Development and Characterization of Ultrasound-assisted PLGA Nanobubbles for the Triggered Delivery of Pemigatinib by Design of Experiments

This study used a central composite design (CCD) to evaluate how the independent attributes—sonication distance (X1), amplitude (X2), time (X3), & power (X4)—impacted two outcomes—droplet-size (Y1) & polydispersity index (PDI) (Y2). With the time-consuming & inefficient "changing one factor at a time" approach would have been an option for this multifactor optimization, but we opted against it since we wanted to be sure we had the best possible values. Here, a mathematical model of the combined influence of the processing elements led to the selection of a CCD, which is known to be significantly more dependable. Using a double emulsion technique, we created and fine-tuned a drug delivery system consisting of nanobubbles loaded with pemigatinib. Pemigatinib nanobubbles were studied for their shape, surface charge, and particle size to determine their physicochemical characteristics and found that they were spherical with the Zeta potential (ZP) and particle size of -25.3±2.98 & 38.53±2.14 respectively. Pemigatinib-loaded nanobubbles were also tested for their release behaviors and drug encapsulation effectiveness. Finally, we tried to study the anti-tumor activity and cellular absorption of poly (lactic-co-glycolic acid) nanobubbles-Pemigatinib in vitro.

Preliminary clinical application of novel magnetic navigation and ultrasound-guided percutaneous transhepatic cholangiography drainage through the right liver duct for malignant obstructive jaundice / 中华内科杂志

Objective:To analyze the clinical application value of a novel magnetic navigation ultrasound (MNU) combined with digital subtraction angiography (DSA) dual-guided percutaneous transhepatic biliary drainage (PTCD) through the right hepatic duct for the treatment of malignant obstructive jaundice.Methods:Randomized controlled trial. The clinical data of 64 patients with malignant obstructive jaundice requiring PTCD through the right hepatic duct at the Hepatobiliary Center of the First Affiliated Hospital of Nanjing Medical University (Jiangsu Province People′s Hospital) from December 2018 to December 2021 were retrospectively analyzed. The MNU group ( n=32) underwent puncture guided by a novel domestic MNU combined with DSA, and the control group ( n=32) underwent puncture guided by traditional DSA. The operation time, number of punctures, X-ray dose after biliary stenting as shown by DSA, patients' tolerance of the operation, success rate of the operation, pre- and post-operative total bilirubin, and incidence of postoperative complications were compared between the two groups. Results:The operation time of the MNU group was significantly shorter than that of the control group [(17.8±7.3) vs. (31.6±9.9) min, t=-6.35, P=0.001]; the number of punctures in the MNU group was significantly lower [(1.7±0.6) vs. (6.3±3.9) times, t=-6.59, P=0.001]; and the X-ray dose after biliary stenting as shown by DSA in the MNU group was lower than that in the control group [(132±88) vs. (746±187) mGy, t=-16.81, P<0.001]; Five patients in the control group were unable to tolerate the operation, and two stopped the operation, however all patients in the MNU group could tolerate the operation, and all completed the operation, with a success rate of 100% (32/32) in the MNU group compared to 93.8%(30/32) in the control group; the common complications of PTCD were biliary bleeding and infection, and the incidence of biliary bleeding (25.0%, 8/32) and infection (18.8%, 6/32) in the MNU group was significantly lower than that in the control group, 53.1% (17/32) and 28.1% (9/32), respectively. Conclusion:Magnetic navigation ultrasound combined with DSA dual-guided PTCD through the right biliary system for the treatment of malignant obstructive jaundice is safe and feasible.

Sonication as a tool for disrupting biofilms and recovering microorganisms in bladder catheters / Sonicação como uma ferramenta para romper biofilmes e recuperar microrganismos em cateteres vesicais

Abstract Introduction: Urinary catheter-related infection is commonly associated with bacterial biofilm. The impact of anaerobes is unknown, but their detection in the biofilm on this device has not been previously reported. This study aimed to evaluate the capability to recovery strict, facultative, and aerobic microorganisms in patients using bladder catheters from ICUs using conventional culture, sonication, urinary analysis, and mass spectrometry. Methods: Parallel, sonicated bladder catheters from 29 critically ill patients were compared with their routine urine culture. Identification was performed using matrix-assisted laser desorption/ionization with time-of-flight mass spectrometry. Results: The positivity rate in urine (n = 2, 3.4%) was lower than that in sonicated catheters (n = 7, 13.8%). Conclusion: Bladder catheter sonication showed more positive culture results than urine samples for anaerobic and aerobic microorganisms. The role of anaerobes in urinary tract infection and catheter biofilm is discussed.

