the CFL1 protein, while no expression was detected in BPH tissues. In addition, CFL1 was also observed to be significantly associated with the Gleason score (GS; <7 vs. ≥7; P<0.0001) and presence of lymph node metastasis (presence vs. absence; P<0.0001). However, there was no association between the expression of CFL1 and other clinicopathological variables, such as age (<69 years vs. ≥69 years; P=0.54), pre-operative prostate specific antigen level (<20 ng/ml vs. ≥20 ng/ml; P=0.45) and pathological stage (T2 vs. ≥T3a; P=0.055). In addition, 35 tissues (31.5%) were observed to possess a CFL1-positive mesenchyme. CFL1 expression was revealed to be an independent predictive factor for a high GS. The status of CFL1 expression in the mesenchyme also found to individually predict extraprostatic extension in PCa patients, based on multivariate analysis. The results of the present study indicated that CFL1 may specifically predict the development of PCa, and that the expression of CFL1 in the mesenchyme may be closely associated with the development of lymph node metastasis.

Pharmacokinetics of rh-IFNalpha-2a-NGR, a tumor targeted-therapy candidate, following intramuscular administration to mice, rats and monkeys.

The compound rh-IFNalpha-2a-NGR can inhibit tumor angiogenesis and could be used for targeted therapy. In the present study, double antibody sandwich ELISA analysis was used to determine the concentration of rh-IFNalpha-2a-NGR in serum after intramuscular administration of various dosages to mice, rats and monkeys. The results showed that the pharmacokinetic properties of rh-IFNalpha2a-NGR after i.m. administration to mice, rats and monkeys were consistent with a one-compartment open model. The main pharmacokinetic parameters in mice (9.36 microg/kg), rats (4.68 microg/kg) and monkeys (2.34 microg/kg) after i.m. rh-IFNalpha2a-NGR were as follows: T(peak) was 0.49, 1.65 and 3.60h, C(max) was 3030.20, 654.49 and 268.13 ng/L, t1/2 was 0.39, 4.52 and 2.70 h, and AUC(0-infinity)) was 4197.65, 5784.58 and 2622.06 ng/L x h, respectively. Also, mice, rats and monkeys had their own distinct metabolic characteristics. These data would provide references for further clinical pharmacokinetic study of rh-IFNalpha2a-NGR.

Examination of the role of DNA polymerase proofreading in the mutator effect of miscoding tRNAs.

We previously described Escherichia coli mutator tRNAs that insert glycine in place of aspartic acid and postulated that the elevated mutation rate results from generating a mutator polymerase. We suggested that the proofreading subunit of polymerase III, epsilon, is a likely target for the aspartic acid-to-glycine change that leads to a lowered fidelity of replication, since the altered epsilon subunits resulting from this substitution (approximately 1% of the time) are sufficient to create a mutator effect, based on several observations of mutD alleles. In the present work, we extended the study of specific mutD alleles and constructed 16 altered mutD genes by replacing each aspartic acid codon, in series, with a glycine codon in the dnaQ gene that encodes epsilon. We show that three of these genes confer a strong mutator effect. We have also looked for new mutator tRNAs and have found one: a glycine tRNA that inserts glycine at histidine codons. We then replaced each of the seven histidine codons in the mutD gene with glycine codons and found that in two cases, a strong mutator phenotype results. These findings are consistent with the epsilon subunit playing a major role in the mutator effect of misreading tRNAs.

Isolation and characterization of human tissue kallikrein produced in Escherichia coli: biochemical comparison to the enzymatically inactive prokallikrein and methionyl kallikrein.

This report describes bacterial expression, isolation, and characterization of human tissue kallikrein recombinantly produced in Escherichia coli. Successful production of enzymatically active recombinant human kallikrein requires the following processes: expression, solubilization and refolding of prokallikrein, thermolysin activation, and chromatographic separation. All experimental data confirmed that bacterially derived human kallikrein is properly folded and exhibits expected biochemical functions. As confirmed by SDS-PAGE and reverse-phase HPLC, recombinant kallikrein is apparently pure and is devoid of reduced or other partially folded kallikrein forms. Recombinant kallikrein behaves as a monomeric molecule in solution and exhibits full enzymatic activity in hydrolyzing peptide substrates. The molecule can bind to aprotinin to form kallikrein-inhibitor complex at a 1:1 molar ratio. Peptide mapping analysis derived from pepsin digestion of recombinant kallikrein assigned five disulfide bonds which match those of porcine kallikrein predicted from X-ray structure. Peptides containing unpaired cysteines or mispaired disulfide bonds were not detected. Both properly folded prokallikrein and methionyl kallikrein, containing a propeptide and an initiator methionine at their N-termini, respectively, were also produced and isolated. These two molecules are structurally similar to recombinant kallikrein, but are not enzymatically active.

Activation of cell-specific expression of rat growth hormone and prolactin genes by a common transcription factor.

In the anterior pituitary gland, there are five phenotypically distinct cell types, including cells that produce either prolactin (lactotrophs) or growth hormone (somatotrophs). Multiple, related cis-active elements that exhibit synergistic interactions appear to be the critical determinants of the transcriptional activation of the rat prolactin and growth hormone genes. A common positive tissue-specific transcription factor, referred to as Pit-1, appears to bind to all the cell-specific elements in each gene and to be required for the activation of both the prolactin and growth hormone genes. The data suggest that, in the course of development, a single tissue-specific factor activates sets of genes that ultimately exhibit restricted cell-specific expression and define cellular phenotype.