Impact of physician-staffed ground emergency medical services-administered pre-hospital trauma care on in-hospital survival outcomes in Japan.

PURPOSE: In Japan, the vehicle used in pre-hospital trauma care systems with physician-staffed ground emergency medical services (GEMS) is referred to as a "doctor car". Doctor cars are highly mobile physician-staffed GEMS that can provide complex pre-hospital trauma management using various treatment strategies. The number of doctor car operations for patients with severe trauma has increased. Considering facility factors, the association between doctor cars and patient outcomes remains unclear. Therefore, this study aimed to examine the relationship between doctor cars for patients with severe trauma and survival outcomes in Japan. METHODS: A nationwide retrospective cohort study was conducted to compare the impact of the doctor car group with the non-physician-staffed GEMS group on in-hospital survival in adult patients with severe trauma. The data were analyzed using multivariable logistic regression models with generalized estimating equations. RESULTS: This study included 372,365 patients registered in the Japan Trauma Data Bank between April 2009 and March 2019. Of the 49,144 eligible patients, 2361 and 46,783 were classified into the doctor car and non-physician staffed GEMS groups, respectively. The adjusted odds ratio (OR) for survival was significantly higher in the doctor car group than in the non-physician staffed GEMS group (adjusted OR = 1.228 [95% confidence interval 1.065-1.415]). CONCLUSION: Using nationwide data, this novel study suggests that doctor cars improve the in-hospital survival rate of patients with severe trauma in Japan. Therefore, doctor cars could be an option for trauma strategies.

Potential biomarker enhancing the activity of tuberculosis, hsa-miR-346.

OBJECTIVES: To determine the usefulness of hsa-miR-346, a potential biomarker enhancing the activity of non-tuberculous mycobacterial diseases, as a biomarker of tuberculosis activity. METHODS: We investigated whether hsa-miR-346 is secreted by human macrophages infected with Mycobacterium tuberculosis (M. tuberculosis) in an in vitro study. In addition, a cross-sectional study was conducted first to evaluate whether serum hsa-miR-346 is elevated in patients with tuberculosis compared with that in healthy individuals. Second, we conducted a retrospective study to evaluate whether anti-tuberculosis treatment reduces serum hsa-miR-346 levels. RESULTS: Log hsa-miR-346 levels were significantly elevated in the supernatant of human macrophages infected with M. tuberculosis in a dose-dependent manner. The mean serum log hsa-miR-346 levels were -15.48 (-15.76 to -15.21) in patients with tuberculosis and -16.12 (-16.29 to -15.95) in healthy volunteers, which significantly differed. In addition, hsa-miR-346 significantly decreased at 2 months from starting an anti-tuberculosis treatment. CONCLUSIONS: We consider hsa-miR-346 as a potential biomarker enhancing the tuberculosis activity.

Prolonged viral shedding of SARS-CoV-2 in an immunocompromised patient.

The duration of viral shedding of SARS-CoV-2 is usually less than 10 days. We experienced a COVID-19 case with prolonged viral shedding for 2 months. His cell mediated immunity has been depressed (CD4+T cell <100/µl) due to advanced malignant lymphoma and chemotherapy which had been completed 4 months prior to the onset of symptoms of COVID-19. We administered several treatments against COVID-19, however the results of Polymerase Chain Reaction (PCR) from nasopharyngeal specimens remained positive to SARS-CoV-2 for 2 months. Moreover, virus isolation assays performed on Day 59 also remained positive. He was finally discharged on Day 69 with two consecutive negative PCR results for SARS-CoV-2. Immunocompromised status may prolong viral shedding and it is therefore important for the clinician to take into account this when assessing such patients.

Advanced glycation end-products enhance calcification in cultured rat dental pulp cells.

