In this work, lanthanum modified lead titanate ceramics are prepared and studied. The giant electrocaloric effect in lanthanum modified lead titanate ceramics is revealed for the first time. Large refrigeration efficiency (27.4) and high adiabatic temperature change (1.67 K) are achieved by indirect analysis. Direct measurements of electrocaloric effect show that reversible adiabatic temperature change is also about 1.67 K, which exceeds many electrocaloric effect values in current direct measured electrocaloric studies. Both theoretical calculated and direct measured electrocaloric effects are in good agreements in high temperatures. Temperature and electric field related energy storage properties are also analyzed, maximum energy-storage density and energy-storage efficiency are about 0.31 J/cm The unique properties and great variety of relaxer ferroelectrics also make them highly attractive for future solid-state refrigeration technologies. During the past decades, intensive research efforts have been conducted to develop solid-state cooling technologies4. The adiabatic temperature change (ΔT) and isothermal entropy change (ΔS) of polar materials are figure of merits of electrocaloric effect (ECE) during application and removal of electric field, which is environment friendly. ECE provides a highly efficient approach to achieve solid-state cooling instead of the existing vapour-compression refrigeration9. Recently, ferroelectrics for future solid-state refrigeration technologies become very hot18. In order to gain higher ΔT, many scholars pay attentions to thin films due to their large breakdown field21. It is a well-known fact that thin films have advantages in small solid state cooling devices, but bulk materials play an important role on larger scale devices, such as: refrigeration23. As a result, ECE of bulk materials are also desired, we should pay more attentions to ECE of bulk materials. Bulk materials including multilayer capacitors, ceramics and single crystals have been reported a lot, such as: 0.9Pb(Mg1/3Nb2/3)O3–0.1PbTiO3 multilayer capacitors16, 0.9PMN-0.1PT single crystal14, Bax1−SrxTiO3 ceramics12. Compared to multilayer capacitors and single crystals, ceramics have the advantages of low-cost and easier fabrications.Since the discovery of ferroelectrics, ferroelectric materials have been exploited in many applications, such as: piezoelectric energy harvesting, optical electronic devices, and Tc) and low dielectric constant25, which make PT based ceramic to be a valuable research object30. In this work, lanthanum modified lead titanate ceramics (Pbx1−Lax)Tix/41−O3  ceramics are prepared and studied. Energy-storage and ECE of PLT ceramics are revealed for the first time. In this work, frequencies and temperatures dependent dielectric permittivity ɛγ and loss tanδ are also investigated to study the relaxer phase transitions and defects related relaxations. Ferroelectric based energy-storage properties are also analyzed. Energy-storage density in this work researches about 0.31 J/cm3, high energy-storage efficiency (91.18%) is also obtained. Large ECE in PLT ceramics is achieved for the first time, maximum value of ΔT is about 1.67 K, and giant refrigeration efficiency is up to 27.4. Additional direct measured electrocaloric effects are analyzed, giant temperature change (1.67 K) is achieved, which indicates that PLT ceramics may be used in future solid-state refrigeration applications.In recent years, lead titanate (PT) based ceramics become one of the most studied and used ferroelectric materials in both scientific and industrial communities due to its high Curie temperature  binder solution and pressed into discs of 10 mm in diameter and 1 mm in thickness uniaxially. These discs were sintered at 1300 °C for 2 h in air. Silver paste was applied on both sides of discs and fired at 650 °C as electrodes for electrical properties measurements. High temperature dielectric behaviours were measured by Agilent E4980A . Low temperature permittivity ɛγ and dielectric loss tanδ of PLT samples were measured using an HP4194A LCR . Complex impedance plots were conducted by Agilent E4980A (0.02–2000 kHz). Ferroelectric hysteresis loops were obtained by a computer-controlled virtual-ground circuit with Precision Premier II Ferroelectric Tester . The direct measurements of ECE were conducted by a customized system: for the direct measurement, ECE change of temperature was monitored by a small thermistor attached to the upper gold electrode of ceramic. In order to reduce the heat exchange with environment, a thermistor and an electric field controlled by a computer were employed to detect the temperature change caused by ECE as the application or withdrawing of an electric field. Also, a high voltage generator controlled by an arbitrary signal generator is used to generator the electric field step signal, which is then applied to the sample. The voltage should be maintained for a few seconds to get into thermal equilibrium with the surrounding. Then the voltage was released immediately. The typical thermal response times along the sample thickness direction is a few milliseconds. Within such a short period, a very fast equilibration of the temperature throughout the whole sample, including the electrodes, attached thermistor and wires, took place, but then the equilibrated sample exchanges the heat on a much longer time scale to the surrounding bath.PLT ceramics were synthesized by a conventional high temperature solid-state fabrication method. Reagent-grade Pbɛγ and loss tanδ for PLT samples are shown in Fig. ɛγ depicts typical relaxer behaviours with a strong dispersion of ɛγ peaks, especially for PLT28 and PLT32 ceramics, Tm (temperature of maximum ɛγ) shift to higher temperatures and maximum ɛγ decrease with increasing frequencies. On the other hand, loss tanδ also exhibits broad peaks clearly, with increasing frequencies, maximum loss tanδ increase as well. Similar results were also reported32. This phenomenon signifies relaxer behaviours33.Temperatures dependent dielectric permittivity ɛγ, at the Curie point Tc of an ideal ferroelectric crystal can be described by the Curie-Weiss law34:C and To are Curie-Weiss constant and Curie-Weiss temperature, respectively. For a first-order phase transition, TC is greater than To, whereas for second-order phase transitions, TC equals To34. In this work, ɛγ of PLT ceramics are analyzed by the Curie-Weiss law, plots of temperatures versus inverse ɛγ (at 10 kHz) are shown in Fig. Tm and To are 385.15 K and 400.00 K, 306.49 and 335.68 K, 207.39 K and 265.75 K, 132.09 and 200.00 K respectively for PLT20, PLT24, PLT28, and PLT32 ceramics. Clearly, both Tm and To decrease sharply with increasing La concentrations.Generally speaking, the maximum value of 35:C1 and γ are assumed to be constant, and ɛm is the maximum permittivity. Parameter γ shows clear information on the character of phase transitions38. Figure ɛγ − 1/ɛm) versus ln(T − Tm) with (at 10 kHz). After fitting the experimental data to the modified Curie-Weiss relationship, we obtain the value of parameter γ = 1.39, 1.47, 1.66, 1.75, respectively for PLT20, PLT24, PLT28 and PLT32 ceramics. Fitting values of γ also support the evidence of relaxer nature.It is well known that dielectric behaviours of relaxer ferroelectrics exhibit to deviate from typical Curie-Weiss behaviour, it can be described by a modified Curie-Weiss relationship7 Hz, 400–800 °C), which are called dielectric relaxation41. In order to give a clear knowledge of high temperature dielectric relaxations, impedance technology is selected as an efficient technique, which has been intensive used in electrical properties of electro-ceramic materials42. The variation of normalized imaginary parts of impedance (Z″/Z″max) are shown in Fig. Z″/Z″max become close gradually in higher frequencies. For PLT24 and PLT28 ceramics, Z″/Z″max can be fitted into 2 separate parts. For a thermally activated relaxation process, relaxation frequency usually follows the Arrhenius law:T, ωo, Ea, kβ are the absolute temperature, characteristic frequency, activation energy and Boltzmann constant, respectively. Relaxation parameter Ea is determined by plotting ln(ω) as a function of the inverse of temperature (1000/T) using Arrhenius law (Zr0.9Ti0.1)O345, and TiO3 ceramics42.From Fig. P-E) hysteresis loops of PLT ceramics under various electric fields . Typical ferroelectric hysteresis loops are observed for PLT20 and PLT24 ceramics, which manifests the ferroelectric phase at room temperatures. For PLT28 and PLT32 ceramics, slim hysteresis loops are achieved indicating the relaxer ferroelectric nature. At room temperature, remnant polarization, and coercive field decrease sharply with increasing La concentrations as shown in Fig. Figure P-E hysteresis loops also reflect energy-storage capacities of dielectric materials. According to the definition of energy-storage density by P-E hysteresis loops, energy-storage density, Jreco, is defined as49:As a well-known fact, Jreco can be obtained by numerical integration of the area between polarization axis and curves of P-E loops easily. In this work, energy-storage density Jreco . From the aspect of practical application, high energy-storage efficiency (η) and low energy-loss density (Jloss) are also significant. Similar to energy-storage density (Jreco), energy-loss density Jloss  (Zr0.65Ti0.35)O3 relaxer ferroelectric thin films54, PbZrO3 antiferroelectric thin films55, HfO2-ZrO2 solid solution thin films56, and HfZrO2 films53, those works report very good results, the energy storage densities are rather large. Compared with bulk materials, our results of Jreco are higher than BaTiO3-SrTiO3 composites57, BaSrTiO3 ceramics58, Sr0.5Ba0.5Nb2O6 glass-ceramics59, and etc. From aspects of applications, the priority among priorities for energy-storage devices are to gain a slim hysteresis loop  or double hysteresis loops, this is the research direction to which we should pay more attentions52.In this work, our results show lower values of ECE of ceramics, ferroelectric properties of PLT20  are showed in Fig. ECE, when electric field increases from E1 to E2, the isothermal entropy change ΔS of an ECE material should be:In order to calculate ECE materials at E1  will affect ECE directly. Assuming the Maxwell relation:Therefore, initial conditions of S and the reversible adiabatic temperature change ΔT are calculated by following relations61:ρ, C, E1 and E2 are mass density, mass heat capacity, initial and final applied electric fields, respectively. Values of (∂P/∂T)E  measured at various temperatures.The corresponding isothermal entropy change ΔS and ΔT calculated at different electric fields are presented in Fig. S and ΔT decrease firstly and then increase with increasing temperatures sharply (<30 kV/cm). On the other hand, ΔS and ΔT increase continuously with increasing temperatures especially for higher fields (>30 kV/cm). Temperatures of maximum ΔT (TECmax) shift toward the higher temperatures with increasing electric fields. Although the ΔT value in this study is lower than PbZr0.95Ti0.05O3 film19, PMN-PT films62. But for bulk materials, our results show higher values of ΔT than Ba(Zr0.2Ti0.8)O3-(Ba0.7Ca0.3)TiO3 ceramics13, and Sr0.75Ba0.25Nb2O6 materials63. More comparison results69 are shown in Table ΔQ and W are isothermal heat and corresponding electrical work per unit volume, and W is equal to ∫EdP71. Our result reveals that value of COP for is about 27.4, which is much higher than previous reports of Pb0.97La0.02(Zr0.75Sn0.18Ti0.07)O3 thick film (COP = 18)70, PbZr0.95Ti0.05O3 film19, P(VDF-TrFE) film72, and PMN-PT (COP = 5.6) films62. Large values of COP suggest the high cooling efficiency, which implies PLT20 ceramics have potential applications in future solid-state refrigeration technologies.In order to give a comparison criterion for electrocaloric refrigeration, refrigeration efficiency is given:E on ECE, electrocaloric coefficient is given:Tmax is the maximum temperature change and ΔEmax is the corresponding electric