Resumo Introdução: A infecção relacionada ao cateter urinário é comumente associada ao biofilme bacteriano. O impacto dos anaeróbios é desconhecido, mas sua detecção no biofilme deste dispositivo não foi relatada anteriormente. Este estudo teve como objetivo avaliar a capacidade de recuperar microrganismos estritos, facultativos e aeróbios em pacientes que utilizam cateteres vesicais de UTIs utilizando cultura convencional, sonicação, análise urinária e espectrometria de massa. Métodos: Paralelamente, foram comparados cateteres vesicais sonicados de 29 pacientes gravemente enfermos com sua urocultura de rotina. A identificação foi realizada utilizando dessorção/ionização a laser assistida por matriz com espectrometria de massa por tempo de voo. Resultados: A taxa de positividade na urina (n = 2; 3,4%) foi inferior à dos cateteres sonicados (n = 7; 13,8%). Conclusão: A sonicação do cateter vesical apresentou resultados de cultura mais positivos do que as amostras de urina para microrganismos anaeróbios e aeróbios. É discutido o papel dos anaeróbios na infecção do trato urinário e no biofilme do cateter.

Epidemiología de las infecciones post osteosíntesis detectadas por baño de ultrasonido en el Servicio de Traumatología del Hospital Universitario de Caracas / Post osteosinthesis infections epidemiology by ultrasound bath in the trauma service of the University Hospital of Caracas

El objetivo de este trabajo es determinar la epidemiología de la infección post osteosíntesis a través de cultivos de fluidos sonicados en los pacientes del Hospital Universitario de Caracas en el período comprendido entre noviembre 2021-noviembre 2022. Se realizó un estudio observacional de tipo, serie de casos, a través de la revisión de historias médicas de todos los casos que acudieron con diagnóstico de infección post osteosíntesis a fin de determinar cuál agente causal fue el más común, factores de riesgo asociados y tratamiento de elección. Se incluyeron 10 pacientes, 70% de sexo masculino y edad promedio de 40,6±17,9 años. Los gérmenes aislados en el cultivo convencional fueron el SAMS, SAMR, Enterobacter cloacae, Staphylococcus coagulasa negativo (10,0% cada uno), el 60,0% de los cultivos en esta modalidad fueron negativos, en el cultivo de fluidos por baño de ultrasonido, el germen más frecuente fue el SAMR en el 30% de los casos, seguido del SAMS con 20%, en menor medida un caso de Staphylococcus coagulasa negativo y una infección polimicrobiana compuesta por K. pneumoniae, E. cloacae y Enterococo sp. El tratamiento médico consistió en antibioticoterapia vía endovenosa, se realizó de acuerdo al antibiograma obtenido del cultivo, el más empleado fue la cefazolina en 30% (en casos de SAMS), seguido de la vancomicina + meropenem y la vancomicina aislada en 20%. Todos los pacientes cumplieron tratamiento al menos por 4 semanas con evolución satisfactoria(AU)

The objective of this work is to determine the epidemiology of post-osteosynthesis infection through sonicated fluid cultures in patients at the Hospital Universitario de Caracas in the period between November 2021 and November 2022. An observational study of type, series of cases, through the review of the medical records of all the cases that presented with a diagnosis of post-osteosynthesis infection in order to determine which causative agent was the most common, associated risk factors and treatment of choice. 10 patients were included, 70% male and mean age 40.6 ± 17.9 years. The germs isolated in the conventional culture were SAMS, SAMR, Enterobacter cloacae, coagulase-negative Staphylococcus (10.0% each), 60.0% of the cultures in this modality were negative, in the culture of fluids by bath of On ultrasound, the most frequent germ was MRSA in 30% of cases, followed by SAMS with 20%, to a lesser extent a case of coagulase-negative Staphylococcus and a polymicrobial infection made up of K. pneumoniae, E. cloacae and Enterococcus sp. The medical treatment consisted of intravenous antibiotic therapy, it was carried out according to the antibiogram obtained from the culture, the most used was cefazolin in 30% (in cases of SAMS), followed by vancomycin + meropenem and vancomycin alone in 20%. All patients complied with treatment for at least 4 weeks with satisfactory evolution(AU)

Investigation on formulation parameters of donepezil HCl loaded solid lipid nanoparticles

