A small-molecule degron with a phenylpropionic acid scaffold.

Targeted protein degradation (TPD), employing proteolysis-targeting chimeras (PROTACs) composed of ligands for both a target protein and ubiquitin ligase (E3) to redirect the ubiquitin-proteasome system (UPS) to the target protein, has emerged as a promising strategy in drug discovery. However, despite the vast number of E3 ligases, the repertoire of E3 ligands utilized in PROTACs remains limited. Here, we report the discovery of a small-molecule degron with a phenylpropionic acid skeleton, derived from a known ligand of S-phase kinase-interacting protein 2 (Skp2), an E3 ligase. We used this degron to design PROTACs inducing proteasomal degradation of HaloTag-fused proteins, and identified key structural relationships. Surprisingly, our mechanistic studies excluded the involvement of Skp2, suggesting that this degron recruits other protein(s) within the UPS.

Chronobioethics: Symphony of biological clocks observed by 7-day/24-hour ambulatory blood pressure monitoring and cardiovascular health.

BACKGROUND: The high prevalence of desynchronized biological rhythms is becoming a primary public health concern. We assess complex and diverse inter-modulations among multi-frequency rhythms present in blood pressure (BP) and heart rate (HR). SUBJECTS: and Methods: We performed 7-day/24-hour Ambulatory BP Monitoring in 220 (133 women) residents (23 to 74 years) of a rural Japanese town in Kochi Prefecture under everyday life conditions. RESULTS: A symphony of biological clocks contributes to the preservation of a synchronized circadian system. (1) Citizens with an average 12.02-h period had fewer vascular variability disorders than those with shorter (11.37-h) or longer (12.88-h) periods (P<0.05), suggesting that the circasemidian rhythm is potentially important for human health. (2) An appropriate BP-HR coupling promoted healthier circadian profiles than a phase-advanced BP: lower 7-day nighttime SBP (106.8 vs. 112.9 mmHg, P=0.0469), deeper nocturnal SBP dip (20.5% vs. 16.8%, P=0.0101), and less frequent incidence of masked non-dipping (0.53 vs. 0.86, P=0.0378), identifying the night as an important time window. CONCLUSION: Adaptation to irregular schedules in everyday life occurs unconsciously at night, probably initiated from the brain default mode network, in coordination with the biological clock system, including a reinforced about 12-hour clock, as "a biological clock-guided core integration system".

[Efficient Commissioning of the Radiotherapy Treatment Planning System with the Golden Beam Data].

Commissioning of a linear accelerator (Linac) and treatment planning systems (RTPs) for clinical use is complex and time-consuming, typically 3-4 months in total. However, based on clinical needs and economics, hospitals desire early clinical starts for patients, and various studies have been conducted for shortening the preparation period. One of the methods to shorten the period is using golden beam data (GBD). The purpose of this study was to shorten the commissioning period without reducing accuracy and to simplify commissioning works while improving safety. We conducted commissioning of the RTPs before installing the Linac using GBD, and carried out verification immediately after the acceptance test. We used TrueBeam STx (Varian Medical Systems) and Eclipse (ver. 13.7, Varian Medical Systems) for RTPs and anisotropic analysis algorithm (AAA) and AcurosXB (AXB) for calculation algorithms. The difference between GBD and the measured beam data was 0.0 ± 0.2% [percentage depth dose (PDDs) ] and -0.1 ± 0.2% (Profiles) with X-ray, and -1.2 ± 1.3% (PDDs) with electrons. The difference between the calculated dose and the measured dose was 0.1 ± 0.3% (AAA) and 0.0 ± 0.3% (AXB) under homogeneous conditions, and 0.7 ± 1.4% (AAA) and 0.6 ± 1.1% (AXB) under heterogeneous conditions. We took 43 days from the end of the acceptance test to the start of clinical use. We found that the preparation period for clinical use can be shortened without reducing the accuracy, by thinning out the number of measurement items using GBD.

The effect of night shift work on the expression of clock genes in beard hair follicle cells.

