Clinical Significance of Cancer Stem Cell Markers in Primary and Metastatic Tissues in Patients With Breast Cancer.

BACKGROUND/AIM: This study aimed to examine the clinical significance of the protein expression of the cancer stem cell (CSC) markers ALDH1A1, CD133, CD44, and MSI-1 in primary and metastatic tissues of patients with breast cancer (BC). PATIENTS AND METHODS: ALDH1A1, CD133, CD44, and MSI-1 protein expression in pairs of primary and metastatic tissues of 55 patients with BC with metastases treated at Kanagawa Cancer Center between January 1970 and December 2016 were evaluated using immunohistochemical assay and their association with clinicopathological factors and survival was examined. RESULTS: There were no significant differences in CSC marker expression rates between primary and metastatic tissues for any CSC markers. Regarding the relationship between CSC marker expression in primary tissues and survival, patients with high CD133 expression had significantly lower recurrence-free survival (DFS) and overall survival. On multivariate analysis, they were also a poor independent predictor of DFS (hazard ratio=4.993, 95%CI=2.189-11.394, p=0.0001). In contrast, there was no significant association between the expression of any CSC marker in metastatic tissues and survival. CONCLUSION: CD133 expression in the primary BC tissue may be a useful risk factor for recurrence in patients with BC.

Lung cavitation in patients with anaplastic thyroid cancer treated with lenvatinib.

Background: Anaplastic thyroid cancer (ATC) is a rare malignancy with a poor prognosis. It accounts for 1-2% of all thyroid cancers. Lenvatinib is an orally administered inhibitor of vascular endothelial growth factor receptor (VEGFR)-1, -2, and -3, fibroblast growth factor receptor (FGFR)-1 to -4, platelet-derived growth factor receptor (PDGFR)-α, rearranged during transfection (RET), and KIT. There have been cases of pneumothorax caused by lung cavitation and collapse after administration of lenvatinib in ATC with lung metastasis. In this study, we investigate lung cavitation during treatment with lenvatinib in ATC patients with lung metastasis. Methods: All ATC patients with lung metastasis treated at our hospital with lenvatinib between November 2015 and May 2021 were selected from our electronic medical records. The primary objective was to determine the incidence of cavitation of lung metastasis of ATC in patients treated with lenvatinib. The secondary objective was to evaluate prognostic factors in ATC patients with lung metastasis treated with lenvatinib. Results: We identified 26 patients treated with lenvatinib for ATC with lung metastasis. Of these, 12 (46.2%) had cavitation with lung metastasis during lenvatinib treatment. The median overall survival (OS) was 128 days (79-228 days), and the cavitation (+) group had significantly longer OS than the cavitation (-) group [186 days (117-355 days) vs. 89 days (59-179 days), P=0.033]. Kaplan-Meier survival curves indicated a significant difference in OS was observed between the two groups (P=0.0293). Univariate analysis demonstrated lung cavitation was a significant prognostic factor (hazard ratio: 0.38, 95% CI: 0.16-0.93). Conclusions: Lung cavitation occurred in 46.2% of patients treated with lenvatinib for ATC with lung metastasis. Patients who developed lung cavitation had a significantly better prognosis than those who did not.

A case of familial hypocalciuric hypercalcemia type 1 due to CASR p.Pro55Leu mutation.

BACKGROUND: Familial hypocalciuric hypercalcemia (FHH) is a rare autosomal dominant disease, which requires differential diagnosis from relatively common primary hyperparathyroidism (PHPT) in order to avoid unnecessary surgery. CASE PRESENTATION: A 16-year-old female had been followed by the department of psychosomatic medicine at our institution. Throughout the follow-up period, her plasma calcium levels were high, plasma Pi levels were relatively low, and plasma intact PTH was relatively high. She was referred to our department to determine the cause of her hypercalcemia. Her 24 h urinary calcium excretion was as low as 100 mg/day, and calcium creatinine clearance ratio was below 0.01. Moreover, she had a family history of hypercalcemia (proband, her brother, and her father). The genetic testing for her family revealed that she, her brother, and her father were definitively diagnosed with FHH type 1 due to the heterozygous calcium-sensing receptor mutation (NM_00388:4:c.164C > T:p.Pro55Leu). CONCLUSION: We experienced a 16-year-old female with FHH, in whom genetic testing identified the heterozygous calcium-sensing receptor mutation (NM_00388:4:c.164C > T:p.Pro55Leu) as pathogenic, permitting a definitive diagnosis of FHH type 1. The genetic testing for calcium sensing receptor is beneficial to distinguish asymptomatic primary hyperparathyroidism from FHH.