INTRODUCTION: Amorphous calcification frequently appears in dental pulp tissues of diabetic patients; however, its pathologic process has not been fully elucidated. We previously found that pulp stones and thickened predentin occurred more frequently in diabetic rats. Recent findings demonstrated that accumulation of advanced glycation end-products (AGE) might be involved in vascular calcification complicated with diabetes. The aim of this study was to determine the effect of AGE on calcified nodule formation by rat dental pulp cells in culture. METHODS: Rat dental pulp cells and gingival fibroblasts were independently cultured with 50 and 100 µg/mL AGE. Alkaline phosphatase activity and calcified nodule formation were measured. Expressions of receptor for AGE, osteopontin (OPN), and osteocalcin (OCN) mRNA were determined by quantitative real-time polymerase chain reaction. Protein levels of OPN and OCN secreted in culture medium were quantified by enzyme-linked immunosorbent assay. RESULTS: AGE (50 and 100 µg/mL) markedly increased both alkaline phosphatase activity and calcified nodule formation in dental pulp cells (P < .01), whereas it did not affect those in gingival fibroblasts. Real-time polymerase chain reaction analysis revealed that AGE increased mRNA expressions of receptor for AGE, OPN, and OCN in dental pulp cells (P < .05). Enzyme-linked immunosorbent assay analysis revealed that the protein levels of OPN and OCN produced by dental pulp cells were higher in AGE-treated than in untreated cells (P < .05). CONCLUSIONS: AGE enhanced the calcification potentials of rat dental pulp cells, suggesting that it may stimulate pathologic calcification of diabetic dental pulp tissues.

CYP2C8 haplotype structures and their influence on pharmacokinetics of paclitaxel in a Japanese population.

OBJECTIVE: CYP2C8 is known to metabolize various drugs including an anticancer drug paclitaxel. Although large interindividual differences in CYP2C8 enzymatic activity and several nonsynonymous variations were reported, neither haplotype structures nor their associations with pharmacokinetic parameters of paclitaxel were reported. METHODS: Haplotype structures of the CYP2C8 gene were inferred by an expectation-maximization based program using 40 genetic variations detected in 437 Japanese patients, which included cancer patients. Associations of the haplotypes and paclitaxel pharmacokinetic parameters were analyzed for 199 paclitaxel-administered cancer patients. RESULTS: Relatively strong linkage disequilibriums were observed throughout the CYP2C8 gene. We estimated 40 haplotypes without an amino-acid change and nine haplotypes with amino acid changes. The 40 haplotypes were classified into six groups based on network analysis. The patients with heterozygous *IG group haplotypes harboring several intronic variations showed a 2.5-fold higher median area under concentration-time curve of C3'-p-hydroxy-paclitaxel and a 1.6-fold higher median value of C3'-p-hydroxy-paclitaxel/paclitaxel area under concentration-time curve ratio than patients bearing no *IG group haplotypes (P<0.001 for both comparisons by Mann-Whitney U-test). No statistically significant differences, however, were observed between patients with and without the *IG group (haplotypes) in clearance and area under concentration-time curve of paclitaxel, area under concentration-time curve of 6alpha-hydroxy-paclitaxel and 6alpha-, C3'-p-dihydroxy-paclitaxel, and area under concentration-time curve ratio of 6alpha-hydroxy-paclitaxel/paclitaxel. CONCLUSION: CYP2C8*IG group haplotypes were associated with increased area under concentration-time curve of C3'-p-hydroxy-paclitaxel and area under concentration-time curve ratio of C3'-p-hydroxy-paclitaxel/paclitaxel. Thus, *IG group haplotypes might be associated with reduced CYP2C8 activity, possibly through its reduced protein levels.

Impact of the haplotype CYP3A4*16B harboring the Thr185Ser substitution on paclitaxel metabolism in Japanese patients with cancer.

OBJECTIVE: Paclitaxel is one of the most important anticancer drugs for the treatment of various tumors such as non-small cell lung cancer. We investigated the association between CYP3A4 haplotypes and pharmacokinetic parameters of paclitaxel metabolism. METHODS: This study enrolled 235 Japanese patients with cancer who were receiving paclitaxel. These patients were screened for CYP3A4 gene polymorphisms by either direct sequencing or pyrosequencing. Plasma concentrations of paclitaxel and its 3 metabolites were determined by HPLC in 229 patients. RESULTS: Median values of paclitaxel clearance, normalized for body surface area, were lower in the high-dose group (>or=175 mg/m2, n = 199) than in the low-dose group (

Polymorphic tandem repeat sequences of the thymidylate synthase gene correlates with cellular-based sensitivity to fluoropyrimidine antitumor agents.