OBJECTIVE: Shift work encompasses a broad range of work time arrangements. However, how shift work affects the circadian expression of clock genes remains to be explored. The objective of this study was to evaluate the pattern of clock gene expression in shift workers in the field. METHODS: We examined clock gene expression in Japanese men who work: (1) one night shift followed by a day off (caregivers: nurses and doctors; the one-night group); (2) three or more consecutive night shifts (factory workers; the consecutive-night group); or (3) daytime only (the daytime group), using beard follicle samples. The expression of Period3, Nuclear Receptor Subfamily 1 Group D Member 1 (Nr1d1), and Nuclear Receptor Subfamily 1 Group D Member 2 (Nr1d2) was examined by real-time polymerase chain reaction. RESULTS: Period3 expression in the daytime and one-night groups together with Nr1d2 expression in the one-night group fitted a 24-h-period cosine curve better than in the consecutive-night group (p = 0.004, 0.012, and 0.001, respectively). The level of overall Period3 gene expression, calibrated with that of 18S-rRNA, was decreased in the consecutive-night group compared with that in the daytime group (p = 0.006). The patterns of Period3 and Nr1d2 expression in the daytime and one-night groups were more coherent than those in the consecutive-night group. CONCLUSIONS: These results suggest that night shift work affects the rhythms and levels of circadian Period3 and Nr1d2 expression dependent on the shift schedule or type of the shift; however, there is substantial variation between individuals.

Control banding assessment of exposure of offset printing workers to organic solvents.

OBJECTIVES: We aimed to assess the exposure of offset printing workers to hazardous substances in the rinsing processes of small-sized companies using a control banding method. METHODS: We obtained half-year amounts of hazardous substances purchased through a questionnaire survey and the hazardous information from the safety data sheets (SDSs) and related literature. RESULTS: The amount of petroleum kerosine and carbon hydride markedly increased in 2013 compared with that in 2010. In contrast, the amount of dichloromethane (DCM) decreased in 2013, and 1,2-dichloropropane (DCP) was not used in either 2010 or 2013. Mineral oil and xylene were allocated to Hazard Group D and judged to require Control Approach 3. In addition to DCM with Global Harmonization System's carcinogenic category 1, mildly treated mineral oil and solvent naphtha, allocated into Hazard Group E, are carcinogenic to humans and were judged to require Control Approach 4. There are two limitations of the control banding assessment: first, only limited and scarce hazard information could be obtained from SDSs, and second, safe-sided judgment for control technology for industrial hygiene. CONCLUSION: Small-sized enterprises are encouraged to implement control banding assessment for hazardous substances and to access expert advice available from Regional Industrial Health Centers. Easy access to appropriate expert advice is important to compensate for the limited and scarce hazard information and safe-sided judgment for control technology for Control Approaches 3 and 4.

[A case of rubella encephalitis presenting as clinically mild encephalitis/encephalopathy with a reversible splenial lesion].

A 26-year-old male was admitted because of a fever, headache and disturbance of consciousness with lymph node swelling of the neck two days after developing a rash. A neurological examination revealed restlessness with irritability in response to sensory stimuli, such as an injection. Diffusion-weighted brain magnetic resonance imaging (MRI) revealed a hyperintense ovoid lesion in the splenium of the corpus callosum, which showed a low coefficient in the ADC map: the lesion disappeared after 22 days. An enzyme immunoassay (EIA) of the serum and cerebrospinal IgM were positive for rubella virus. The patient was therefore diagnosed with rubella encephalitis. He recovered gradually and was discharged on day 19 after the onset of symptoms without any sequelae. To our knowledge, this is the first case of rubella encephalitis presenting as clinically mild encephalitis/encephalopathy with a reversible splenial lesion (MERS). Although the exact mechanism underlying the development of rubella encephalitis is not well established, this case indicated that our patient had an immune-mediated secondary encephalitis. According to the survey of the pandemic of rubella from 2012 to April 2013 in Japan, the incidence of rubella encephalitis is thought to be relatively higher than was previously noted. This emphasizes the importance of vaccination for preventing encephalitis.