Efficacy of Clinical Guidelines in Identifying All Japanese Patients with Hereditary Breast and Ovarian Cancer.

Clinical screening using the National Comprehensive Cancer Network (NCCN) testing criteria may fail to identify all patients with hereditary breast and ovarian cancers. Thus, this study aimed to evaluate the strategy of expanding target patients for genetic testing among Japanese patients. We reviewed the medical records of 91 breast cancer patients who underwent genetic testing. Among 91 patients, eight were diagnosed with pathogenic or likely pathogenic variants: BRCA1 (n = 4) and BRCA2 (n = 4). Among 50 patients meeting the testing criteria of the guidelines, 6 (12%) were diagnosed with pathogenic or likely pathogenic variants. The sensitivity and specificity of screening using the testing criteria were 75% and 47%, respectively. Expanding the NCCN criteria to include all women diagnosed with breast cancer aged ≤65 years achieved 88% sensitivity but 8% specificity. The expansion of the NCCN criteria could benefit Japanese patients; however, larger studies are necessary to change clinical practice.

Role of Ras in regulation of intestinal epithelial cell homeostasis and crosstalk with Wnt signaling.

Cross talk between different signaling pathways is thought to be important for regulation of homeostasis of, as well as oncogenesis of, the intestinal epithelium. Expression of an active form of K-Ras specifically in intestinal epithelial cells (IECs) of mice (IEC-RasDA mice) resulted in the development of hyperplasia in the small intestine and colon of mice. IEC-RasDA mice also manifested the increased proliferation of IECs. In addition, the number of goblet cells markedly increased, while that of Paneth cells decreased in IEC-RasDA mice. Development of intestinal organoids was markedly enhanced for IEC-RasDA mice compared with control mice. Whereas, the expression of Wnt target genes was significantly reduced in the in intestinal crypts from IEC-RasDA mice compared with that apparent for the control. Our results thus suggest that K-Ras promotes the proliferation of IECs as well as generation of goblet cells. By contrast, Ras counter-regulates the Wnt signaling and thereby contribute to the proper regulation of intestinal epithelial cell homeostasis.

Effects of physician's diabetes self-management education using Japan Association of Diabetes Education and Care Diabetes Education Card System Program and a self-monitoring of blood glucose readings analyzer in individuals with type 2 diabetes: An exploratory, open-labeled, prospective randomized clinical trial.

AIMS/INTRODUCTION: This 6-month, single-center, prospective, open-labeled, randomized trial was designed to investigate whether physicians' diabetes self-management education using an education tool developed by the Japan Association of Diabetes Education and Care and a self-monitoring of blood glucose (SMBG) analyzer improves glycemic control in individuals with type 2 diabetes receiving insulin and SMBG. MATERIALS AND METHODS: Participants were randomized into intervention (I) and control (C) groups. Both groups received physicians' diabetes self-management education at each hospital visit, whereas the Japan Association of Diabetes Education and Care education tool and the SMBG readings analyzer was used in group I, but not group C. All participants filled out a diabetes treatment-related quality of life form and an original questionnaire on SMBG use with five questions (Q1-Q5) before and after the study period. RESULTS: A total of 76 individuals were recruited and randomized. Glycated hemoglobin (HbA1c) was significantly improved during the study period in group I, whereas no significant change was observed in group C. The change in HbA1c was greater in group I, although it did not reach statistical significance. The diabetes treatment-related quality of life total score was not changed in either group. Interestingly, the score of Q1 ("How important is SMBG to you?") in the SMBG questionnaire was unchanged in group I, whereas it was significantly decreased in group C. HbA1c change was independently associated with changes in insulin dose and SMBG Q1 score. CONCLUSION: Greater HbA1c-lowering by physicians' diabetes self-management education using the Japan Association of Diabetes Education and Care education tool and SMBG analyzer in individuals with type 2 diabetes receiving insulin and SMBG was suggested, but not confirmed.

Effects of glucagon-like peptide-1 receptor agonists on secretions of insulin and glucagon and gastric emptying in Japanese individuals with type 2 diabetes: A prospective, observational study.