field change70. Clearly, maximum §max achieved in this work is 0.033 K·cm·kV−1, which is higher than (Pb0.97La0.02)(Zr0.67Sn0.38Ti0.05)O3 thick films (0.030 K·cm·kV−1)73, BaZr0.2Ti0.8O3 ceramic74, and 0.94Bi0.5Na0.5TiO3-0.06KNbO3 ceramic75, 0.7Pb(Mg1/3Nb2/3)O3-0.3PbTiO3 (0.03 K·cm·kV−1)15, 0.9Pb(Mg1/3Nb2/3)O3-0.1PbTiO3 single crystal (0.025 K·cm·kV−1)14, 0.68Pb(Mg1/3Nb2/3)O3-0.32PbTiO3 thin films (0.022 K·cm·kV−1)20.In order to evaluate the quantitative effect of electric field ΔECE in higher temperature region, so the directly measured ΔT (40 kV/cm) are analyzed from 353.15 to 393.15 K as shown in Fig. §max calculated from direct measurements was about 0.050 K·cm·kV−1. Figure T exhibits the maximum value at about 373.5 K. Although the temperature of maximum ΔT from theoretical calculation is higher than that of direct measurement, but they show the similar behaviours and are in good agreements. Compared to previous studies (direct measured ECE) on P(VDF-TrFE-CFE) film77, and P(VDF-TrFE) film78, the ΔT value in this study is smaller by one order of magnitude. But compared to that (directly measured ECE) of bulk materials, our measured results showed higher values than BaHfTiO3 ceramics79, PbMg1/3Nb2/3O3–30PbTiO3 single crystals80, PbZrO3 ceramics81. Figure ECE reported here with some bulk materials91. For ECE researches on ceramics, ΔT (direct measurement) is usually very low, mostly below 1 K. Our research (maximum adiabatic temperature change) shows nearly 1.67 K, both electric field and temperatures dependent ΔT show high ECE values, and it may open more opportunities for practical application in refrigeration devices. The high ΔT value in this study indicates that PLT ceramics have potential applications in future solid-state refrigeration technologies.As PLT20 ceramic exhibits higher ECE74. Some strategies to enhance the ECE applications are possible, such as: maximizing the number of close-energy phases near a critical point in the temperature-composition phase diagram74, combining conventional and inverse caloric responses in a single refrigeration cycle93, introducing extra available degree of freedom like strain via mechanical stress94, and multicaloric effect driven by either single stimulus or multiple stimuli 95. Though promising, in bulk ferroelectrics, ΔT is usually less than a few kelvins, the obtained ΔT is still insufficient for practical application.Ferroelectrics that are characterized by the existence of an electric-field switchable polarization whose appearance is accompanied by structural phase transition have attracted increasing attention for the last 10 years especially in the field of 3 and 91.2%, respectively. Temperatures and electric fields influenced energy-storage properties are analyzed. ECE is studied. High refrigeration efficiency (27.4) and large electrocaloric coefficient are achieved by theoretical calculation, maximum value of ΔT is about 1.67 K. Direct measurements of ECE shows that large ΔT (1.67 K) is obtained, such high value of directly measured ΔT is rare in previous reports.In this work, PLT ceramics are prepared and studied. Relaxer phase transitions and high temperature relaxations are studied. Room temperature energy-storage density and energy-storage efficiency are about 0.31 J/cm


Current status and clinical significance of interventional nephrology has not been reported from Japan.Questionnaires were mailed twice to the directors of all 534 Japanese certificated nephrology training institutions in 2014. The main questions were current performance, categorized annual procedure volume and managers of peritoneal dialysis (PD) access, vascular access (VA) surgery, endovascular intervention, and kidney biopsy. Frequencies of nephrologist involvement between high volume center and low volume center and association between the level of nephrologists’ involvement to each procedure and annual procedure volume were examined.p < 0.01). Nephrologists involvement to VA surgery associated with procedure volume increase in not only VA surgery, but also PD catheter insertion (p < 0.01) and kidney biopsy (p < 0.05). And nephrologists involvement to PD catheter insertion also associated with surgical volume increase in both VA surgery (p < 0.01) and endovascular intervention (p < 0.05).332 (62.2%) institutions answered performance of all procedures and 328 (61.4%) institutions answered all procedure volume. Kidney biopsy, VA surgery, endovascular intervention and PD access surgery were performed by any doctors in 94.2, 96.3, 88.4, and 76.2% and each involvement of nephrologist was 93.9, 54.1, 53.1 and 47.6%, respectively. Cochran–Armitage analyses demonstrated significant increases in all 4 procedure volume with greater management by nephrologists (Main manager of all 4 procedures was nephrologist in Japan. Each procedure volume increased as nephrologists become more involved. Acquisition of one specific procedure by nephrologist associated with increase not only in this specific procedure volume, but also the other procedure volume. Most fellows in non-nephrology internal medicine subspecialties have never considered nephrology as a career choice and the lack of procedural opportunities may be a major reason for not selecting nephrology as a career choice –3. “Don’In 2000, the American Society of Diagnostic and Interventional Nephrology (ASDIN) was established to change the views of nephrologists regarding the practice of nephrology by promoting procedural aspects , 5. RespNo meetings comparable to ASDIN have been organized in Japan. We therefore, established the Japanese Meeting for Interventional Nephrology in 2013, and conducted the first questionnaire survey of all 534 Japanese adult nephrology training institutions to clarify the actual conditions of interventions related to nephrology. This questionnaire asked what types and how many procedures were performed by what medical specialties in each hospital.The results of this survey firstly demonstrated variability between institutions in the performance of procedures related to nephrology by various specialists in Japan, and revealed main managers of this field are nephrologists in Japan and procedure volume increases with increasing levels of involvement by nephrologists.The survey was written jointly by the authors based on their experience in clinical nephrology, performance and assessment of skills in nephrology-related procedures. The general distribution of case numbers for each procedure underwent preliminary field testing with 15 authors, all of whom were members of Japanese nephrology training institutions. Details of this survey were decided by the Japanese Meeting for Interventional Nephrology in July 2013 and February 2014. To improve response rates, we selected a minimal number of procedures for the present questionnaire and excluded placement of hemodialysis catheters from the present survey. The ethics committee for clinical research at Jikei University School of Medicine approved all protocols in this study [Permission no. 26-003 (7508)].Certification in procedures for nephrology fellows in Japan requires that training in these skills be obtained in a nephrology training institution. We thus chose Japanese certificated adult nephrology training institutions as target institutions for this study. Questionnaires were mailed to the directors of all 534 Japanese nephrology training institutions at that time in July 2014. A follow-up questionnaire was posted again in November 2014. All responses were collected by fax.The questionnaire comprised 4 sections, with questions about access procedures for peritoneal dialysis (PD)  , 10; hemStaff of the Japanese Meeting for Interventional Nephrology reviewed the questionnaires and developed the final analysis.We divided institutions into 3 groups: N-institutions, with only nephrologists perform a specific procedure; non-N-institutions, with only non-nephrologists performing the specific procedure; and C-institutions, with both nephrologists and non-nephrologists cooperatively performing the specific procedure.p < 0.05 were considered significant. Cochran–Armitage analysis was used to compare case numbers for procedures among the 3 groups of N-institutions, non-N-institutions and C-institutions.Statistical analyses were performed using JMP version 9.0 . Data are expressed as mean ± standard deviation or numbers and percentages of institutions. Comparisons across groups were performed using the Pearson’s Chi-square test for categorical data. All tests were two-tailed, and values of p < 0.05], there was no significant differences among procedure performance, occupancy rates of nephrologists and procedure volume between Hokkaido region and the other regions.Of the 534 Japanese certificated nephrology training institutions contacted, 236 (44.2%) institutions responded to the initial letters in July 2014, and additional 96 (18.0%) institutions responded to the follow-up letter in November 2014, resulting in responses from a total of 332 (62.2%) institutions. We examined differences of responding rate among each Japanese region and found minimum responding rate was 54.6% (18/33) in Chugoku region without significant differences. Though significant difference of responding rate was seen only in Hokkaido region [Hokkaido region 86.7% (13/15) vs the other regions 61.5% (319/519), n = 214; total responses: 328); 6–10 cases, 20.4% (n = 67); 11–20 cases, 11.3% (n = 37); 21–40 cases, 2.1% (n = 7); and >40 cases, 0.9% (n = 3). The majority of institutions (65.2%) performed <6 cases annually by any doctors.Table n = 285; total responses: 328); 6–10 cases, 11.3% (n = 37); 11–20 cases, 1.2% (n = 4); 21–40 cases, 0.3% (n = 1); and >40 cases, 0.3% (n = 1). The majority of institutions (86.9%) performed <6 cases annually by any doctors.Annual case numbers of PD catheter removal were categorized, and numbers and frequencies were as follows: <6 cases, 86.9% ; 6–10 cases, 0.9% (n = 3); and >10 cases, 0% (n = 0). The majority of institutions (99.1%) performed <6 cases annually by any doctors.Annual case numbers of PD catheter unroofing were categorized, and numbers and frequencies were as follows: <6 cases, 99.1% ; 11–50 cases, 41.5% (n = 136); 51–150 cases, 40.9% (n = 134); 151–300 cases, 6.1% (n = 20); and >300 cases, 2.1% (n = 7). The majority of institutions (82.4%) performed VA surgery 11–150 cases per year by any doctors.Annual case numbers of VA surgery were categorized, and numbers and frequencies were as follows: <11 cases, 9.5% ; 11–50 cases, 36.6% (n = 120); 51–150 cases, 21.6% (n = 71); 151–300 cases, 9.8% (n = 32); and >300 cases, 4.6% (n = 15). The majority of institutions (64.0%) performed endovascular interventions <51 cases annually by any doctors.Annual numbers of endovascular interventions were categorized, and numbers and frequencies were as follows: <11 cases, 27.4% ; 11–50 cases, 54.6% (n = 179); 51–100 cases, 17.7% (n = 58); 101–150 cases, 7.3% (n = 24); and >150 cases, 0.9% (n = 3). Over half of institutions (54.6%) performed 11–50 kidney biopsies annually by any doctors.Annual numbers of kidney biopsies were categorized, and number and frequencies were as follows: <11 cases, 19.5%  vs 52.4% (n = 131) vs 17.6% (n = 44); for PD catheter removal, 28.8% (n = 72) vs 56.0% (n = 140) vs 15.2% (n = 38); for PD catheter unroofing, 35.6% (n = 89) vs 54.0% (n = 135) vs 10.4% (n = 26). Among non-nephrologists, surgeons  and urologists  were involved in higher degree of PD catheter insertions.Among 250 institutions which were offering PD access surgery by any doctors, frequencies of involvement to each PD access surgery by nephrologists, non-nephrologists or their collaboration were as follows: for PD catheter insertion, 30.0% (n = 110) vs 45.8% (n = 145) vs 19.3% ; for endovascular interventions, 39.0% (n = 113) vs 46.9% (n = 136) vs 14.1% ; and for kidney biopsy, 85.1% (n = 263) vs 6.1% (n = 19) vs 8.7%  , urologists , surgeons  and others  performed VA surgery. Radiologists , cardiac surgeons , cardiologists , surgeons  and others  performed endovascular interventions. Urologists  and others  performed kidney biopsies.Among non-nephrologists, cardiac surgeons  institutions could complete all 4 procedures in their own institutions by any doctors. And 22.9% (75/328) institutions could complete all 4 procedures by nephrologists (including nephrologists in cooperative action with non-nephrologists). The resultant 46.0% (151/328) institutions needed other specialists to complete all 4 procedures.n = 306). 