Characteristics of fluid composition of left displaced abomasum in beef cattle fed high-starch diets.

To clarify the pathophysiology of left displaced abomasum (LDA), beef cattle fed high-starch diets were examined. The abomasal pH in beef cattle with LDA was lower than that in non-LDA reference animals (data from beef cattle at an abattoir), suggesting that it facilitated acidity. Bacteriological examinations of the abomasal fluid in cattle with LDA revealed the presence of Pseudomonas spp., Clostridium spp. and Candida spp., presumably reflecting the accelerated influx of ruminal fluid into the abomasum. Biochemical analyses of serum revealed that LDA cattle had higher lactic acid and lower vitamin A and E levels than non-LDA reference animals. These results indicate that beef cattle with LDA may suffer from vitamin A and E deficiencies due to maldigestion of starch and the high acidity of abomasal fluid.

Administration of angiotensin receptor II blockade improves vascular function, urinary albumin excretion, and left ventricular hypertrophy in low-risk essential hypertensive patients receiving antihypertensive treatment with calcium channel blockers.

We examined calcium channel blockers (CCBs) and angiotensin receptor II blockers in low-risk hypertensives to evaluate renal, vascular function and left ventricular mass (LVM) from the viewpoint of salt intake (SI). Low-risk hypertensives who had not met blood pressure (BP) goals with CCB were administered telmisaltan. Office and home BP, urinary albumin excretion (uAE), brachial ankle pulse wave velocity (baPWV) and LVM were significantly reduced. uAE and baPWV correlated with SI. It is therefore necessary to evaluate the organ-protecting effects from the viewpoint of SI. In low-risk hypertensives, telmisartan with CCB improves renal, vascular function and LVM.

Switching to tacrolimus extended-release improved the effectiveness of immunosuppressive therapy in a heart transplant patient: A case report.

We report on a 25-year-old female heart transplant patient who presented with recurrent episodes of cellular rejection due to decreased adherence to immunosuppressive therapy. She received a heart transplantation in 1994 when she was 10 years old. In order to improve her adherence to immunosuppressive therapy, switching to the once-daily extended-release formulation of tacrolimus was performed in a step-wise fashion. First, the twice-daily formulation of cyclosporin A was replaced with the twice-daily preparation of tacrolimus. When the trough blood levels of tacrolimus reached a plateau in the range of 5.0 ng/mL, it was changed to the once-daily extended-release formulation of tacrolimus after confirming the absence of new rejection episodes. There were no significant changes in renal function before and after the switch. After being discharged from the hospital, the patient made significant advancements in adherence to immunosuppressive therapy. Her subsequent clinical course was uneventful, with no adverse events observed. Most patients who undergo solid organ transplantation must receive lifelong immunosuppressive therapy. This case demonstrates that conversion to the extended-release formulation of tacrolimus from other calcineurin inhibitor preparations is a reasonable choice to consider in the management of compromised immunosuppressive therapy adherence in heart transplant patients during the late posttransplant period.

Chemotactic properties and absence of the formyl peptide receptor in ferret (Mustela putorius furo) neutrophils.

This study describes a chemotaxis assay of ferret polymorphonuclear cells (PMNs). The optimal conditions for this chemotaxis assay were investigated for three chemoattractants: zymosan activated serum (ZAS), recombinant human interleukin-8 (rhIL-8) and N-formyl-Met-Leu- Phe (fMLF). In this study, ferret polymorphonuclear cells (PMNs) reacted to ZAS and rhIL-8, but not fMLF. The optimal concentration of ZAS and rhIL-8 were 5% and 100 ng/ml, respectively. The optimal incubation time of each reagent was 60 min. Due to the lack of response shown from fMLF, the existence of formyl peptide receptors (FPR) on ferret PMNs was investigated by evaluating FPR binding using flow cytometry. The receptor was not detected, implying that ferret neutrophils may lack FPR. This study confirms the fundamental experimental conditions for ferret PMNs chemotaxis and elucidates new findings concerning FPR in ferret neutrophils.