PURPOSE: Thymidylate synthase (TS) is one of the target molecules for the antitumor effects of fluoropyrimidine drugs. The cellular thymidylate synthase level is one of the determining factors for the antitumor activity of fluoropyrimidines. TYMS, which encodes TS, has been reported to possess 28-bp tandem repeat sequences in its 5'-untranslated region, the number of which varies. In addition, single nucleotide polymorphisms have also been shown in a triple repeat sequence. In this study, correlation between the polymorphic tandem repeat sequences of the TYMS gene and the antitumor activities of 5-fluorouracil (5-FU) and 5-fluoro-2'-deoxyuridine (FUdR) were investigated with 30 established human cell lines derived from solid tumors. METHODS: A reporter assay system was developed in order to compare the ability of the transactivation mediated by the double (2R) and triple (c- or g-type, 3Rc or 3Rg, respectively) repeat sequences using a human colon cancer cell line, DLD-1. The 50% inhibitory concentration (IC(50)) of cell growth by 5-FU and FUdR was measured with 30 different established cell lines of human solid tumors. Genotypes based on the number of the 28-bp TYMS tandem repeat for the above cell lines were determined by electrophoretical analysis of PCR products containing the repeat sequences and nucleotide sequencing. RESULTS: The reporter activity mediated by the 3Rg sequence was significantly higher than that by the 2R and 3Rc sequences. Activities mediated by the 2R and 3Rc sequences were comparable. According to the reporter assay, 2R and 3Rc were judged as low TS expression alleles and 3Rg as a high TS expression allele. On the basis of IC(50) values, cells possessing the 2R/2R and 2R/3R repeat of TYMS were significantly more sensitive to FUdR than those with the 3R/3R repeat. Cells possessing 3Rg/3Rg (a high TS expression genotype) were significantly less sensitive to FUdR than cells with 2R/2R, 2R/3Rc, and 3Rc/3Rc (low TS expression genotypes). CONCLUSIONS: Our results of the reporter assays using 2R, 3Rc, and 3Rg repeat sequences prompted us to classify 3Rg as a high TS expression allele, and 2R and 3Rc as low TS expression alleles. The cells with low TS expression alleles were shown to exhibit significantly higher FUdR sensitivity than the cells with high TS expression alleles for the first time. These results were consistent with numerous previous in vitro and in vivo findings that tumors showing high TS expression were less sensitive to fluoropyrimidines. These results support the idea that genotyping the tandem repeat sequences of TYMS in the 5'-untranslated region is useful for individualized therapy involving fluoropyrimidine antitumor drugs.

Cytosolic and microsomal activation of doxifluridine and tegafur to produce 5-fluorouracil in human liver.

PURPOSE: The enzymatic formation of 5-fluorouracil (5-FU) from two fluoropyrimidine prodrugs, doxifluridine (5'-DFUR) and tegafur (FT), was compared in vitro in order to determine whether there are differences between the metabolic profiles of the two prodrugs. METHODS: Conversion of the two fluoropyrimidine prodrugs to 5-FU was measured by high-performance liquid chromatography at a concentration of 500 micro M using the microsomal and cytosolic fractions of 12 human livers. The degree of correlation between the 5-FU-forming activities was determined using various cytochrome P450-dependent reactions. RESULTS: Liver microsomes catalyzed 5-FU formation from 5'-DFUR at rates of 10.0-160.1 pmol/min per mg protein and correlated well with CYP2A6-dependent coumarin 7-hydroxylase activity. The rates of microsomal 5-FU formation from FT ranged from 44.9 to 808.3 pmol/min per mg protein and also correlated with coumarin 7-hydroxylase activity. The cytosol fractions catalyzed 5-FU formation from 5'-DFUR at rates of 3,164.6 to 6,026.6 pmol/min per mg protein, almost two orders of magnitude higher than the rates of cytosolic 5-FU formation from FT (46.8-219.0 pmol/min per mg protein). CONCLUSIONS: The cytosolic enzymes in livers appear to be important for 5-FU formation from 5'-DFUR. Both cytosolic and microsomal enzymes were involved almost equally in 5-FU formation from FT. The increased formation of 5-FU from 5'-DFUR might provide an answer to the question of why similar blood 5-FU levels were retained despite blood 5'-DFUR levels lower than blood FT levels.