Hinokitiol impedes tumor drug resistance by suppressing protein kinase B/mammalian targets of rapamycin axis.

Chemotherapy is a treatment method commonly used for cancer and that patients showing low to no response to the treatment often developed drug resistance via multiple mechanisms. Natural products have been shown to reduce tumor drug resistance. Hinokitiol, a natural tropolone derivative, has potential as an antitumor agent. To improve the efficacy and safety of hinokitiol, a further understanding of hinokitiol interactions with the tumor microenvironment is necessary. The presence of plasma membrane multidrug resistance protein P-glycoprotein (P-gp) is favorable for tumor cells to elicit chemotherapeutic resistance. Here, we showed that hinokitiol dose-dependently decreased P-gp expression and suppressed the P-gp-driven efflux activity based on Rhodamine 123 assay. The protein expression levels of phosph-protein kinase B (P-AKT), phosph-mammalian targets of rapamycin (P-mTOR), and phosph-p70 ribosomal s6 kinase (P-p70s6K) in tumor cells were likewise reduced after hinokitiol treatment. The transfection of cells with active P-AKT rescued hinokitiol-induced downregulation of P-gp, suggesting the involvement of Akt/mTOR/p70s6K signaling in P-gp expression. Our results showed that hinokitiol can chemosensitize cancer cells. These findings indicate that hinokitiol could enhance 5-Fluorouracil therapeutic effects in murine B16F10 and CT26 tumor cells via downregulation of the AKT/mTOR pathway.

Characterization of CRISPR-Cas Systems in Clinical Klebsiella pneumoniae Isolates Uncovers Its Potential Association With Antibiotic Susceptibility.

Prokaryotic CRISPR-Cas systems limit the acquisition of genetic elements and provide immunity against invasive bacteriophage. The characteristics of CRISPR-Cas systems in clinical Klebsiella pneumoniae isolates are still unknown. Here, 97 K. pneumoniae genomes retrieved from the Integrated Microbial Genomes & Microbiomes genome database and 176 clinical isolates obtained from patients with bloodstream (BSI, n = 87) or urinary tract infections (UTI, n = 89) in Taiwan, were used for analysis. Forty out of ninety-seven genomes (41.2%) had CRISPR-Cas systems identified by the combination of CRISPRFinder and cas1 gene sequence alignment. The phylogenetic trees revealed that CRISPR-Cas systems in K. pneumoniae were divided into two types (type I-E, 23; subtype I-E∗, 17) based on the sequences of Cas1 and Cas3 proteins and their location in the chromosome. The distribution of type I-E and I-E∗ CRISPR-Cas systems was associated with the multilocus sequence typing and the pulsed-field gel electrophoresis results. Importantly, no CRISPR-Cas system was identified in published genomes of clonal complex 258 isolates (ST11 and ST258), which comprise the largest multi-drug resistant K. pneumoniae clonal group worldwide. PCR with cas-specific primers showed that 30.7% (54/176) of the clinical isolates had a CRISPR-Cas system. Among clinical isolates, more type I-E CRISPR-Cas systems were found in UTI isolates (BSI, 5.7%; UTI, 11.2%), and subtype I-E∗ CRISPR-Cas systems were dominant in BSI isolates (BSI, 28.7%; UTI, 15.7%) (p = 0.042). Isolates which had subtype I-E∗ CRISPR-Cas system were more susceptible to ampicillin-sulbactam (p = 0.009), cefazolin (p = 0.016), cefuroxime (p = 0.039), and gentamicin (p = 0.012), compared to the CRISPR-negative isolates. The strains containing subtype I-E∗ CRISPR-Cas systems had decreased numbers of plasmids, prophage regions, and acquired antibiotic resistance genes in their published genomes. Here, we first revealed subtype I-E∗ CRISPR-Cas system in K. pneumoniae potentially interfering with the acquisition of phages and plasmids harboring antibiotic resistance determinants, and thus maintained these isolates susceptible to antibiotics.

Primary Sjögren's syndrome associated with Gitelman's syndrome presenting with muscular paralysis.

A 38-year-old woman presented with muscle cramping of 4 extremities and paralysis for months. Laboratory results showed an elevated antinuclear antibody titer; antibodies to the ribonucleoprotein antigen Ro; hypokalemia; hypomagnesemia with hyperreninemia, but abnormally high urine potassium and magnesium levels and low urine calcium levels; and a blunted diuretic effect to thiazide, but not furosemide, which met the criteria for Gitelman's syndrome (GS) and led to the diagnosis of primary Sjögren's syndrome (pSS). She received medical treatment, including a potassium supplement and aldosterone antagonist. GS as a presentation of pSS has never been reported in the literature. The features of renal diseases related to SS are reviewed. SS is the underlying cause of GS, which may precede the onset of the well-known sicca complex.