Abstract Donepezil-HCl is a member of the acetylcholinesterase inhibitors that is indicated for the symptomatic treatment of Alzheimer's disease (AD) and has many side effects. In this study, to reduce the side effects of Donepezil-HCl and increase the penetration of the drug through the blood-brain barrier, we aimed to design a solid lipid nanoparticle (SLN) formulation. The effects of the different formulation parameters, such as homogenization speed, sonication time, lipid and drug concentration, surfactant type and concentration, and volume of the aqueous phase, were assessed for optimization. The particle size and PDI increased with increasing lipid concentration but decreased with increasing amounts of surfactant (Tween 80) and co-surfactant (lecithin). When the homogenization rate and sonication time increased, the particle size decreased and the encapsulation efficiency increased. The optimized formulation exhibited particle size, PDI, encapsulation efficiency, and zeta potential of 87.2±0.11 nm; 0.22±0.02; 93.84±0.01 %; -17.0±0.12 mV respectively. The in vitro release investigation revealed that approximately 70% of Donepezil-HCl was cumulatively released after 24 hours. TEM analysis proved that spherical and smooth particles were obtained and formulations had no toxic effect on cells. The final optimized formulation could be a candidate for Donepezil-HCl application in Alzheimer's treatment with reduced side effects and doses for patients

Detection of Microorganisms in Clinical Sonicated Orthopedic Devices Using Conventional Culture and qPCR / Detecção de microrganismos em dispositivos ortopédicos sonicados clínicos usando cultura convencional e qPCR

Abstract Objective To evaluate the sensitivity and specificity of the quantitative real-time polymerase chain reaction (qPCR) for 16S rDNA gene screening using sonicated fluid from orthopedic implants. Methods A retrospective study was conducted on 73 sonicated fluids obtained from patients with infection associated with orthopedic implants. The samples were subjected to conventional culture and molecular testing using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) and qPCR for 16S rDNA. The cycle threshold values were used to define a cut-off of the qPCR of the 16S rDNA for negative and positive cultures. Results No statistical differences were observed between the positive and negative culture groups based on the time from the first surgery to infection (p= 0.958), age (p =0.269), or general comorbidities. Nevertheless, a statistical difference was found between the mean duration of antibiotic use before device removal (3.41 versus 0.94; p =0.016). Bacterial DNA was identified in every sample from the sonicated fluids. The median cycle thresholds of the positive and negative cultures were of 25.6 and 27.3 respectively (p< 0.001). As a diagnostic tool, a cycle threshold cut-off of 26.89 demonstrated an area under the curve of the receiver operating characteristic of 0.877 (p≤ 0.001). Conclusion The presence of antimicrobial agents for more than 72 hours decreased culture positivity, but did not influence the qPCR results. Despite this, amplification of the 16S rDNA may overestimate infection diagnosis.

Resumo Objetivo Avaliar a sensibilidade e a especificidade da reação em cadeia de polimerase em tempo real quantitativa (quantitative real-time polymerase chain reaction, qPCR, em inglês) para a triagem do gene rDNA 16S, com a utilização do fluido sonicado de implantes ortopédicos. Métodos Um estudo retrospectivo foi realizado em 73 fluidos sonicados obtidos de pacientes com infecção associada aos implantes ortopédicos. As amostras foram submetidas a cultura convencional e a teste molecular utilizando ionização e dessorção a laser assistida por matriz com espectrometria de massa por tempo de voo (matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, MALDI-TOF MS, em inglês) e qPCR para o gene rDNA 16S. Os valores limiares do ciclo foram usados para definir um ponto de corte para a qPCR do gene rDNA 16S para culturas negativas e positivas. Resultados Não foram observadas diferenças estatísticas entre os grupos de cultura positiva e negativa com base no tempo desde a primeira cirurgia até a infecção (p= 0,958), na idade (p= 0,269), ou nas comorbidades em geral. No entanto, uma diferença estatística foi encontrada entre a duração média do uso de antibióticos antes da remoção do dispositivo (3,41 versus 0,94; p= 0,016). O DNA bacteriano foi identificado em todas as amostras dos fluidos sonicados. Os limiares do ciclo médio de culturas positivas e negativas foram de 25,6 e 27,3, respectivamente (p< 0,001). Como uma ferramenta de diagnóstico, um corte do limite do ciclo de 26,89 demonstrou uma área sob a curva da característica de operação do receptor de 0,877 (p ≤ 0,001). Conclusão A presença de agentes antimicrobianos por mais de 72 horas diminuiu a positividade da cultura, mas não influenciou os resultados da qPCR. Apesar disso, a amplificação do rDNA 16S pode sobrestimar o diagnóstico de infecção.