AIMS/INTRODUCTION: Differences in the glucose-lowering mechanisms of glucagon-like peptide-1 receptor agonists (GLP-1RAs) have been noted. Clarifying these differences could facilitate the choice of optimal drugs for individuals with type 2 diabetes and requires investigation in a clinical setting. MATERIALS AND METHODS: A single-arm, prospective, observational study was conducted to evaluate the effects of various GLP-1RAs on postprandial glucose excursion, secretions of insulin and glucagon as well as on the gastric emptying rate. Participants were subjected to meal tolerance tests before and 2 weeks and 12 weeks after GLP-1RA initiation. Effects on postprandial secretions of glucose-dependent insulinotropic polypeptide (GIP) and apolipoprotein B48 were also investigated. RESULTS: Eighteen subjects with type 2 diabetes received one of three GLP-1RAs, i.e., lixisenatide, n = 7; liraglutide, n = 6; or dulaglutide, n = 5. While 12-week administration of all of the GLP-1RAs significantly reduced HbA1c, only lixisenatide and liraglutide, but not dulaglutide, significantly reduced body weight. Postprandial glucose elevation was improved by all of the GLP-1RAs. Postprandial insulin levels were suppressed by lixisenatide, while insulin levels were enhanced by liraglutide. Postprandial glucagon levels were suppressed by lixisenatide. The gastric emptying rate was significantly delayed by lixisenatide, while liraglutide and dulaglutide had limited effects on gastric emptying. GIP secretion was suppressed by lixisenatide and liraglutide. Apolipoprotein B48 secretion was suppressed by all of the GLP-1RAs. CONCLUSIONS: All of the GLP-1RAs were found to improve HbA1c in a 12-week prospective observational study in Japanese individuals with type 2 diabetes. However, differences in the mechanisms of the glucose-lowering effects and body weight reduction were observed.

Quality Control of Breast Cancer Surgery Samples: Introducing Time Stamp Checking.

Background: Analytical information obtained from clinical tissue samples has recently become more important due to recent advancements in the clinical practice of medicine, for example, gene panel testing. However, acquiring and managing the sample quality, which greatly influences the analyses, are not sufficient and hence requires immediate attention. We introduced time stamp (TS) recording and documentation using the Standard PREanalytical Code (SPREC) for breast cancer surgery samples to monitor and control their quality. Materials and Methods: The TS recording used SPREC for quality control of each sample by recording seven factors: type of sample, type of collection, warm ischemia time (WIT), cold ischemia time (CIT), fixation type, fixation time (FT), and long-term storage. The responsibilities to record each factor were assigned among group members (breast surgeons, anesthesiologists, pathologists, operating room nurses, and medical technologists in pathology). Results: Records based on SPREC were recorded for 393 surgical cases of first-time breast cancer patients performed at the Kanagawa Cancer Center from May 2018 to April 2019. The vascular clamp time was defined as when skin flap formation was completed, regardless of the surgical procedure. An anesthesiologist recorded the vascular clamp time and sample collection time, and the pathologist recorded the fixation start time and fixation end time. WIT was 23 (3-116) minutes (breast-conserving surgery, 11 [3-38] minutes; mastectomy, 26 [5-116] minutes; and nipple-sparing mastectomy, 39 [31-43] minutes), CIT was 37 (3-1052) minutes, and FT was 43 (17-115) hours. The median CIT and FT were significantly shortened after introducing the TS system, and the variabilities were reduced. Conclusion: A TS system for quality control of breast cancer surgical sample functions well due to the establishment of highly versatile WIT and a working group consisting of multiple members of different occupations who shared roles.

Effects of ChREBP deficiency on adrenal lipogenesis and steroidogenesis.