Prediction of number of apheresis procedures necessary in healthy donors to attain minimally required peripheral blood CD34+ cells.

BACKGROUND: Allogeneic peripheral blood stem cell (PBSC) transplantation is widely performed as a curative therapy for hematopoietic malignancies. Donors for PBSC harvest (PBSCH) are usually healthy subjects and undergo granulocyte-colony-stimulating factor treatment and apheresis procedures. A considerable proportion of donors experience poor mobilization, necessitating additional harvesting or marrow collection or remobilization. Although some characteristics have been reported to correlate with poor mobilization, they may not be taken into account in selecting PBSC donors. To protect healthy donors, it is preferable to predict the number of apheresis procedures needed for PBSCH before the procedure is initiated. STUDY DESIGN AND METHODS: A retrospective cohort study of 83 subjects was conducted, using statistical models to predict the probability of obtaining a sufficient number of CD34+ cells (>or=2.0 x 10(6)/kg) in the first to the third apheresis procedures and the probability of failure to obtain sufficient cells within three apheresis sessions. This study explored potential candidate factors in an ordinal probit regression analysis. RESULTS: Significant factors predicting successful PBSCH were donor age, donor sex, and body weight difference between donor and recipient. The predictive model showed good agreement with the observed number of apheresis sessions. Simulation tables are presented with this model. CONCLUSION: The statistical model developed to predict the number of apheresis procedures for PBSCH may be useful for planning PBSCH in clinical practice.

Dock2 participates in bone marrow lympho-hematopoiesis.

Dock2 has been shown to be indispensable for chemotaxis of mature lymphocytes as a critical Rac activator. However, the functional expression of Dock2 in immature hematopoietic cells is unclear. In this study, we demonstrate that Dock2 is broadly expressed in bone marrow (BM) hematopoietic compartment, including hematopoietic stem/progenitor cell (HSC/HPC) fraction. Response of Dock2-/- HPCs to CXCL12 in chemotaxis and actin polymerization in vitro was impaired, although alpha4 integrin activation by CXCL12 was not altered. Myelosuppressive stress on HSCs in vivo, such as consecutive 5-FU administration and serial bone marrow transplantation, did not show hematopoietic defect in Dock2-/- mice. Long-term engraftment of transplanted Dock2-/- BM cells was severely impaired in competitive reconstitution. However, this was not intrinsic to HSCs but originated from the defective competition of Dock2-/- lymphoid precursors. These results suggest that Dock2 plays a significant role in BM lymphopoiesis, but is dispensable for HSC engraftment and self-renewal.

Rapid hematopoietic progenitor mobilization by sulfated colominic acid.

Hematopoietic progenitor cells (HPCs) can be mobilized from bone marrow (BM) to the blood by G-CSF. In this process, CXCR4 and CD26 play critical roles. Sulfated colominic acid (SCA) inhibits HIV entry, the step which requires CXCR4 and CD26 as co-receptors. Thus, we hypothesized that SCA would modulate HPC trafficking. We first found that SCA mobilized HPCs rapidly via CD26-independent mechanism. In vitro progenitor migration toward chemokine SDF-1 was significantly enhanced by SCA, and it was completely abrogated by CXCR4 inhibition. This likely originated from the inhibition of CXCR4 down-regulation after interaction with SDF-1. Serum SDF-1 level increased after SCA injection, whereas no change was observed in BM and bone. These results suggest that SCA induces HPC mobilization by modulating CXCR4 function resulting in attraction toward increased SDF-1 in the circulation. Furthermore, we confirmed an additive effect with G-CSF in mobilization. SCA may provide an efficacy in clinical mobilization.

Subdural hematoma associated with dural metastasis of gastric carcinoma: report of two cases.