92.2% (n = 285) used US-guided procedures alone, 6.1% (n = 19) used both US-guided or open biopsy, and 0.6% (n = 2) used either US- or X-ray-guided procedures. The remaining institutions used open biopsy alone  or some other method details unknown . There were no institutions performing X-ray-guided kidney biopsy alone.In this survey, we also investigated method of kidney biopsy in Japanese certificated adult nephrology training institutions. Kidney biopsy  was performed by ultrasound (US)-guided in 99.0%  and 30.7% (35/114), p < 0.01 for >5 cases and no more than 5 cases per year of PD catheter insertion, 69.5% (66/95) and 42.9% (54/126), p < 0.01 for >50 cases and no more than 50 cases per year of endovascular intervention, 96.5% (191/198) and 73.9% (17/23), p < 0.01 for >10 cases and no more than 10 cases per year of kidney biopsy, respectively.To clarify association between frequencies of nephrologist involvement to each intervention and procedure volume, we compared the frequencies of nephrologist involvement between high volume center and low volume center in each 4 procedure Fig. . FrequenThus, there were significant differences in the frequencies of nephrologist involvement for clinical intervention among the groups categorized by the number of volume of cases. And higher volume centers were offering significant higher frequencies of nephrologist involvement in all 4 procedures.n = 250), VA surgery (n = 316), endovascular intervention (n = 290) and kidney biopsy (n = 309).Next, to examine whether interventional nephrologists have the potential to increase procedure volumes as nephrologists get more deeply involved in each procedure, we analyzed associations between levels of independence from non-nephrologists and annual procedure volume. Frequencies of institutions that performed >5 PD catheter insertions, >50 VA surgery, >50 endovascular interventions and >10 kidney biopsies annually were compared among N-institutions, C-institutions and non-N-institutions Fig. . Institu>5 PD catheter insertions were performed in 68.0% of N-institutions (51/75), 54.6% of C-institutions (24/44) and 29.0% of non-N-institutions (38/131) among 250 institutions that were performing PD access procedures. Over 50 VA surgery were performed annually in 66.4% of N-institutions (73/110), 52.5% of C-institutions (32/61) and 38.6% of non-N-institutions (56/145) among 316 institutions that were performing VA surgery. Over 50 endovascular interventions were performed annually in 57.5% of N-institutions (65/113), 36.6% of C-institutions (15/41) and 27.9% of non-N-institutions (38/136) among 290 institutions that were performing endovascular interventions. More than 10 kidney biopsies annually were performed in 85.1% of institutions (263/309), comprising 87.8% of N-institutions (231/263), 88.9% of C-institutions (24/27) and 42.1% of non-N-institutions (8/19).n = 250 (p < 0.0001); >50 VA surgery/year, n = 316 (p = 0.0055); >50 endovascular interventions/year, n = 290 (p = 0.0083); and >10 kidney biopsies/year, n = 309 (p = 0.0001). These analysis suggested procedure volume may increase with the levels of independence from non-nephrologists.Cochran–Armitage analysis showed annual numbers of PD catheter insertions, VA surgery, endovascular interventions and kidney biopsies significantly increased in a stepwise manner from non-N-institutions to C-institutions to N-institutions as nephrologist involved more as follows: >5 PD catheter insertions/year, p = 0.0138; for >50 VA surgery, 71.9 vs 40.9%, p < 0.0001; for >50 endovascular interventions, 47.7 vs 36.6%, p = 0.0999; and for >10 kidney biopsy, 95.3 vs 81.7%, p = 0.0011, respectively.When nephrologists acquire a specific procedure, whether it has positive effects on the other procedure volume is worth examination. In this analysis, we included only 221 institutions which completed all 4 procedures in their own institutions  of institutions could complete all 4 procedures by nephrologists (in cooperative action with non-nephrologists), indicating that majority of nephrology fellows could rarely train all 4 procedures even in Japanese certificated nephrology training institutions.Nephrologists’ participation to VA surgery did not only associate with surgical volume increase in VA surgery, but also associated with procedure volume increase for PD catheter insertion and kidney biopsy. And nephrologists’ participation to PD catheter insertion associated with surgical volume increase in not only PD catheter insertion, but also both VA surgery and endovascular intervention. Thus, acquisition of one interventional procedure by nephrologists may enhance skill acquisition and volume increase in the other procedures. This chain reaction will activate interventional nephrology in each institution and may promote fellows to select nephrology as a carrier choice.Japanese certificated nephrology training institutions were offering VA surgery, kidney biopsy, and endovascular intervention with higher performance rates of over 88.4% by any doctors. On the other hand, performance rate of PD access surgery by any doctors was relatively lower (76.2%) and non-nephrologists managed over half of PD access surgery, revealing many young nephrologists could not train PD access surgery even in certificated nephrology training institutions.Present study revealed main manager of VA surgery was nephrologist, followed by cardiac surgeon, urologists and surgeon in Japan. 54.1% of nephrology training institutions were performing VA surgery by nephrologists alone (34.8%) or with the help of a non-nephrologist (19.3%), relatively high rates compared to reports from both the United States and Europe.Arteriovenous fistula (AVF) established by nephrologists represented only 35% in Europe and under 11% in the United States . BeatharTo date, HD access surgery by nephrologists and its main effects have also been reported , 15–17. In this survey, 76.2% institutions answered they could offer PD access surgery by any doctors. But over 10 cases/annually of PD catheter insertion were performed by only 14.3% institutions in Japan, relative fewer than 23% in USA (23%) . ContrasThis poor surgical volume of PD catheter insertion by non-nephrologists is a characteristic in Japan and may cause the poor penetration rate of PD in Japan , 24. So,Several reports described PD catheter insertion by nephrologists and the effects on clinical nephrology, such as utilization of PD –28 or prPerformance rates for kidney biopsy performed by any type of doctor were 94.2% (309/328 institutions) in present study, broadly comparable to the high rate in the United States (99%) . AdditioThis study involved several limitations. First, participation was voluntary, with 37.8% of directors were not responding. Second, we could not analyze details about which kinds of VA surgery and endovascular intervention were performed in responding institutions. Third, no attempt was made to verify the accuracy of the responses. In this regard, other surveys conducted in this field were similar to the other survey , 31. ForThis survey offers the first comprehensive picture of interventional nephrology in Japanese nephrology training institutions. Main manager of all 4 procedures was nephrologist in Japan. All procedure volume increased as nephrologists become more involved. Acquisition of one specific procedure by nephrologist associated with increase in not only this specific procedure volume, but also the other procedure volume.


Upon addition of molecular probes of Victoria blue B (VBB), the surface-enhanced Raman scattering (SERS) peak linearly decreased at 1612 cm−1 with the Hg(II) concentration increasing in the range of 0.013–0.5 μmol/L. With its good selectivity and good accuracy, the RRS method is expected to be a promising candidate for determining mercury ions in water samples.Mercury (Hg) is a heavy metal pollutant, there is an urgent need to develop simple and sensitive methods for Hg(II) in water. In this article, a simple and sensitive resonance Rayleigh scattering (RRS) method was developed for determination of 0.008–1.33 µmol/L Hg, with a detection limit of 0.003 μmol/L, based on the Hg(II) regulation of gold nanoenzyme catalysis on the HAuCl They activity ,9,10,11.activity ,13. Goldity [2O2 . Combini2+ becomes very important. At present, several methods have been reported to determine Hg2+, such as atomic fluorescence spectroscopy (AFS), atomic absorption spectrometry (AAS), and inductively coupled plasma mass spectrometry (ICP-MS) [2+ detection, however, these methods have a number of weaknesses such as expensive instruments, long analysis period, need for professional operators, and so on. It is of great significance to develop a simple, rapid, and low-cost method for mercury detection. In recent years, some new methods for the detection of Hg2+ have been reported such as aptamer and functionalized gold nanoparticle biosensors [4-ascorbic acid reactions [4-H2O2, while Hg2+ exhibits strong inhibition on this nanocatalytic reaction, and the nanoreaction product of formed AuNPs demonstrated RRS effect and SERS activity in the presence of Victoria blue B (VBB) probes. Thus, two simple, rapid, low cost, high sensitivity, and good selectivity RRS and SERS methods were established for detection of Hg2+.Mercury is a highly toxic heavy metal element which is very widely distributed in nature. Therefore, the monitoring of Hg(ICP-MS) ,15,16. Aosensors ,18,19. Rosensors ,22,23,24osensors ,29,30,31osensors  and HAuC2O2-HAuCl4 was relatively slow. While nanocatalyst of AuNP was added, a large number of HAuCl4 adsorbed on the AuNP surface, and H2O2 was also adsorbed on the surface that there had many free electrons, which can enhance the redox electron-transfer between H2O2-HAuCl4. Thus, it enhanced the reduction of Au3+ to form elemental gold, and led to the gold nanoparticles forming greatly. The formed gold nanoparticles have strong SERS and RRS signal. Therefore, the nanogold catalytic reaction can be used to establish SERS and RRS detection methods for AuNPs (3+ + AuNP = AuNP-Au3+ (adsorb)Au2O2 + AuNP-Au3+ = Au3+-AuNP-H2O2 (adsorb)H3+-AuNP-H2O2 = Au+-AuNP-H2O2 + O2Au+-AuNP-H2O2 = AuNP + Au + O2Aun = AuNPsnAu = (Au)Nano-catalysis is an important way to amplify analysis signal. It has significance in developing a new nano-catalysis reaction. From the experiment, we found that, in hydrochloric acid medium, the reaction of Hor AuNPs .Au3+ + 2+ would form in the presence of Hg2+. Similarly, in the analytical system, the formed AuNP-HgCl42− can adsorb on the surface of AuNP, which greatly inhibits the red electron-transfer, and decreases the AuNP catalytic reaction. With the Hg2+ connection increased, the catalytic reaction rate slowed, and the RRS and SERS intensity decreased   .b exhibited a surface plasmon resonance (SPR) peak at 510 nm, it catalyzed H2O2 reduced HAuCl4 to form gold nanoparticles with a SPR peak at 570 nm. With AuNPb increased, the color changed from colorless to red  (n = 5) for the water samples after adding a certain amount of Hg2+ were determined. The recovery was in the range of 97–102%, and the relative standard deviation (RSD) was in the range of 4.5–5.2%. This indicated the method is accuracy and reliable.Three water samples—including tap, river, and pond—were collected from Guangxi Normal University, and filtrated according to the reference . The folry (AAS) . The aveA DXR smart Raman spectrophotometer  with a power of 2.5 mW, laser wavelength of 633 nm, and slit width of 50 mm; a Cary Eclipse fluorescence spectrophotometer ; a model of TU-1901 double-beam UV—visible spectrophotometer ; a C-MAG