Antitumor effect of low-frequency ultrasound combined with autophagy blockade in the mouse model of 4T1 breast cancer / 中华超声影像学杂志

Objective:To investigate the effect of low-frequency ultrasound combined with hydroxychloroquine (HCQ) on the growth and invasion of 4T1 breast cancer in mice.Methods:4T1 cells in logarithmic growth phase were divided into 4 groups: control group, ultrasound group, HCQ group and ultrasound combined with HCQ group. Western blot was performed to detect the effects of ultrasound combined with HCQ on the protein expression levels of autophagy-related proteins LC3, p62 and matrix metalloproteinase 9 (MMP-9). Transmission electron microscopy was used to observe the formation of autophagic vesicles. The cell proliferation and cell viability were determined by EdU and CCK-8. Transwell assay was performed to detect the effect of ultrasound combined with HCQ on the invasive ability of 4T1 cells. Flow cytometry was performed to detect the effect of ultrasound combined with HCQ on the apoptosis of 4T1 cells. The transplantation tumor model of 4T1 breast cancer in BALB/c mice was constructed.The mice were randomly divided into 4 groups ( n=5 for each group), including control group, ultrasound group, HCQ group, ultrasound combined with HCQ group. The tumor volume and mice body weight were evaluated and measured in each group every 2 days. Results:The expression of LC3-Ⅱ and p62 protein levels increased in the ultrasound combined with HCQ group, and intracellular autophagosome accumulation was evident by transmission electron microscopy. In cellular experiments, compared with the other groups, the ultrasound combined with HCQ group showed stronger growth inhibition, significantly decreased cell proliferation rate, decreased expression of MMP-9, and significantly inhibited invasion (all P<0.050). In the in vivo experiments, compared with the control group, the tumor growth rate of all 3 inter vention groups of mice decreased (all P<0.050), and the ultrasound combined with HCQ group had better therapeutic effects than the ultrasound and HCQ groups. The treatment effect of ultrasound combined with HCQ group was better than that of ultrasound and HCQ groups (all P<0.001). Conclusions:The combination of low-frequency ultrasound and hydroxychloroquine could synergistically inhibit tumor cell proliferation, promote apoptosis, and significantly inhibit tumor growth in 4T1 tumor-bearing mice.

Comparison between ultrasound-assisted and landmark-guided paramedian spinal anesthesia for hip fracture surgery in the elderly / 中华老年医学杂志

Objective:To investigate the efficiency of the ultrasonic-assisted positioning technique for lumbar anesthesia in elderly patients with hip fractures through the paramedian approach compared with body surface labeling.Methods:Patients(aged ≥65 years)with hip fractures were randomized(1∶1)to receive either ultrasound-assisted or landmark-guided paramedian spinal anesthesia in a lateral position.The primary outcome was the number of needle passes needed for a successful dural puncture.The secondary outcomes included one-pass success rate, number of needle attempts, one-attempt success rate, total time of spinal anesthesia and adverse effects.Results:A total of 88 subjects were randomized.The ultrasound-assisted approach significantly reduced the number of needle passes, compared with the landmark-guided approach[2.0(1.0-3.0) vs.5.0(3.0-8.8); Z=-4.708, P<0.001]. The one-pass success rate was higher in the ultrasound-assisted approach than in the landmark-guided approach[40.9%(18/44) vs.4.5%(2/44); χ2=16.565, P<0.001]. There was no statistical difference in the number of needle attempts and one-attempt success rate between the two groups( P>0.05 for both). The total time of spinal anesthesia was longer in the ultrasound-assisted group than in the landmark-guided group[252(218-317) s vs.168(143-195) s; Z=-5.592, P<0.001]. In the ultrasound-assisted group, fewer patients developed bloody cerebral spinal fluid taps than in the landmark-guided group[0%(0/44) vs.18.2%(8/44); χ2=6.738, P=0.009]. Conclusions:In elderly hip fracture patients, ultrasound-assisted paramedian spinal anesthesia is superior to the landmark-guided approach in reducing the number of needle passes and should be recommended for these patients.