We treated two cases of a subdural hematoma associated with dural metastasis of gastric cancer, from which both patients died. Case 1: A 60-year-old female patient was hospitalized with a diagnosis of type 4 gastric cancer of the antrum. The patient suddenly collapsed, and, subsequently, left hemiplegia and a depressed level of consciousness were noted. A head computed tomography (CT) scan revealed a subdural hematoma with midline shift. The patient was diagnosed with chronic subdural hematoma and underwent emergency burr hole irrigation. Case 2: A 73-year-old man was diagnosed with type 4 gastric cancer and a total gastrectomy plus splenectomy were performed together with dissection of the N1 and N2 lymph node groups (D2 dissection) in March 2006 (T3, N2, P0, H0, INFgamma, ly3, v0, por2). Postoperative adjuvant chemotherapy was performed using oral TS-1; following tests revealed no recurrence in the abdomen. In December 2006, gingival bleeding was noted with disseminated intravascular coagulation (DIC) and 10 days later, the patient was hospitalized with chief complaints of impaired consciousness and anorexia. CT scan revealed a right subdural hematoma with a midline shift. The patient was diagnosed with chronic subdural hematoma and underwent emergency burr hole irrigation and drainage. The dural biopsy of the two cases revealed adenocarcinoma noted in the dural blood vessel. Special staining revealed CEA-positive adenocarcinoma, and a diagnosis of the dural metastasis of gastric cancer was made. These patients' level of consciousness significantly improved postoperatively. However, DIC developed concurrently, and the patients died on the 13th and 14th postoperative day, respectively.

Chronic lymphoproliferative disorder with regulatory T-cell phenotype.

We report a case of T-cell chronic lymphoproliferative disorder (CLPD) that shows neither features of T-cell prolymphocytic leukemia nor disease progression for more than 34 months. Flow cytometric analyses of the lymphocytes revealed high expression of CD4 and CD25. Up-regulation of Foxp3, a master regulatory gene for developmental differentiation of regulatory T cells (Treg), was confirmed at mRNA and protein levels. To our knowledge, this is the first case of extremely indolent CLPD with Treg phenotype.

[Acute myelogenous leukemia complicated with severe tumor lysis syndrome after a single day of idarubicin/cytarabine chemotherapy].

We describe a patient with acute myelogenous leukemia (AML) who was complicated with severe tumor lysis syndrome (TLS) after a single day of chemotherapy; a previously unreported occurrence. A 48-year-old man was admitted because of fever and bleeding tendency, and was diagnosed as having AML: M1. His white blood cell count (WBC) was 66, 300/microliter with 96% myeloblasts. Chemotherapy with idarubicin and cytarabine was started. On the second day of the therapy, he developed acute renal failure (ARF) and deterioration of disseminated intravascular coagulation (DIC) with a marked reduction in his WBC (8500/microliter). Immediately after a diagnosis of tumor lysis syndrome (TLS), the following chemotherapy was discontinued. Hemodiafiltration and hemofiltration were initiated to treat the ARF, concomitant with anticoagulant and antifibrinolysis therapy for the DIC. Chemotherapy with daunorubicin and cytarabine was resumed on the 12th day, recognizing a regrowth of leukemic cells. Thereafter a complete remission was achieved without recurrence of the TLS.

Hybrid respiration in the denitrifying mitochondria of Fusarium oxysporum.

Induction of the mitochondrial nitrate-respiration (denitrification) system of the fungus Fusarium oxysporum requires the supply of low levels of oxygen (O(2)). Here we show that O(2) and nitrate (NO(3)(-)) respiration function simultaneously in the mitochondria of fungal cells incubated under hypoxic, denitrifying conditions in which both O(2) and NO(3)(-) act as the terminal electron acceptors. The NO(3)(-) and nitrite (NO(2)(-)) reductases involved in fungal denitrification share the mitochondrial respiratory chain with cytochrome oxidase. F. oxysporum cytochrome c(549) can serve as an electron donor for both NO(2)(-) reductase and cytochrome oxidase. We are the first to demonstrate hybrid respiration in respiring eukaryotic mitochondria.