HS7 magnetic stirrer with heating , and a constant temperature magnetic stirrer were used. A FEI Quanta 200 FEG Field emission scanning electron microscope  was recorded the SEM of nanoparticles, the sample preparation was as follows: put 1.5 mL of the reaction solution of the procedure into a 2 mL centrifuge tube and centrifuge for 20 min (150 × 100 r/min), discard the supernatant and add water to 1.5 mL, then dispers for 30 min with ultrasonication. After centrifuging again, add 1 mL water, and disperse for 30 min. Place 2 µL of the sample solution by pipette dripping on silicon slices and dry naturally for use.4·4H2O , 1% trisodium citrate, 10 mmol/L sodium borohydride, 0.01 mol/L HCl solution and 0.3% H2O2 (0.1 mol/L) were prepared. Victoria blue B solution: 0.025 g VBB was dissolved in 5.0 mL ethyl alcohol before diluting to 50.0 mL with water, and the concentration was 1.0 × 10−3 mol/L. It was stepwise diluted to 1.0 × 10−5 mol/L before use. A 5.23 × 10−5 mol/L RhS and 5.0 × 10−5 mol/L safranine T solution were prepared. Preparation of gold nano sol (AuNPb): At room temperature, 40 mL of water was added into a conical flask, 0.5 mL 1.0% HAuCl4 and 3.5 mL 1.0% trisodium citrate were added into the flask in order by stirring. Then, 4.0 mL 0.05% NaBH4 was added slowly. The mixture was stirred for 10 min, to obtain a concentration of 58 μg/mL 5 nm AuNPb . Preparation of AuNPc: 50 mL of water was added into a conical flask, heated to boiling. Then, 0.5 mL 1% HAuCl4 and 3.5 mL 1% trisodium citrate were added rapidly into the boiling water successively. After boiling for 10 min while stirring, the color went from colorless to wine red. The mixture was stirred continuously to room temperature, and then diluted to 50.0 mL to obtain about 10 nm of AuNPc at a concentration of 58 μg/mL. Preparation of silver nanosol (AgNPs): 40 mL of water was added into a conical flask, 385 μL 2.4 × 10−2 mol/L AgNO3 and 3.5 mL 10 g/L trisodium citrate were added into the flask in order with stirring. Then, 4.0 mL 0.5 mg/mL NaBH4 was added slowly. The color turned from pale yellow to deep yellow. The mixture was stirred for 10 min, diluted to 50.0 mL to obtain a concentration of 20.0 μg/mL AgNPs about 9 nm in size, and stored at 4 °C. All reagents were of analytical grade and the water was doubly distilled.A 1% HAuClb, and a certain amount of Hg2+ were added and diluted to 200 µL, then left to rest for about 20 min. 80 µL 0.1% HAuCl4 (84 μmol/L), 100 μL 0.01 mol/L HCl, and 50 µL 0.3% (0.1 mol/L) H2O2 were added sequentially, diluted to 1.5 mL and mixed well. Then the mixture heated for 15 min in a 60 °C water bath and cooled with tap water. The RRS spectra were recorded by means of synchronous scanning excited wavelength exλ and emission wavelength emλ (exλ − emλ = Δλ = 0) on fluorescence spectrophotometer, with a photomultiplier tube (PMT) voltage of 400 v, and both excited and emission slit widths were 5 nm, emission filter = 1% T attenuator. The reaction solution RRS intensity at 370 nm 370 nmI and the blank solution without Hg2+ (370 nmI)0 were recorded. The value of 370 nmΔI = 370 nmI − 370 nm)0 and the blank value (0I) without Hg2+ were recorded. The value of ΔI = I − 0I was calculated.In a 5 mL marked test tube, 100 µL 0.57 µg/mL AuNP4-H2O2 is slow. Upon addition of nanocatalyst of AuNPb, the nanoreaction enhanced greatly to form gold nanoparticles with strong RRS, SERS and SPR absorption effects. When analyte of Hg2+ was added, the catalysis was greatly inhibited and the SPR effect decreased. Thus, two new RRS and SERS methods were developed for determination of Hg2+, with simplicity, high sensitivity, and good selectivity.The gold nanoreaction of HAuCl


Bacillus subtilis-infecting bacteriophages on soybean fermentation is poorly-studied. In this study, we isolated a B. subtilis-infecting bacteriophage BSP10 from Meju (a brick of dried fermented soybean) and further characterized it. This Myoviridae family bacteriophage exhibited a narrow host range against B. subtilis strains . The genome of bacteriophage BSP10 is 153,767 bp long with 236 open reading frames and 5 tRNAs. Comparative genomics  and phylogenetic analysis strongly suggest its incorporation as a new species in the Nit1virus genus. Furthermore, bacteriophage BSP10 was efficient in the growth inhibition of B. subtilis ATCC 15245 in liquid culture and in Cheonggukjang (a soybean fermented food) fermentation. Artificial contamination of as low as 102 PFU/g of bacteriophage BSP10 during Cheonggukjang fermentation significantly reduced bacterial numbers by up to 112 fold in comparison to the control (no bacteriophage). Moreover, for the first time, we experimentally proved that B. subtilis-infecting bacteriophage greatly enhanced poly-γ-glutamic acid degradation during soybean fermentation, which is likely to negatively affect the functionalities of Cheonggukjang.While the harmful effects of lactic acid bacterial bacteriophages in the dairy industry are well-established, the importance of  Bacillus subtilis is a Gram-positive, endospore-forming bacterium commonly found in soil and the human gut  × 100). EOP was classified into three categories, high (>50%), medium (10–50%), and low (<10%).Host range analysis was performed initially by dotting 5 µL of PEG-precipitated bacteriophage sample  and incubated at 37 °C. When the optical density at 600 nm (OD600) was reached at 0.3 (approximately 1 × 107 CFU/mL), the bacteriophage was added at MOI of 0.01 and incubated for 5 min, followed by centrifugation at 26,200× g for 10 min. The pellet was resuspended in the same volume of fresh TA medium. Then 1 mL of this mixture was aliquoted into micro-centrifuge tubes (1.5 mL) followed by incubation at 37 °C with shaking (160 rpm). At every 10 min, one tube was taken out from the incubator and centrifuged at 12,300× g for 3 min. The supernatant was then collected and used for PFU counting by using TA soft agar.A one-step growth assay was performed as previously described with some modifications . To summGenomic DNA was extracted using PEG precipitated bacteriophage stock (filter-sterilized) and an ExgeneTM Cell SV DNA purification kit  after DNase and SDS treatment. Whole genome sequencing was done by Macrogen Inc., Geumcheon-gu, Korea using a MiSeq sequencing system . Raw reads were trimmed and de novo assembled using A5-miseq pipeline with approximately 575× depth of coverage among the 297,028 reads.Bacillus bacteriophage BSP10 was deposited in GenBank (Accession number MF422185).Open reading frames (ORF) were predicted using RAST 2.0 , GlimmerBacillus-infecting bacteriophages  was calculated using the BLASTN algorithm and the following equation: ([query coverage × identity]/100) /100) . Dot ploty]/100) . Progresty]/100)  and Corety]/100)  were useThe whole genome sequence of bacteriophage BSP10 was analyzed using the CLuster Analysis of Sequences (CLANS) software package  and GegeB. subtilis ATCC 15245 was added to 25 mL fresh TSB media and incubated at 37 °C with shaking (160 rpm). Bacteriophage BSP10 was added at a multiplicity of infection (MOI) of 1 and 0.1 when OD600 was approximately 0.3. Bacterial growth  and plaque forming units (PFU) were monitored at the designated time points for 72 h. The experiment was performed for three times and the values were represented as the mean ± Standard Deviation (SD).Liquid culture inhibition of bacteriophage BSP10 was performed as described by Shin, Bandara, Shin, Ryu, and Kim , with soB. subtilis ATCC 15245 growth inhibition by bacteriophage BSP10 was evaluated by modifying the method used for B. cereus inhibition by bacteriophage in Cheonggukjang  Glycerol in TAE buffer) and allowed to electrophorese in 1.5% agarose gel in TAE buffer  at 50 V for 90 min. γ-PGA was visualized by staining the gel with 0.5% methylene blue in 3% acetic acid for 30 min followed by destaining in H2O.Extracted γ-PGA and a standard poly-L-γ-glutamic acid sodium salt  were electrophoresed following the method of Mamberti, et al. . BrieflyB. subtilis ATCC 15245 were grown in TSB media at 37 °C in shaking condition (160 rpm) for 24 h followed by centrifugation at 26,200× g for 10 min. The cytosolic contents of the bacteria were prepared after lysis of the control bacteria (without bacteriophage infection) by lysozyme (at 37 °C for 1 h) and sonication  for 10 min, followed by centrifugation at 12,300× g for 5 min to remove the cell debris. The bacteriophage-infected bacterial lysate was prepared by the infection of bacteriophage BSP10 (MOI 1) to B. subtilis ATCC 15245 culture in TSB media followed by incubation at 37 °C for 24 h in shaking condition (160 RPM), and the supernatant was collected after centrifugation at 26,200× g for 10 min.In order to find out the source of the γ-PGA hydrolase enzyme, the bacterial culture supernatant (without bacteriophage infection), cytosolic contents of the bacteria (without bacteriophage infection), and bacteriophage-infected bacterial lysate were used for γ-PGA hydrolase enzyme assay. For bacterial culture supernatant preparation, γ-PGA hydrolase activity was assayed following the method of Kimura and Itoh . BrieflyMeju, by using B. subtilis ATCC 15245 as a host. The bacteriophage showed a clear plaque in TA soft agar after 12 h of incubation at 37 °C. Bacteriophage BSP10 was found to be a member of the Myoviridae family with an isometric head  and a contractile tail   . Similar, n = 6) A,B.B. subtilis (17 out of 52 strains) (B. subtilis), EOP was classified as high , medium , and low . Other tested bacterial strains  were not infected by bacteriophage BSP10.Bacteriophage BSP10 showed a narrow host range against strains) . DependiB. subtilis ATCC 15245  was observed in the region between nucleotides 78,776–84,660 (5885 bp) using Geneious software, suggesting the presence of a long terminal repeat similar to phiNIT1 and Grass in the us genus ,35,36. ANit1virus genus, phiNIT1, Grass, and SPG24, respectively, whereas no significant similarity was observed with the representative members of the genus Agatevirus, B4virus, Bastillevirus, Bc431virus, Cp51virus, or Wphvirus  with the ICTV-classified members of Wphvirus . It alsoWphvirus  in the CBacillus bacteriophages using CLANS, it was clustered with the members of previously-proposed Bastille-like group bacteriophages [Nit1virus genus  and 48 (20.3%) of ORFs had a high resemblance (≥99%) with bacteriophage phiNIT1 and Grass, respectively. On the other hand, 25 and 32 ORFs of bacteriophage BSP10 were found to be unique (spanned throughout the genome) compared with phiNIT1 and Grass, respectively, while 16 ORFs of phiNIT1 and 33 ORFs of Grass are absent in bacteriophage BSP10 genome .The bacteriophage BSP10 genome contained a poly-gamma-glutamate hydrolase (γ-PGA hydrolase) homolog , which might function to breakdown γ-PGA, which is responsible for the sticky appearance as well as the functional properties of soybean-based fermented foods. It showed high similarities (BLASTP) with those of Grass  and phiNIT1 .B. subtilis ATCC 15245 in liquid culture. Clear lysis was observed 3 h postinfection for MOI 1 (OD600 of 0.00) and 4 h postinfection for MOI 0.1 (0.00), whereas the OD600 of the control (no bacteriophage addition) was 1.32 ± 0.10 after 5 h (corresponding to 3 h postinfection in the bacteriophage-treated sample) and 1.42 ± 0.04 after 6 h (corresponding to 4 h postinfection) of bacterial inoculation (Bacteriophage BSP10 effectively inhibited the growth of culation A.10 CFU/mL) and 3.1 ± 1.3 log10 CFU/mL (4.0 ± 0.6 and 3.9 ± 0.6 log10 CFU/mL) for the control (without bacteriophage) and the MOI 0.1 treated set, respectively. Interestingly, no bacterial regrowth was detected for MOI 1 after 24, 48, and 72 h. With regard to bacteriophage growth, PFU counts rapidly increased, and after 3 h postinfection reached 10.0 ± 0.2 log10 PFU/mL (MOI 1) and 9.8 ± 0.1 log10 PFU/mL (MOI 0.1) (After 24 h (48 and 72 h) incubation, the bacterial CFU counts were 8.6 ± 0.2 (8.0 ± 0.4 and 7.7 ± 0.1 logMOI 0.1) B. No sigCheonggukjang was prepared to check the effect of bacteriophage BSP10 on