Carbohydrate response element-binding protein (ChREBP) is critical in the regulation of fatty acid and triglyceride synthesis in the liver. Interestingly, Chrebp-/- mice show reduced levels of plasma cholesterol, which is critical for steroid hormone synthesis in adrenal glands. Furthermore, Chrebp mRNA expression was previously reported in human adrenal glands. Thus, it remains to be investigated whether ChREBP plays a role directly or indirectly in steroid hormone synthesis and release in adrenal glands. In the present study, we find that Chrebp mRNA is expressed in mouse adrenal glands and that ChREBP binds to carbohydrate response elements. Histological analysis of Chrebp-/- mice shows no adrenal hyperplasia and less oil red O staining compared with that in WT mice. In adrenal glands of Chrebp-/- mice, expression of Fasn and Scd1, two enzymes critical for fatty acid synthesis, was substantially lower and triglyceride content was reduced. Expression of Srebf2, a key transcription factor controlling synthesis and uptake of cholesterol and the target genes, was upregulated, while cholesterol content was not significantly altered in the adrenal glands of Chrebp-/- mice. Adrenal corticosterone content and plasma adrenocorticotropic hormone and corticosterone levels were not significantly altered in Chrebp-/- mice. Consistently, expression of genes related to steroid hormone synthesis was not altered. Corticosterone secretion in response to two different stimuli, namely 24-h starvation and cosyntropin administration, was also not altered in Chrebp-/- mice. Taking these results together, corticosterone synthesis and release were not affected in Chrebp-/- mice despite reduced plasma cholesterol levels.

Direct evidence that KNDy neurons maintain gonadotropin pulses and folliculogenesis as the GnRH pulse generator.

The gonadotropin-releasing hormone (GnRH) pulse is fundamental for mammalian reproduction: GnRH pulse regimens are needed as therapies for infertile women as continuous GnRH treatment paradoxically inhibits gonadotropin release. Circumstantial evidence suggests that the hypothalamic arcuate KNDy neurons expressing kisspeptin (encoded by Kiss1), neurokinin B (encoded by Tac3), and dynorphin A serve as a GnRH pulse generator; however, no direct evidence is currently available. Here, we show that rescuing >20% KNDy neurons by transfecting Kiss1 inside arcuate Tac3 neurons, but not outside of these neurons, recovered folliculogenesis and luteinizing hormone (LH) pulses, an indicator of GnRH pulses, in female global Kiss1 knockout (KO) rats and that >90% conditional arcuate Kiss1 KO in newly generated Kiss1-floxed rats completely suppressed LH pulses. These results first provide direct evidence that KNDy neurons are the GnRH pulse generator, and at least 20% of KNDy neurons are sufficient to maintain folliculogenesis via generating GnRH/gonadotropin pulses.

Effect of Carbon Black Nanoparticle on Neonatal Lymphoid Tissues Depending on the Gestational Period of Exposure in Mice.

Introduction: Particulate air pollution, containing nanoparticles, enhances the risk of pediatric allergic diseases that is potentially associated with disruption of neonatal immune system. Previous studies have revealed that maternal exposure to carbon black nanoparticles (CB-NP) disturbs the development of the lymphoid tissues in newborns. Interestingly, the CB-NP-induced immune profiles were observed to be different depending on the gestational period of exposure. It is important to identify the critical exposure period to prevent toxic effects of nanoparticles on the development of the immune system. Therefore, the present study was aimed to investigate the effect of CB-NP on the development of neonatal lymphoid tissues in mice, depending on the gestational period of exposure. Methods: Pregnant ICR mice were treated with a suspension of CB-NP (95 µg/kg body weight) by intranasal instillation; the suspension was administered twice during each gestational period as follows: the pre-implantation period (gestational days 4 and 5), organogenesis period (gestational days 8 and 9), and fetal developmental period (gestational days 15 and 16). The spleen and thymus were collected from offspring mice at 1, 3, and 5-days post-partum. Splenocyte and thymocyte phenotypes were examined by flow cytometry. Gene expression in the spleen was examined by quantitative reverse transcription-polymerase chain reaction. Results: The numbers of total splenocytes and splenic CD3-B220- phenotype (non-T/non-B lymphocytes) in offspring on postnatal day 5 were significantly increased after exposure to CB-NP during the organogenesis period compared with other gestational periods of exposure and control (no exposure). In contrast, expression levels of mRNA associated with chemotaxis and differentiation of immune cells in the spleen were not affected by CB-NP exposure during any gestational period. Conclusion: The organogenesis period was the most susceptible period to CB-NP exposure with respect to lymphoid tissue development. Moreover, the findings of the present and previous studies suggested that long-term exposure to CB-NP across multiple gestational periods including the organogenesis period, rather than acute exposure only organogenesis period, may more severely affect the development of the immune system.

Regulation of colonic epithelial cell homeostasis by mTORC1.