Effect of ultrasound combined with 4-hydroxyphenyl-retinamide lipid microbubbles on type Ⅰ collagen α1 chain expression in keloid-derived fibroblasts / 中华皮肤科杂志

Objective:To investigate the effect of ultrasound combined with 4-hydroxyphenyl-retinamide (4-HPR) lipid microbubbles on type Ⅰ collagen α1 chain (COL1A1) protein expression in keloid-derived fibroblasts.Methods:In vitro cultured keloid-derived fibroblasts were divided into 3 groups: control group receiving conventional culture with incomplete Dulbecco′s modified Eagle′s medium (DMEM) , 4-HPR lipid microbubble group cultured with incomplete DMEM containing 15 mg/L 4-HPR lipid microbubbles, and ultrasound + 4-HPR lipid microbubble group cultured with incomplete DMEM containing 15 mg/L 4-HPR lipid microbubbles under ultrasound treatment. After 24-hour treatment, reverse transcription (RT) -PCR and Western blot analysis were performed to determine the mRNA and protein expression of COL1A1 in keloid-derived fibroblasts in each group. Intergroup comparison was carried out by using t test. Results:The mRNA relative expression level of COL1A1 was 1.00 ± 0.18, 0.69 ± 0.15 and 0.35 ± 0.18 in the control group, 4-HPR lipid microbubble group and ultrasound + 4-HPR lipid microbubble group respectively, and the protein relative expression level of COL1A1 was 0.93 ± 0.03, 0.74 ± 0.07 and 0.44 ± 0.06 in the above 3 groups respectively. Moreover, the mRNA and protein expression of COL1A1 was significantly lower in the 4-HPR lipid microbubble group and ultrasound + 4-HPR lipid microbubble group than in the control group ( P < 0.05 or 0.001) , and lower in the ultrasound + 4-HPR lipid microbubble group than in the 4-HPR lipid microbubble group ( P < 0.05) . Conclusion:Ultrasound combined with 4-HPR lipid microbubbles could markedly inhibit the mRNA and protein expression of COL1A1 in keloid-derived fibroblasts.

Efeito do BioGran® funcionalizado ao raloxifeno através do processamento ultrassônico no reparo ósseo em defeitos críticos de calvárias de ratos / Effect of BioGran® functionalized to Raloxifene through ultrasonic processing in bone repair in critical defects in rat calvaria

Este estudo teve como objetivo analisar o processo de formação óssea, bem como a microarquitetura óssea promovido pela associação entre o BioGran® e diferentes concentrações de raloxifeno através do método da sonoquímica em defeitos críticos realizados em calvária de rato. Em um total de doze ratos machos, um defeito crítico de 5 mm de diâmetro foi feito e preenchido com BioGran® 100% (Bg), BioGran® 90% associado à Raloxifeno 10%, e BioGran® 80% associado à Raloxifeno 20%. Aos 14 e 24 dias pós-operatório, foram aplicados os fluorocromos calceína e alizarina, de modo respectivo. A eutanásia ocorreu aos 30 dias após a realização do procedimento cirúrgico para enxertia dos biomateriais. Análise de microtomografia computadorizada (micro-CT) através dos parâmetros de superfiície de intersecção (i.S), fração de volume ósseo (BV/TV) e densidade de conectividade (Conn.Dn) e por microscopia confocal a laser por meio da dinâmica óssea, superfície de mineralização ativa e a taxa de aposição mineral (MAR). Os dados foram analisados através de análise estatística utilizando o teste de normalidade Shapiro-Wilk e o pós teste de Turkey (p<0.05). Para os parâmetros de micro-CT avaliados os menores valores foram encontrados no grupo BG+RL10% (p<0,05), valores similares foram encontrados entre os grupos BG e BG+RL20%. A microscopia confocal evidenciou melhor mineralização óssea e maior taxa de aposição mineral (MAR) para o Grupo BG+RL20% (p<0,05%). Conclusão: A concentração de Raloxifeno a 20% combinado ao BioGran® pelo método da sonoquímica parece ter acelerado o reparo ósseo(AU)

This study aimed to analyze the bone formation process, as well as the bone microarchitecture promoted by the association between BioGran® and different concentrations of raloxifene through the sonochemistry method in critical defects performed in rat calvaria. In a total of twelve male rats, a critical defect of 5 mm in diameter was made and filled with BioGran® 100% (Bg), BioGran® 90% associated with Raloxifene 10%, and BioGran® 80% associated with Raloxifene 20%. At 14 and 24 days postoperatively, the fluorochromes calcein and alizarin were applied, respectively. Euthanasia occurred 30 days after the surgical procedure for grafting biomaterials. Analysis of computed microtomography (microCT) through the parameters of intersection surface (iS), bone volume fraction (BV / TV) and connectivity density (Conn.Dn) and by confocal microscopy through bone dynamics, surface of active mineralization and the mineral apposition rate (MAR). Data were analyzed through statistical using the Shapiro-Wilk normality test and the Turkey post-test (p <0.05). For the me microCT parameters evaluated, the lowest values were found in the BG + RL10% group (p<0.05), similar values were found between the BG and BG + RL20% groups. Confocal microscopy showed better bone mineralization and higher mineral apposition rate (MAR) for the BG + RL20% Group (p < 0.05%). Conclusion: The concentration of Raloxifene at 20% combined with BioGran® by the sonochemistry method seems to have accelerated bone repair(AU)