B. subtilis growth inhibition during soybean fermentation. Bacteriophage BSP10 was artificially contaminated at 102, 104, and 106 PFU/g in autoclaved soybean which contained 104 CFU/g (which is the standard concentration used in factories) of B. subtilis ATCC 15245. After 24 h incubation, statistically significant differences (p < 0.05) were observed among all the treatments and the bacterial numbers were 8.85 ± 0.06, 8.34 ± 0.10, and 7.11 ± 0.13 log10 CFU/g for 102, 104, and 106 PFU/g addition of bacteriophage BSP10, respectively, as compared to 9.16 ± 0.04 log10 CFU/g of the control (no bacteriophage)  and 106 PFU/g (8.29 ± 0.21 log10 CFU/g) of bacteriophage BSP10-treated sets as compared to control (9.51 ± 0.09 log10 CFU/g) and 102 PFU/g treated sets (9.31 ± 0.16 log10 CFU/g) (6 PFU/g treated flasks). No statistically significant difference in the bacterial numbers was observed between control (9.78 ± 0.09 log10 CFU/g) and 102 PFU/g bacteriophage BSP10-treated set (9.63 ± 0.18 log10 CFU/g), whereas the numbers were significantly different in 104 PFU/g (9.10 ± 0.14 log10 CFU/g) and 106 PFU/g (8.26 ± 0.23 log10 CFU/g) treated sets as compared to the control (The fast-fermented soybean product iophage) A. B. sub0 CFU/g) A. Simila control A.10 PFU/g for an initial bacteriophage contamination level of 102, 104, and 106 PFU/g, respectively   B.Cheonggukjang, γ-PGA was extracted from the bacteriophage-treated and -untreated Cheonggukjang samples, and electrophoresed in 1.5% agarose gel. The fragmentation of γ-PGA (low molecular weight) was observed in the bacteriophage-treated Cheonggukjang samples, while a clear band of high molecular weight γ-PGA was noticed for the bacteriophage-untreated sample  and the bacterial cytosolic content. On the other hand, the degraded γ-PGA was observed in the bacteriophage-treated bacterial lysate B.B. subtilis is used in different soybean-based fermented food industries, and is responsible for the fermentation and qualities of the final products [B. subtilis-infecting bacteriophage might create a serious problem, such as the delay of fermentation process leading to economic losses and changes in the functional properties of the final product. The knowledge about the prevalence, diversity, classification, and properties of bacteriophages is essential for the fermentation industry to develop antiphage strategies and minimize the risk of bacteriophage infections [B. subtilis-infecting bacteriophage whole genome sequences (24 sequences including bacteriophage BSP10) are available in the NCBI database until now.As previously acknowledged, products ,5. Consifections . The avafections ,10. In cMyoviridae family bacteriophage, BSP10 from a dried soybean-fermented brick, Meju   . Moreoveus genus . The aboNit1virus. The whole genome sequence comparison (BLASTN) of bacteriophage BSP10 with phiNIT1 (87.30%), Grass (84.48%), and SPG24 (92.12%)  , suggestB. subtilis-infecting bacteriophages    and one Previously, it was proposed that the genes within the terminal redundancy region of the bacteriophage are the first to be expressed in the infected cell . It was Nit1virus bacteriophages were found in a wide range of environments  and different geographical locations  [Nit1virus bacteriophages, it can be postulated that these widespread bacteriophages might be a threat to soybean-based fermented food industries and other industries where B. subtilis is used as a fermenting bacteria., Korea) ,36. ConsNit1virus genus are not available), possessing a long, contractile tail and an isometric head; baseplates moved upwards and the tail tube was extended from the baseplates and tail sheath  [The latency period (40 min) and burst size of bacteriophage BSP10 (approximately 185)  were foutely 50) , SPO1 (ltely 50) , and vB_tely 50) . The distely 50) ,43,44.Listeria monocytogenes, Escherichia coli, Salmonella spp., Campylobacter spp., Enterobacter sakazakii, and B. cereus [B. subtilis ATCC 15245 in liquid culture  after 24 h  collected after cheese fermentation [High bacteriophage titer was observed during the liquid culture inhibition B and Cheentation B, possibentation  and/or centation .B. subtilis) fermentation [B. subtilis-infecting bacteriophage directly affects γ-PGA qualities during fermentation in the soybean medium has not been experimentally proven.γ-PGA, a sticky material, is produced in soybean-based fermented foods by bacterial (especially entation . It has entation ,54,55. Oentation . A numbeentation ,42. HoweCheonggukjang fermentation (In this study, we found the fragmentation of γ-PGA in the bacteriophage BSP10-treated samples during entation A. Furtheentation B. Taken B. subtilis-infecting bacteriophage in Cheonggukjang fermentation not only retards bacterial growth but also significantly lowers product qualities.The production of a low molecular weight γ-PGA in bacteriophage-treated samples could negatively affect the functionalities of γ-PGA. Taken together with the reports on the detrimental effects of lactic acid bacterial bacteriophage in dairy environments, such as reduced acidification of milk in presence of lactic acid bacterial bacteriophage ,56, our Myoviridae family B. subtilis-infecting bacteriophage BSP10 was isolated from Meju and found to be a new species in the Nit1virus genus. We proved that bacteriophage BSP10 infection in Cheonggukjang fermentation not only affected bacterial growth but also decreased the quality of the soybean-based fermented food by degrading the functional compound, γ-PGA. This study will provide useful information for future research on the host–bacteriophage interaction and help to establish a clear classification scheme of B. subtilis-infecting bacteriophage.A


Although several methods to create an effective counter traction for safer endoscopic submucosal dissection (ESD) have been reported, these methods do not overcome problems regarding delivery and ease of use. This randomized prospective study assessed the usefulness of ring-shaped thread counter traction, which not only allowed the safer colorectal ESD but also the easiest and lower cost counter traction without any special devices.n = 22) and the ring-shaped thread counter traction ESD group (RE) (n = 21). The ring-shaped thread was hooked and lifted up to the contralateral mucosa with a hemoclip. The primary outcome was the dissected area per minute during ESD (cm2/min) (UMIN000020160).Forty-five patients diagnosed with colorectal lateral spreading tumors over 20 mm were allocated to the conventional ESD group (CE) (P = 0.001). For the dissected areas per minute (cm2/min), there was a significant difference, with 0.125 (0.1–0.18) versus 0.235 (0.16–0.36) min (P = 0.003) for the CE and RE groups, respectively. There were 1 cases of perforation during ESD in the CE compared to 0 for the RE, and this was no significantly different (P = 0.31). The procedure time of producing and setting the ring-shaped thread counter traction was approximately 1.80 (0.80–3.30) min only.There were significant differences in the dissection time (min), with 130.0 (56.0–240.0) versus 80 (35.0–130.0) min for the CE and RE groups, respectively  contains supplementary material, which is available to authorized users. Inclusion criteria were colorectal tumors with diameters over 20 mm in diameter [lateral spreading tumors (LSTs)] that were classified in the JSCCR (Japanese Society for Cancer of the Colon and Rectum) as follows: LST-G (granular type) and LST-NG (non-granular type). Exclusion criteria were definitely protruded-type tumors, such as (0–1), and a diagnosis of advanced colorectal (SM massive) cancer with image-enhanced magnified endoscopic examination. Tumor diameters were measured using measuring forceps. A total of two patients were excluded due to diagnoses of advanced colorectal (SM massive) cancer with image-enhanced magnified endoscopic examination. Finally, a total of 43 of 45 patients were included with colorectal tumors larger than 20 mm.n = 22) were allocated to the conventional ESD group (CE), and even-numbered patients (n = 21) were allocated to the ring-shaped thread counter traction ESD group (RE)  of 1.5, and heparin was discontinued 3 h before ESD. Heparin was resumed 3 h after ESD, and anticoagulants resumed the following day. Patients taking antiplatelet drugs consulted a cardiologist. Patients taking ticlopidine hydrochloride, clopidogrel sulfate or aspirin were changed to cilostazol 3 days before ESD, and they discontinued cilostazol the day of ESD. All antiplatelet drugs were resumed the day after ESD.Colon pretreatment consisted of ingestion of 2 L of a polyethylene glycol solution . All patients were discharged 6 days after ESD.2, adjusting the strength of counter traction with the ring-shaped thread was possible n) (min) was defined as the dissecting time of submucosal layer only, except for the other ESD procedure times, which were measured by a nurse with a stopwatch who measured only the dissecting time when the ESD operator used electric knives during the ESD.In both groups, the dissection time (DTn) (cm) and longer axis (abbreviated Ln) of the ellipsoid resected specimen were measured after ESD. The ellipsoid resected area (abbreviated An) was defined as the area calculated by the following formula:The shorter axis  according to the Declaration of Helsinki. The patients provided verbal and written informed consent.University Hospital Medical Information Network (UMIN000020160) following the CONSORT check list.2/min). Based on the results of pilot studies (DAn) (cm2/min), the sample size was calculated by performing a statistical analysis using GraphPad Prism with the sample size (23 patients) using the effective size of 0.80 (standard deviation 0.0351) . (http://biostat.mc.vanderbilt.edu/wiki/Main/PowerSampleSize).After we conducted the pilot study for 8 patients in the CE and 8 patients in the RE , we found significant differences between the two groups in the dissection speed .Incisional knife: IT knife 2 .Hemostatic forceps: Coagrasper .Injection needle: NM-4U .®, Johnson & Johnson K.K., Tokyo, Japan) and 10% glycerin (mixing ratio 1:1).Local injection solution: hyaluronic acid .n) (cm2/min) (n = 1–43).The primary outcome was the dissected area per minute during ESD .Data between groups were analyzed using Fisher’s exact test or the P = 0.23 and 0.48). For the location of lesions, in the CE group (22 lesions), we found 3 lesions in the cecum, 13 in the colon and 6 in the rectum. In the RE groups (21 lesions), 5 lesions were in the cecum, 9 in the colon and 7 in the rectum. There was no significant difference in the location of lesions (P = 0.15).Of the characteristic backgrounds in the CE and RE groups, there were no significant differences in age and gender (P = 0.13) (Table Macroscopic findings of lesions (JSCCR classification) revealed that LST-G (granular type) accounted for 12 and 14 lesions, and LST-NG (non-granular type) for 10 and 7 lesions in the CE and RE, respectively. There was no significant difference in the macroscopic findings (n) (cm2), with 27.6 (10.3–50.20) versus 27.3 (11.0–49.9) cm2 for the CE and RE groups, respectively (P = 0.54) (Table n) (min): 130.0 (56.0–240.0) versus 80 (35.0–130.0) (min) for the CE and RE groups, respectively (P = 0.001)  (cm2/min), there was a significant difference, with 0.125 (0.1–0.18) versus 0.235 (0.16–0.36) (cm2/min) (P = 0.003) . There was 1 case of perforation during ESD in the CE groups compared to 0 in the RE group, which was also of no significant difference (P = 0.31) (Table There were no significant differences in the approximate ellipsoid resected area (An) cm, with 277.3 11.0–9.9 cm2 f7.3 11.0–9.9 cm2 f7.3 11.0–9.9 cm2 fP = 0.42) (Table The procedure time of making and setting ring-shaped thread counter traction to the lesions was approximately only 1.80 (0.80–3.30) min. Histopathological examinations revealed no significant differences in the proportions of adenocarcinoma, adenomas with high-grade atypia and adenomas with low-grade atypia P = 0.4 Below is the link to the electronic supplementary material.