Cell signaling important for homeostatic regulation of colonic epithelial cells (CECs) remains poorly understood. Mammalian target of rapamycin complex 1 (mTORC1), a protein complex that contains the serine-threonine kinase mTOR, mediates signaling that underlies the control of cellular functions such as proliferation and autophagy by various external stimuli. We here show that ablation of tuberous sclerosis complex 2 (Tsc2), a negative regulator of mTORC1, specifically in intestinal epithelial cells of mice resulted in increased activity of mTORC1 of, as well as increased proliferative activity of, CECs. Such Tsc2 ablation also reduced the population of Lgr5-positive colonic stem cells and the expression of Wnt target genes in CECs. The stimulatory phosphorylation of the kinase Akt and inhibitory phosphorylation of glycogen synthase kinase 3ß were both markedly decreased in the colon of the Tsc2 conditional knockout (CKO) mice. Development of colonic organoids with cryptlike structures was enhanced for Tsc2 CKO mice compared with control mice. Finally, Tsc2 CKO mice manifested increased susceptibility to dextran sulfate sodium-induced colitis. Our results thus suggest that mTORC1 activity promotes the proliferation of, as well as the expression of Wnt target genes in, CECs and thereby contributes to colonic organogenesis and homeostasis.

Development of an Anti-Sheep Podoplanin Monoclonal Antibody PMab-256 for Immunohistochemical Analysis of Lymphatic Endothelial Cells.

Sensitive and specific monoclonal antibodies (mAbs) targeting podoplanin (PDPN) are needed for immunohistochemical analyses as a marker for lymphatic endothelial cells. We recently have developed anti-PDPN mAbs against many species, including human, mouse, rat, rabbit, dog, cat, bovine, pig, Tasmanian devil, alpaca, tiger, whale, goat, horse, and bear. However, anti-sheep PDPN (sPDPN) has not yet been established. In this study, we used the Cell-Based Immunization and Screening method for the development of anti-sPDPN mAbs. RAP14 tag was added to N-terminus of sPDPN, and anti-RAP14 tag mAb (PMab-2) was used to detect the expression level of sPDPN in flow cytometry and western blot. We immunized mice with sPDPN-overexpressing Chinese hamster ovary (CHO)-K1 (CHO/sPDPN) cells and screened mAbs against sPDPN using flow cytometry. One of the mAbs, PMab-256 (IgG1, kappa), specifically detected CHO/sPDPN cells by flow cytometry and western blot. Furthermore, PMab-256 stained type I alveolar cells of lung, renal glomerulus and Bowman's capsule, and lymphatic endothelial cells of lung and colon. Our findings suggest the potential usefulness of PMab-256 for the functional analyses of sPDPN.

Development of Novel Mouse Monoclonal Antibodies Against Human CD19.

CD19 is a type I transmembrane glycoprotein belonging to the immunoglobulin superfamily. It is expressed in normal and neoplastic B cells, and it modulates the threshold of B cell activation for amplifying B cell receptor signaling. Blinatumomab (a CD3-CD19-bispecific T cell-engaging antibody) and tisagenlecleucel (genetically modified T cells that express a CD19 chimeric antigen receptor [CART-19]) provide significant benefits for patients with CD19-positive relapsed or refractory B cell malignancies. In this study, we first employed the Cell-Based Immunization and Screening (CBIS) method to produce anti-CD19 monoclonal antibodies using CD19-overexpressing cells for both immunization and screening. One established clone-C19Mab-1-proved to be useful in flow cytometry assays against lymphoma cell lines, such as BALL-1, P30/OHK, and Raji. Second, the extracellular domain of CD19 was immunized into mice, and enzyme-linked immunosorbent assays were performed for the first screening. One established clone-C19Mab-3-was determined to be useful for Western blotting and immunohistochemical analysis. Due to their complementary utility, a combination of C19Mab-1 (established using CBIS) and C19Mab-3 (established using conventional method) could be useful for the pathological analysis of CD19.

Epitope Mapping of PMab-241, a Lymphatic Endothelial Cell-Specific Anti-Bear Podoplanin Monoclonal Antibody.