To evaluate if the new Discovery Molecular Insights (DMI) PET/CT scanner provides equivalent results compared to the standard of care PET/CT scanners (GE Discovery 600 or GE Discovery 690) used in our clinic and to explore any possible differences in semi-quantitative measurements.18F-FDG injection and two scans: the clinical standard PET/CT followed immediately by the DMI PET/CT scan. We measured SUVmax and SUVmean of different background organs and up to four lesions per patient from data acquired using both scanners.The local Institutional Review Board approved the protocol and written informed consent was obtained from each patient. Between September and November 2016, 50 patients underwent a single 18F-FDG uptake. The SUVmax values for all 107 lesions ranged 1.2 to 14.6 (mean ± SD: 2.8 ± 2.8), higher on DMI PET/CT compared with standard of care PET/CT. The mean lesion:aortic arch SUVmax ratio and mean lesion:liver SUVmax ratio were 0.2–15.2 (mean ± SD: 3.2 ± 2.6) and 0.2–8.5 (mean ± SD: 1.9 ± 1.4) respectively, higher on DMI PET/CT than standard PET/CT. These differences were statistically significant  and not correlated to the delay in acquisition of DMI PET data (P < 0.0001).DMI PET/CT identified all the 107 lesions detected by standard PET/CT scanners, as well as additional 37 areas of focal increased Our study shows high performance of the new DMI PET/CT scanner. This may have a significant role in diagnosing and staging disease, as well as for assessing and monitoring responses to therapies. Positron emission tomography (PET) plays a significant role in the diagnostic workup of cancer patients, evaluating various biological processes, including tumor cells metabolism. Indeed, PET has increasingly been recognized as a powerful tool for diagnosis, prognosis and determination of response to therapy in oncology .18F-FDG), an analogue of glucose, is the most used PET radiotracer in clinical practice, providing information based on the increased glucose uptake and glycolysis of cancer cells fluoro-D-glucose , time between injection and scan (uptake time) as well as data reconstruction methods .Bland-Altman plot was used to evaluate the degree of agreement between different scanners. We used Partial Correlation and Linear Regression analysis to evaluate the relationship between different SUV18F FDG PET for Oncologic Conditions from the Centers for Medicare & Medicaid Services , performed on each of the normal tissues matched per patient are presented in On a per patient analysis, 12 of the 50 patients had no abnormal uptake on either standard of care PET/CT or DMI PET/CT. DMI PET/CT identified all the lesions seen using the standard of care PET/CT, as well as additional 37 FDG-avid lesions in 14 of 50 patients (28%). DMI PET/CT identified more lymph nodes than standard of care PET/CT in 12 patients. A similar finding was noted for lung nodules in 5 patients, liver lesions in 2 patients and other areas of additional focal uptake in 1 patient. Same number of osseous lesions were detected on both scans for all patients. Representative cases are shown in Figs max concordance is graphically depicted with a Bland-Altman Plot in On a per lesion analysis a total of 144 lesions were detected, of which 37 only by DMI PET/CT. Within 107 lesions detected by both scans there were 55 lymph nodes, 15 lung nodules, 9 liver lesions, 10 bone lesions and 18 other lesions . The agreement between lesion SUVmax were 1.2–17.4 (mean ± SD: 6.1 ± 3.3) for standard PET/CT, 1.7–24.9 (mean ± SD: 8.9 ± 4.8) for DMI PET/CT with TOF and Q.Clear® and 1.7–23.6 (mean ± SD: 7 ± 4) for DMI PET/CT without TOF and Q.Clear®. SUVmax differences between standard and DMI (with TOF and Q.Clear® on or off) were statistically significant for all 107 lesions (P < 0.0001) and not correlated to the delay to acquisition of DMI PET data (P < 0.0001)  Figs –7.max measurements for all 107 lesions were 1.2–14.6 (mean ± SD: 2.8 ± 2.8) and 3.3–7.4 (mean ± SD: 0.8 ± 1.4), higher on DMI PET/CT  compared with standard of care PET/CT.The SUVP < 0.05) when comparing separately differences in SUVmax between D600 and D690 vs DMI PET/CT (TOF and Q.Clear® on or off).For 14 of the 50 participants standard PET/CT used D600  and for 36 of the 50 participants it used D690 . The differences stayed statistically significant (max of 0.6–9.8 (mean ± SD: 3.8 ± 2.4).The 37 lesions detected only by DMI included 24 lymph nodes, 10 lung nodules, 2 liver lesions and 1 other lesion (pleura). These additional lesions had SUVmax for lymph nodes (P < 0.0001), lung nodules (P = 0.0003), bone lesions (P < 0.02), liver nodules (P < 0.01) and other lesions (P = 0.0004) when DMI PET images were reconstructed using TOF and Q.Clear®. When TOF and Q.Clear® were not used, the differences in SUVmax between standard PET/CT and DMI PET/CT remained statistically significant for lymph nodes (P = 0.0004), lung nodules (P = 0.02), bone lesions (P = 0.04) and other lesions (P = 0.002), but not for liver (P = 0.16). This data is shown in Tables The single organs analyses in 107 lesions identified on both scans showed a statistically significant difference in SUVs P < 0.0, liver nP = 0.000, bone len = 107) and backgrounds tissues such as the aortic arch and liver. The lesion:aortic arch ratios ranged 0.6–11.8 (mean ± SD: 3.5 ± 2.2) for standard PET/CT and 1.3–20.5 (mean ± SD: 6.7 ± 4.2) for DMI PET/CT (P < 0.0001). The lesion:liver ratios ranged 0.4–8.9 (mean ± SD: 2.6 ± 1.6) for standard PET/CT and 0.8–12.2 (mean ± SD: 4.4 ± 2.5) for DMI PET/CT (P < 0.0001). The mean lesion:aortic arch SUVmax ratio and mean lesion:liver SUVmax ratio were 0.2–15.2 (mean ± SD: 3.2 ± 2.6) and 0.2–8.5 (mean ± SD: 1.9 ± 1.4), respectively, higher in DMI PET/CT than standard PET/CT.We also analyzed the ratios of uptake in lesions  testing results for D600, D690 and DMI is shown in The meaningful role of hybrid imaging PET/CT in oncology has strongly been demonstrated . InformaIn this study we have shown the potentialities of a new generation PET/CT scanner (DMI PET/CT) compared to standard of care PET/CT (D600 and D690 PET/CT) in a cohort of 50 patients. DMI PET/CT results are at least comparable to D600 and D690 in terms of lesion detectability, with DMI PET/CT identifying all the 107 lesions found using standard of care PET/CT.®. This results in increased spatial and contrast resolution, potentially allowing for the detection of smaller lesions and more accurate quantification of uptake.The DMI PET/CT scanner uses small lutetium-based scintillator crystal arrays combined with a silicon photomultiplier (SiPM) bloc design, leading to a high NEMA sensitivity . The SiPmax measurements from standard PET/CT to DMI PET/CT for all 107 lesions detected by both scanners; this difference was statistically significant and not correlated with the time delay. Similar results were assessed by Nguyen et al [max in 52 avid FDG lesions from standard PET/CT to digital PET/CT.Our semi-quantitative analysis showed an increase of 53.3% in SUVen et al , who evamax differences per single organ were statistically significant comparing standard PET/CT to DMI PET/CT with TOF and Q.Clear® on; when DMI images were not reconstructed using TOF and Q.Clear®, differences in SUVmax between standard and DMI were statistically significant for all organs except the liver. We also compared separately D600 (non TOF enabled) vs DMI and D690 (TOF capable) vs DMI. DMI results (with TOF and Q.Clear® on or off) were superior to standard of care PET/CT, independent to the type of scanner used; equivalence between D600 and D690 was already shown [SUVdy shown .18F-FDG PET/CT for tumor detection [It is known that time delay can improve the diagnostic specificity of etection . Comparietection , 22 we r18F-FDG uptake compared to standard of care PET/CT in 14 of 50 patients. These findings are aligned to results showed by Nguyen et al [18F-FDG avid lesions comparing digital to standard PET/CT in 5 of 21 patients.DMI PET/CT detected 37 more areas of focal increased 18F-FDG uptake seen only by DMI PET/CT had a diameter < 0.7 cm and mean SUVmax of 0.6–4.9 (mean ± SD: 2.6 ± 2). These findings suggest better small lesion detectability using DMI PET/CT compared to standard of care PET/CT.The higher sensitivity provided by the DMI scanner is likely to allow for reduction of the injected dose of radiopharmaceutical without an impact on image quality. It is worth noting that 8 of the 37 areas of focal increased In our study, DMI PET/CT had better lesion conspicuity as indicated by higher lesion-to-background ratios compared to standard of care PET/CT for the lesion-to-aortic arch and lesion-to-liver.P value < 0.05. As demonstrated in We also demonstrated the similarity of background FDG uptake between scanner types using an equivalence testing and empirically choosing ± 0.7 g/mL SUV measurement to define non-equivalent with a 18F-FDG detected only by DMI PET/CT. Furthermore, we could not randomize the order of the standard PET/CT and DMI PET/CT, due to DMI pending FDA approval for clinical use at the time of the study.Our study had some limitations: the relatively small sample size of participants and the lack of histopathological confirmation for foci of Further studies in a larger cohort of patients and with histopathology as reference standard are needed to confirm these preliminary data.Discovery Molecular Insights is a new generation PET/CT scanner that brings together the sensitivity of SiPM detectors and innovative reconstruction technology. Our pilot study demonstrated that DMI PET/CT is more sensitive and can better detect smaller lesions compared to standard of care PET/CT scanners. It may make possible a reduction of the injected dose or the scan time (or a combination of both) without a reduction in image quality. Further studies will confirm its performance and its potential role in helping clinicians to diagnose and stage disease earlier and to better guide treatment strategies.S1 Dataset(XLS)Click here for additional data file.S1 FigA) Standard Maximum-intensity-projection (MIP) B) DMI MIP C) Standard coronal fused image D) DMI coronal fused image.(TIF)Click here for additional data file.S2 Fig18F-FDG injection. A1) D690 PET; A2) D690 CT; A3) D690 fused image B1) DMI PET; B2) DMI CT; B3) DMI fused image.DMI images were acquired 49.8 minutes after standard acquisition and 96.3 minutes after (TIF)Click here for additional data file.S3 FigA1) Standard PET, axial view B1) Standard CT, axial view C1) Standard fused image, axial view A2) DMI PET, axial view B2) DMI CT, axial view C2) DMI fused image, axial view.(TIF)Click here for additional data file.S1 TableM = male; F = female; DLBCL = diffuse large B-cell lymphoma; HL = Hodgkin lymphoma; NSCLC = non-small cell lung cancer; HCC = Hepatocellular carcinoma; FL = follicular lymphoma; UP = unknown primary.(DOCX)Click here for additional data file.


Acute CNS damage is commonly studied using rat and mouse models, but increasingly, molecular analysis is finding species differences that might affect the ability to translate findings to humans. Microglia can undergo complex molecular and functional changes, often studied by in vitro responses to discrete activating stimuli. There is considerable evidence that pro-inflammatory (M1) activation can exacerbate tissue damage, while anti-inflammatory (M2) states help resolve inflammation and promote tissue repair. However, in assessing potential therapeutic targets for controlling inflammation, it is crucial to determine whether rat and mouse microglia respond the same.+ channels —were examined  and specific channel blockers were applied to determine their contributions to microglial migration in the different activation states.Primary microglia from Sprague-Dawley rats and C57BL/6 mice were cultured, then stimulated with interferon-γ + tumor necrosis factor-α , interleukin (IL)-4 , or IL-10 . To profile their activation responses, NanoString was used to monitor messenger RNA (mRNA) expression of numerous pro- and anti-inflammatory mediators, microglial markers, immunomodulators, and other molecules. Western analysis was used to measure selected proteins. Two potential targets for controlling inflammation—inward- and outward-rectifier KPro-inflammatory molecules increased after I + T treatment but there were several qualitative and quantitative differences between the species . Several molecules commonly associated with an M2a state differed between species or they were induced in additional activation states . Resting levels and/or responses of several microglial markers  differed with the activation state, species, or both. Transcripts for several Kir2 and Kv1 family members were detected in both species. However, the current amplitudes (mainly Kir2.1 and Kv1.3) depended on activation state and species. Treatment-induced changes in morphology and migratory capacity were similar between the species . In both species, Kir2.1 block reduced migration and Kv1.3 block increased it, regardless of activation state; thus, these channels might affect microglial migration to damage sites.Caution is recommended in generalizing molecular and functional responses of microglia to activating stimuli between species.The online version of this article (doi:10.1186/s12974-017-0941-3) contains supplementary material, which is available to authorized users. Rats have been used for many years to model CNS damage and disease because they have many physiological similarities to humans and can learn a wide variety of tasks, which makes them useful for behavioral studies . More reWhen the CNS is injured, brain cells release “damage-associated molecular pattern” molecules  and other soluble mediators, including cytokines, high-mobility group box 1, purine metabolites, and nucleic acids. In response, microglia “activate”, and this is accompanied by dramatic morphological and molecular changes , 11. TheThe terminology for microglial activation is evolving , 13–16. + channels. Following acute CNS injury, rodent microglia in situ express inward-rectifier and outward-rectifier K+ currents , or with 20 ng/mL IL-4 [M(IL-4)] or 20 ng/mL IL-10 [M(IL-10)] to induce anti-inflammatory states. The recombinant cytokines  were specific to the rodent species; e.g., mIL-4 was used on mouse microglia. For messenger RNA (mRNA), protein, and functional analyses , the cells were stimulated for 24 h, which was chosen to facilitate comparisons with our previous studies of rat microglia  In mouse microglia, I + T significantly increased many of the same pro-inflammatory genes, but differences (bold numbers) were the increases in Ifng, Ifngr2, Il1r1, and Casp1, and lack of change in Tnfrsf1b. Other species differences were the greater induction of Nos2 (4.7-fold higher) and Ptk2b (5.1-fold higher) in rat, while mouse showed a greater induction of Tnf (3.2-fold) and Tnfrsf1a (1.5-fold). Induction of Ptgs2 by I + T was also greater in mouse cells (8.4-fold), and although it did not reach statistical significance, rat cells showed a nearly 26-fold increase (p = 0.07). M(IL-4). IL-4 did not induce expression of pro-inflammatory genes in either species. Instead, Ifngr1 decreased in mouse only and Tnfrsf1b showed opposite changes in expression in the two species . M(IL-10). There were no changes in pro-inflammatory transcript levels, except for an increase in Il1b (mouse only).As previously shown for rat microglia , 19, 56,ry state , with elOverall, only M(I + T) cells had increased mRNA expression of common pro-inflammatory genes. However, because quantitative differences were seen between rat and mouse, we next used Western analysis to examine protein changes for some key molecules. For iNOS, both mRNA and protein were induced, and only by I + T in both species Fig. . Importa. To further investigate anti-inflammatory responses, we also examined IL-4, IL-10, TGF-β1, and their receptors. Unstimulated. Both species had low transcript levels (<100 counts/200 ng sample) of the alternative-activation markers, Arg1, Ccl22, Retnla (FIZZ1), and Cd163, as well as Il4 and Il10 , Tgfbr1 (3.8-fold), Il1rn (8.4-fold), Il4r (4.2-fold), and Pparg (8.6-fold). M(I + T). Both species showed significantly decreased transcript expression of Myc, and increased expression of Il4r, Il13ra1, Il10ra, and Tgfbr2. However, there was greater induction of IL4r, IL10ra, and IL13ra1 in mouse; and Tgfbr2 in rat. More prominent species differences were that in rat, Il10rb increased, and Tgfb1 and Pparg decreased, while in mouse Il1rn and Tgfbr1 increased. There were other apparent changes in “anti-inflammatory” genes that did not reach significance, likely because of the small sample size. These included reduced expression of Mrc1 in both species, increased Il1rn (3.3-fold), Arg1 (3.9-fold), and Ccl22 (2.8-fold) in rat, and increased Cd163 (2.9-fold) and Ccl22 (21.3-fold) in mouse. [A pilot study of rat microglia at 6 h after I + T showed increases in Ccl22 and Arg1 (data not shown).] M(IL-4). Both species showed increased expression of Mrc1 and Myc, but with 1.5-fold greater induction in rat. Ccl22 induction was also greater in rat (220.8-fold increase) than mouse (128-fold). Conversely, the increase in Arg1 was much greater in mouse (1005-fold) than rat (6.3-fold). Species-specific changes were that Il10rb decreased in rat only, Il10ra and Tgfbr2 decreased in mouse only; Il1rn, Retnla, Pparg, and Tgfb1 increased in mouse only, and Cd163 increased in rat only. We examined several genes known to be upregulated by IL-4 in mouse microglia: ARG1, “found in inflammatory zone” 1 (FIZZ1), MRC1/CD206, CCL22, CD163, and peroxisome proliferator-activated receptor gamma (PPAR-γ) . To furtM(IL-10). Responses were very different from IL-4. In both species, there was increased expression of Tgfbr2, but there were several species differences. In mouse, Il4r, Il13ra1, and Mrc1 increased, while the only rat-specific change was an increase in Il10rb. There was a trend toward a decrease in Myc expression in mouse cells (p = 0.06).Arg1 in rat only and Ccl22 in mouse only. Therefore, we examined protein changes for some key anti-inflammatory markers. As expected, CD206/MRC1 was induced by IL-4 treatment in both species and at both the mRNA  are, in fact, good markers for both rodent species. For instance, IL-4 treatment showed several species differences, and the M1 stimulus (I + T) increased RNA Fig.  and protRNA Fig. . HoweverRNA Fig. , and a hRNA Fig.  correspoRNA Fig. . As expeRNA Fig.  was muchUnstimulated. Microglia of both species expressed low transcript levels of Socs1  and Socs3 (Table Itgam (CD11b), Cd68, Cx3cr1, toll-like receptor 2 (Tlr2), Tlr4, Nfkbia , Tspo (translocator protein), Nr3c1 , and Ptpn6 . Interestingly, most control transcript levels were higher in rat than in mouse microglia, except for Cx3cr1, which was higher in mouse. Surprisingly, mRNA for Aif1  was highly expressed in rat cells  but very low in mouse. This species difference was confirmed at the protein level, where Iba1 was much higher in unstimulated rat microglia . Both species showed increased transcript expression of Nr3c1, Nfkbia, Socs1, and Tspo, and decreased Cd68 and Cx3cr1. The main species differences included the higher increase in mouse cells for Socs1 (5.7-fold) and Tspo (4-fold), and the mouse-specific decrease in Itgam and increases in Aif1, Ptpn6, Socs3 and Tlr4 (which decreased in rat). Although both species showed modest increases in Iba1 protein , they did not reach statistical significance . Both species showed increased expression of Tlr4 and decreased Tlr2. [A pilot study of rat microglia also showed an increase in Tlr4 at 6 h (not shown)]. Socs1 increased in rat, and despite a 33-fold increase in mouse, it did not reach significance (p = 0.08). Notable species differences were increased Cx3cr1 in rat but a dramatic decrease in mouse, decreased Cd68 in rat but an increase in mouse, lower Itgam in rat, and lower Ptpn6 in mouse. M(IL-10). Both species showed increased expression of Itgam and Tlr4, and decreased Cx3cr1. Species differences were increases in Cd68 and Tspo (rat only), and increase in Nc3r1 (mouse only). Mouse had increased Socs3, but the 9.4-fold increase in rat was not significant. Interestingly, Itgam was the only gene selectively upregulated by IL-10 treatment in both species.We next examined several molecules routinely used to identify “activated” microglia after acute brain injury, and several immunomodulatory molecules. s3 Table : Fig. S4nce Fig. . M(IL-4)Nfkbia. Several genes increased with I + T but were not selective pro-inflammatory markers. I + T increased Nc3r1 and Socs3 but they were also slightly increased with IL-10 in mouse; Socs1 and Tspo were elevated in both species but also increased by IL-4 or IL-10 in rat. Several genes were not selective activation state markers in either species at the mRNA level: Aif1, Itgam, Cd68, Cx3cr1, Tlr2, Tlr4, and Ptpn6. Of note, I + T treatment decreased Cx3cr1, which is often used to identify microglia, and Cd68, which is often used to identify their phagocytic state.Overall, the pro-inflammatory state in both species was marked by increased mRNA expression of Unstimulated. Microglia from both species had modest transcript levels (<500 mRNA counts/200 ng RNA) of Nox1, Nox4, P2rx7, P2ry2, and P2ry12; and higher levels (>2500 mRNA counts) of Cybb (NOX2), Fcgr2b (CD32), Fcgr3a (CD16), and Msr1 (SR-A) , Ncf1 (9.3-fold), Trem2 (31.2-fold), and P2ry12 (3.4-fold). M(I + T). Both species had decreased transcript levels of Trem2, and increased Ncf1 , Cybb (1.8-fold greater in mouse), and Fcgr3a (1.5-fold greater in rat). Other species differences were the opposite changes in Fcgr1a expression, the increase in P2ry2 in rat only, and decreases in Msr1 and P2ry12 in rat only. M(IL-4). Both species showed decreased expression of Cybb. All other changes were species dependent, with most genes altered in rat cells but few in mouse. In rat, Fcgr2b, P2rx7, and P2ry12 increased, while Trem2 and Msr1 decreased. Mouse-specific changes were decreased P2ry12 and increased Nox1, which were opposite to the trends in rat. M(IL-10). In both species, Fcgr2b and Fcgr3a increased. Species differences were increases in P2ry2 and Trem2 in rat only, increased Msr1 and P2ry12 in mouse only, and opposite changes in Cybb . Nox4 transcripts remained low in both species and were unaffected by any treatment tested.We conducted a species comparison of transcript levels of several molecules to follow up on our recent study of myelin phagocytosis and consequent production of reactive oxygen species (ROS) in rat microglia . We alsocgr3a CD1, and MsrKcnj2 (Kir2.1), Kcnj12 (Kir2.2), Kcnj4 (Kir2.3), and Kcnj14 (Kir2.4). . Inted Fig. . In a pi members .Fig. 7K+Kcna2), Kv1.3 (Kcna3), and Kv1.5 (Kcna5) mRNA and protein have been detected in primary rat microglia  M(IL-10). There were no significant changes. Overall, the most notable species differences were the opposite changes evoked by I + T in Kir2.1 and Kv1.2, and the IL-4-evoked increase in Kv1.3 in rat only and Kv1.5 in mouse only.Kv1.2  relation (not shown). Thus, to examine whether microglial activation states affect channel activity, we compared the total Kir current density. In both species, the I-V relations showed inward rectification and reversal at about − 82 mV after junction potential correction . The Kir2.1 current decreased in mouse cells but was unchanged in rat. M(IL-4). The Kir2.1 current decreased substantially in rat cells but was unchanged in mouse. M(IL-10). The Kir2.1 current decreased in mouse cells but was unchanged in rat. Overall, the Kir2.1 current amplitude is not a reliable indicator of the microglial activation state, but instead depends on both the rodent species and activating stimulus.To quantify the Kir2.1 component of the whole-cell current, we used ML133, a membrane-permeant blocker  that acticroglia . Regardlion Fig. , which ient Fig.  despite ent Fig. . Speciesent Fig. . M(I + TI-V relation without time dependence during steps (not shown) and was not identified. In all activation states, the total Kv current and AgTx-2-sensitive Kv1.3 component activated in a time- and voltage-dependent manner above about − 60 mV , and increased with depolarization . The shape of the cells changed dramatically, but with subtle species differences. Rat microglia were clustered into chains of cells, while mouse microglia were more evenly distributed. For both species, but especially in mouse microglia, individual cells often appeared flat and round or amoeboid. There was no apparent cell polarity but the cells often bore short, spiky