Anti-bear podoplanin (bPDPN) monoclonal antibodies (mAbs), including PMab-247 and PMab-241, have been previously established. Although PMab-247 has shown positive immunostaining for lymphatic endothelial cells (LECs), type I alveolar cells of the lung, and podocytes of the kidney, PMab-241 stains LECs but does not react with lung type I alveolar cells. PDPN possesses three platelet aggregation-stimulating (PLAG) domains (PLAG1, PLAG2, and PLAG3) and the PLAG-like domain (PLD). The binding epitope of PMab-247 was previously determined to include bPDPN residues Asp76, Arg78, Glu80, and Arg82. Among these, Glu80 and Arg82 are included in PLD of bPDPN. The purpose of this study is to determine the binding epitope of PMab-241 and to clarify the difference between these two anti-bPDPN mAbs. Analysis of bPDPN deletion mutants revealed that the N-terminus of the PMab-241 epitope exists between amino acids (aa) 75 and 80 of bPDPN. In addition, analysis of bPDPN point mutants demonstrated that the critical epitope of PMab-241 includes Thr75, Asp76, and Arg78 of bPDPN. The binding epitopes of PMab-241 and PMab-247 seem to overlap, but this slight difference may be sufficient to provide the specificity of PMab-241 to discriminate LECs from type I alveolar cells of the lung.

Antibody-Drug Conjugates Using Mouse-Canine Chimeric Anti-Dog Podoplanin Antibody Exerts Antitumor Activity in a Mouse Xenograft Model.

Antibody-drug conjugates (ADCs), which consist of a monoclonal antibody (mAb), a linker, and a payload, can deliver a drug to cancer tissues. We previously produced an anti-dog podoplanin (dPDPN) mAb, PMab-38, which reacts with dPDPN-expressing canine melanomas and squamous cell carcinomas (SCCs), but not with dPDPN-expressing canine type I alveolar cells or lymphatic endothelial cells, indicating that PMab-38 possesses cancer specificity. In this study, we developed an ADC, P38B-DM1, using the mouse-canine chimeric anti-dPDPN antibody, P38B as the antibody, a peptide linker, and emtansine as the payload using the chemical conjugation by affinity peptide (CCAP) method. We investigated its cytotoxicity against dPDPN-overexpressed Chinese hamster ovary (CHO/dPDPN) cells in vitro and its antitumor activity using a mouse xenograft model of CHO/dPDPN cells. P38B-DM1 showed cytotoxicity to CHO/dPDPN cells in a dose-dependent manner in vitro. Furthermore, P38B-DM1 exhibited higher antitumor activity than P38B in the mouse xenograft model. These results suggest that P38B-DM1, developed using the CCAP method, is useful for antibody therapy against dPDPN-expressing canine SCCs and melanomas.

ß-N-methylamino-L-alanine (BMAA) suppresses cell cycle progression of non-neuronal cells.

ß-N-methylamino-L-alanine (BMAA), a natural non-proteinaceous amino acid, is a neurotoxin produced by a wide range of cyanobacteria living in various environments. BMAA is a candidate environmental risk factor for neurodegenerative diseases such as amyotrophic lateral sclerosis and Parkinson-dementia complex. Although BMAA is known to exhibit weak neuronal excitotoxicity via glutamate receptors, the underlying mechanism of toxicity has yet to be fully elucidated. To examine the glutamate receptor-independent toxicity of BMAA, we investigated the effects of BMAA in non-neuronal cell lines. BMAA potently suppressed the cell cycle progression of NIH3T3 cells at the G1/S checkpoint without inducing plasma membrane damage, apoptosis, or overproduction of reactive oxygen species, which were previously reported for neurons and neuroblastoma cells treated with BMAA. We found no evidence that activation of glutamate receptors was involved in the suppression of the G1/S transition by BMAA. Our results indicate that BMAA affects cellular functions, such as the division of non-neuronal cells, through glutamate receptor-independent mechanisms.

Relationship between deterioration of glycated hemoglobin-lowering effects in dipeptidyl peptidase-4 inhibitor monotherapy and dietary habits: Retrospective analysis of Japanese individuals with type 2 diabetes.