processes. M(IL-4). Both species showed many unipolar cells, and many of them contained a podonut. The morphology of rat microglia was more heterogeneous and lamellae were often smaller than in unstimulated rat cells or M(IL-4) mouse cells. M(IL-10). For rat microglia, we previously reported that IL-10 increases the proportion of cells containing podonuts [Very little is known about migration of mouse microglia in different activation states or roles of Kir2.1 and Kv1.3 channels in migration in either rodent species see “”. Therefnut Fig. . There wpodonuts ; howeverRat. M(I + T) cells migrated 2.9-fold less than unstimulated cells, while M(IL-4) and M(IL-10) migrated more, by 1.8-fold and 2.4-fold, respectively  cells, increased 1.9-fold in M(IL-4) cells, and 3.3-fold in M(IL-10) cells. Blocking Kir2.1 also inhibited migration of mouse microglia: by 7.2-fold in M(IL-4) cells and 14.2-fold in M(IL-10) cells. Again, while ML133 apparently reduced migration by 5.8-fold in unstimulated mouse microglia and by 20.8-fold in M(I + T) cells, ANOVA did not show statistical significance. Similar to rat cells, blocking Kv1.3 with AgTx-2 increased migration of unstimulated mouse microglia (2.9 fold), M(IL-4) cells (1.9 fold), and M(IL-10) cells (1.4 fold). While AgTx-2 appeared to increase migration of M(I + T) cells (3.2 fold), it was not statistically significant. For both species, we ruled out changes in the number of cells available to migrate. That is, a CyQuant NF assay showed that neither channel blocker altered the cell density over the 24-h migration period, regardless of activation state (data not shown).Next, we quantified migration in each activation state. ely Fig. . Blockinely Fig. . Althougely Fig. . Mouse. Together, our results suggest that in microglia of both species, Kir2.1 activity facilitates migration and Kv1.3 activity inhibits it, regardless of the activation state.We compared gene expression, six proteins, two ion currents, and the role of these channels under four defined activation states in primary rat and mouse microglia. We found both similarities and differences between these species. Because several activation paradigms and outcomes were examined; for clarity, the most relevant literature will be discussed under four topics: molecular profiling of activation responses; Kir2.1 channel expression and current; Kv1.3 channel expression and current; and contributions of Kir2.1 and Kv1.3 channels to migration. In addition, for ease of comparison with the literature, protein names will be used if the gene names are not commonly used . The information obtained can stand on its own or be used to decide which genes are interesting candidates for further study, including protein analysis by immunohistochemistry or Western blots. Most DNA is transcribed , and thuWhile most studies of microglial activation have used primary rat or mouse cells, very few direct species comparisons have been made and the cell activation state was often not determined. We found one older study that directly compared primary rat and mouse microglia but it was restricted to glutamate secretion from unstimulated cells . An inteImportantly, we first used numerous gene markers to validate that their starting  state was similar. For instance, as expected for “resting” microglia, both species showed very low mRNA levels of numerous pro-inflammatory mediators  and anti-inflammatory mediators . Both species also had similar (moderate to high) mRNA levels of several cytokine receptors  as well as ICE, MRC1, MYC, PYK2, TGF-β1 and TNF-α.M(I + T). This pro-inflammatory stimulus evoked several similar gene responses in rat and mouse microglia, although there were quantitative differences. There were increased transcript levels of several well-known pro-inflammatory genes , NOX enzymes , and some less frequently examined genes (PYK2). Previously, increased PYK2 immunoreactivity was seen in rat microglia in vivo after transient middle cerebral artery occlusion or kainate-induced seizures [seizures , and altseizures . Similarseizures .M(IL-4). IL-4 evoked several important species differences in gene expression. We found that two markers commonly used to identify IL-4-mediated alternative activation in mice, FIZZ1 and PPAR-γ [d PPAR-γ , 77, werd PPAR-γ , and a md PPAR-γ . We found PPAR-γ . Thus, mM(IL-10). In both rodent species, responses to IL-10 often differed from IL-4. For instance, IL-10 did not reduce transcript expression of pro-inflammatory mediators  or increase anti-inflammatory mediators . Opposite effects were sometimes seen in the two species . Surprisingly, M(IL-10) and M(I + T) microglia shared many similarities, but not in induction of pro-inflammatory mediators. Instead, both stimuli induced transcripts of molecules known to modulate inflammatory responses , but the IL-10 responses were either comparable or lower than I + T responses. Induction of these modulatory genes suggests that self-limiting feedback responses occurred. Similarly, an earlier study of mouse microglia showed that IL-10 increased IL-1RN and SOCS3 mRNA levels and production of IL-6, CXCL1 and CCL2, but generally to a lesser extent than stimulation with LPS, TNF-α, or IL-1β [or IL-1β .3CR1 expression was surprisingly malleable and species dependent. This receptor is restricted to microglia in the CNS, where it binds neuron-derived CX3CL1, and helps maintain microglial quiescence [3CR1 transcript levels increased slightly after IL-4 treatment only in rat microglia, the most striking change was a decrease following all cytokine treatments in mouse. It is difficult to predict the outcome of these changes. CX3CR1-null mice are said to display microglial hyper-activation [3CR1 deletion was beneficial in models of transient ischemia [There are important implications of these molecular profiles of microglial inflammatory responses. (i) Some molecules routinely used to identify “activated” microglia/macrophages in vivo  were not selectively increased by the pro-inflammatory stimulus, I + T. However, Iba1 was selectively increased in both species. (ii) The only genes induced selectively by IL-10 were CD11b (rat and mouse), CD68, and TREM2 (rat only), and FcγRIIb, P2Y12, and SR-A (mouse only). CD11b and CD68 are often used as markers of microglial “activation” in vivo , 80, butiescence . While ttivation  but its ischemia  and Alzhischemia  but harmischemia .Previously reported changes in Kir and Kv currents with microglial activation have been inconsistent and hinted at species differences.The first reports of Kir2.1 mRNA transcripts in microglia were in the mouse cell lines, BV-2 and C8-B4 , 88, and+-dependent relaxation at very hyperpolarized potentials, a single channel conductance of 25–30 pS [2+. It is important to note that Ba2+ is a poor blocker at depolarized potentials [While the mRNA results suggest that the Kir current in rodent microglia is produced by Kir2.1, numerous patch-clamp studies have not confirmed its identity. It is useful to compare electrophysiological and pharmacological properties of the microglial current with cloned Kir2.1 channels. Like cloned Kir2.1, the microglial current shows fast activation kinetics, Na25–30 pS , 89, and25–30 pS , 90. Thetentials , and thitentials . In the tentials , its efftentials  but the Published responses of the Kir current to microglial activating stimuli have been quite variable. While the Kir current is present in most unstimulated rat and mouse microglia, effects of pro-inflammatory stimuli in rat microglia range from no change to a small increase (with LPS) or a large increase (with IFN-γ) , 92, 93.Kv1.2, Kv1.3, and Kv1.5 transcripts and protein have been detected in primary rat and mouse microglia , 63–65. There is considerable variability in the prevalence of Kv currents reported in primary rodent microglia. For unstimulated microglia, early reports showed an absence in rat , 97, preSome variability in the prevalence and amplitude of Kv currents is very likely due to differences in voltage protocols; e.g., using a depolarized holding potential, which is well known to inactivate Kv1.3 , 28, 93 Both Kv1.3 and Kv1.5 currents have been reported in ex vivo rat microglia  and in MLittle is known about Kv currents in anti-inflammatory states. Mouse microglia stimulated with TGF-β1 or TGF-β2 had increased Kv currents that were blocked by kaliotoxin, charybdotoxin, or MgTx , 88 but + currents) with MgTx in rat microglia reduced chemotaxis induced by monocyte chemoattractant protein 1 (CCL2) or ADP but, again, the activation state was not identified [For rat microglia, we previously reported that M(LPS) cells migrate less than unstimulated cells, while M(IL-4) and M(IL-10) cells migrate more –20, 30, entified .2+ signaling, which is involved in cytoskeletal remodeling, adhesion and migration [2+ entry through Ca2+ release-activated Ca2+ (CRAC) channels is increased with hyperpolarization [2+ entry and migration [2+ prolonged depolarization and reduced the amplitude of ATP-induced Ca2+ transients [+ flux and membrane potential. For instance, cell migration requires integrins, which regulate adhesion to the extracellular matrix, and there is evidence that integrins and K+ channels can co-exist in signaling complexes [1 integrin moiety was reported in T lymphocytes and melanoma cells [One potential role for Kv and Kir channels is to regulate the membrane potential and subsequent Caigration , 108. Forization , and thaigration . Similaransients . Blockinansients . This isansients . One posomplexes . A physima cells , 113 but1. Many molecules were analyzed using gene profiling to assess microglial activation responses. While mRNA levels are often well linked to protein levels, this is not always the case. Several interesting changes were further assessed at the protein and functional level. While it is important that channel proteins were assessed at the level of functional expression (ion currents), their role in only one cell function was examined (migration). In future, it would be valuable to assess roles of proteins in other microglial functions in the different activation states; e.g., relating ECM-degrading enzymes to invasion (as in ), relatiStudies of disease mechanisms rely on animal models, especially rodents. To fulfill the larger goal of translating such results to humans, it is crucial to understand whether rats and mice respond the same and, if not, which species is a more reliable model. There is considerable debate as to how closely mouse models resemble human responses in inflammatory diseases , 124. WhMicroglial properties are increasingly being investigated in human tissue—often in surgical biopsies from epileptic patients—but the cell activation state is usually not determined and they cannot be assumed to be normal, resting cells. Further complications include the potential for strain differences in rodents , and gen+ channels can be targeted to modulate neuroinflammation, it will be crucial to undertake species comparisons of other microglia functions using selective inhibitors or cell-targeted channel depletion, and to extend the studies to models of acute CNS injury.The present study contributes considerable comparative data concerning primary rat and mouse microglia, and it highlights species similarities and differences in the inflammatory response following stimulation with pro- and anti-inflammatory cytokines. The search for molecular targets to control microglial activation and specific functions will also require a better understanding of species differences. For instance, Kv1.3 is considered a promising target in autoimmune diseases, such as multiple sclerosis, rheumatoid arthritis, type 1 diabetes, and psoriasis . HoweverAdditional file 1:n = 4–6 individual cultures). *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001 (PDF 386 kb)Transcript expression of pro-inflammatory mediators. Rat and mouse microglia were unstimulated (CTL) or stimulated with IFN-γ and TNF-α (I + T), IL-4 or IL-10 for 24 h. mRNA counts for each gene were normalized to two housekeeping genes (described in Methods) and are shown as mean ± SEM . *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001 (PDF 364 kb)Transcript expression of anti-inflammatory mediators. Rat and mouse microglia were unstimulated (CTL) or stimulated with IFN-γ and TNF-α (I + T), IL-4 or IL-10 for 24 h. mRNA counts for each gene were normalized to two housekeeping genes (described in Methods) and are shown as mean ± SEM . *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001 (PDF 380 kb)Transcript expression of anti-inflammatory cytokines and their receptors. Rat and mouse microglia were unstimulated (CTL) or stimulated with IFN-γ and TNF-α (I + T), IL-4 or IL-10 for 24 h. mRNA counts for each gene were normalized to two housekeeping genes (described in Methods) and are shown as mean ± SEM . *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001 (PDF 473 kb)Transcript expression of microglia markers and immune modulators. Rat and mouse microglia were unstimulated (CTL) or stimulated with IFN-γ and TNF-α (I + T), IL-4 or IL-10 for 24 h. mRNA counts for each gene were normalized to two housekeeping genes (described in Methods) and are shown as mean ± SEM . To facilitate species comparisons, the Y-axis was the same except as indicated by blue boxes. *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001 (PDF 322 kb)Transcript expression of phagocytosis and purinergic receptors and NOX enzymes. Rat and mouse microglia were unstimulated (CTL) or stimulated with IFN-γ and TNF-α (I + T), IL-4 or IL-10 for 24 h. mRNA counts for each gene were normalized to two housekeeping genes (described in Methods) and are shown as mean ± SEM (