The present study was designed to assess possible relationships between deterioration of the glycated hemoglobin (HbA1c)-lowering effects in dipeptidyl peptidase-4 inhibitor (DPP4i) monotherapy and macronutrient intake among individuals with type 2 diabetes. Type 2 diabetes patients who began and continued DPP4i monotherapy without any prescription change for 1 year were retrospectively stratified into two groups: (i) patients who maintained their HbA1c levels during the 0.5- to 1-year period after DPP4i initiation (group A, ΔHbA1c [1-0.5 year] <0.4%, n = 53); and (ii) those whose HbA1c levels increased [group B, ΔHbA1c (1-0.5 year] ≥0.4%, n = 10). Group B had significantly higher ΔHbA1c (1-0.5 year), Δbodyweight (1-0.5 year) and fat intake, especially of saturated and monounsaturated fats; the carbohydrate and protein intake were similar between groups. Multiple regression analyses showed that fat intake, especially saturated fat intake, was significantly correlated with ΔHbA1c (1-0.5 year). Thus, dietary habits, especially saturated fat intake, might well contribute to deterioration of the HbA1c-lowering effects in DPP4i monotherapy.

Cytotoxicity of Tirapazamine (3-Amino-1,2,4-benzotriazine-1,4-dioxide)-Induced DNA Damage in Chicken DT40 Cells.

Tirapazamine (TPZ) is an anticancer drug with highly selective cytotoxicity toward hypoxic cells. TPZ is converted to a radical intermediate under hypoxic conditions, and this intermediate interacts with intracellular macromolecules, including DNA. TPZ has been reported to indirectly induce DNA double-strand breaks (DSBs) through the formation of various intermediate DNA lesions under hypoxic conditions. Although the topoisomerase II-DNA complex has been identified as one of these intermediates, other lesions have not yet been defined. In order to obtain a deeper understanding of the mechanisms responsible for the selective cytotoxicity of TPZ toward hypoxic cells, its cellular sensitivity was systematically examined with genetically isogenic DNA-repair-deficient mutant DT40 cell lines. Our results showed that tdp1-/-, tdp2-/-, parp1-/-, and aptx1-/- cells displayed hypersensitivity to TPZ only under hypoxic conditions. These results strongly suggest that the accumulation of the topoisomerase I-trapped DNA complex, topoisomerase II-trapped DNA complex, and abortive ligation products with 5'-AMP are the potential causes of TPZ-induced hypoxic cell death. Furthermore, our genetic analysis revealed that under normoxic conditions (as well as hypoxic conditions), TPZ exhibited significant cytotoxicity toward cell lines deficient in homologous recombination, nonhomologous end joining, base excision repair, and translesion synthesis. Ascorbic acid, a radical scavenger, suppressed TPZ-induced cytotoxicity toward normoxic cells. These results suggest the involvement of oxidative DNA damage and DSBs produced by reactive oxygen species generated from superoxide, a byproduct of the oxidation of TPZ radical intermediates in normoxic cells. Collectively, our results demonstrate that TPZ induces oxidative DNA damage under normoxic and hypoxic conditions and selectively introduces abortive topoisomerase-DNA complexes and unligatable DNA ends under hypoxic conditions.

Comparison between procalcitonin and C-reactive protein in predicting bacteremias and confounding factors: a case-control study.

BACKGROUND: The detection of infectious bacteria in blood culture samples is important for diagnosis and treatment, but this requires 1-2 days at least, and is not adequate as a rapid test. Therefore, we have investigated the diagnostic ability and the optimal cutoff value of procalcitonin (PCT) and C-reactive protein (CRP) for predicting the bacteremias using receiver operating characteristic (ROC) curves and relative cumulative frequency distribution (RCD) curves. METHODS: A case-control study was performed in inpatients (852 subjects: 426 positive cultures and 426 negative cultures) from January 1 to December 31, 2014. We retrospectively investigated their blood culture and blood chemistry findings recorded in this period using electronic medical records. RESULTS: Area under the ROC curve of PCT and CRP were 0.79 and 0.66, respectively. The optimal cutoff values were 0.5 µg/L with a sensitivity of 70% and specificity of 70% for PCT and 50.0 mg/L with a sensitivity of 63% and specificity of 65% for CRP. When the optimal cutoff value was treated as a reference, the odds ratio (OR) was 71.11 and the hazard ratio (HR) was 6.27 for PCT >2.0 µg/L, and the risk of blood culture positivity was markedly elevated. PCT levels were significantly higher in the population with Gram-negative rod (GNR) infections than in the population with Gram-positive coccal (GPC) infections. CONCLUSIONS: The elevation of CRP and PCT were significantly associated with bacteremias. PCT was superior to CRP as a diagnostic indicator for predicting bacteremias, for discriminating bacterial from nonbacterial infections, and for determining bacterial species.