Improvement of Mycobacterium tuberculosis detection in sputum using DNA extracted by sonication

Abstract Tuberculosis (TB) is one of the infectious diseases with high mortality in the world. DNA amplification techniques have been used to overcome barriers to the diagnosis of this disease. However, the success of these methodologies is highly dependent on the DNA obtained from the sample. This study was carried out to verify whether the DNA extracted by sonication (in house method) could yield suitable DNA for amplification by real-time PCR (qPCR). Sixty sputum samples were submitted to DNA extraction using sonication compared to a commercial method (Detect-TB kit, Labtest/MG-Brazil). All DNA samples were amplified by qPCR for IS6110 region (IS6110-qPCR/SYBR Green assay). Out of 60 samples, 40 were positive for TB; of these, all had positive results when extracted by sonication (100%) and 80% when extracted by the commercial method. The limit of detection (LOD) of Mycobacterium tuberculosis (H37Rv strain) by qPCR was 14CFU/mL when the DNA was extracted by sonication, compared to countless colonies when extracted by commercial kit. In conclusion, the sonication protocol (without purification step) proved to be a simple, fast, and suitable method for obtaining DNA for use in qPCR from sputum samples.

Estandarización y utilidad de la contrainmunoelectroforesis para el diagnóstico de la bartonelosis / Standardization and utility of counterimmunoelectrophoresis for the diagnosis of bartonellosis

RESUMEN Introducción. La enfermedad de Carrión, causada por Bartonella bacilliformis, es una enfermedad reemergente en el Perú, que se diagnostica convencionalmente mediante el frotis sanguíneo y el cultivo, los cuales son métodos poco sensibles, necesitándose métodos diagnósticos alternativos. Objetivos. Determinar la sensibilidad y especificidad de la contrainmunoelectroforesis (CIEF) utilizando un antígeno sonicado obtenido de una cepa de Bartonella sp., para detectar anticuerpos contra la bacteria comparado con el cultivo como estándar de referencia. Métodos. El antígeno para la prueba se obtuvo por sonicación de un aislado de Bartonella sp., cultivado en un medio bifásico con y sin sangre de carnero. La reactividad del antígeno sonicado fue evaluada por la CIEF empleando 123 sueros de personas, de los cuales 60 fueron de pacientes con diagnóstico clínico y bacteriológico de enfermedad de Carrión, 54 de personas con otras infecciones y 9 de personas sanas. Para la estandarización de la prueba de CIEF se evaluaron el tamaño y la distancia entre los pocillos, así como la concentración del antígeno y los volúmenes de los reactivos usados. Resultados. La concentración óptima del antígeno fue de 0,64 mg/mL, la distancia entre los pocillos de 3 mm, el tamaño de los pocillos de 3 mm y el volumen de los reactivos de 12 μL. La CIEF estandarizada tuvo una sensibilidad de 43,3% y una especificidad de 98,4%. Conclusiones. Los resultados de la CIEF revelan una baja sensibilidad de la prueba, pudiéndose usar como una prueba confirmatoria dada su elevada especificidad, pero no puede ser utilizada como prueba de tamizaje serológico por su escasa sensibilidad.

ABSTRACT Introduction. Carrion's disease, caused by the bacterium Bartonella bacilliformis, is a reemerging disease in Perú, which is conventionally diagnosed by blood smear and culture, which are not very sensitive methods, requiring alternative diagnostic methods. Objectives. To determine the sensitivity and specificity of the counterimmunoelectrophoresis (CIEP) using a sonicated antigen obtained from a Bartonella sp. strain, to detect antibodies against the bacteria compared to the culture as reference standard. Methods. The test antigen was obtained by sonication of an isolate of Bartonella sp., grown in a biphasic medium with and without sheep blood. The reactivity of the sonic antigen was evaluated by the CIEP using 123 sera from people, of which 60 were from patients with a clinical and bacteriological diagnosis of Carrion's disease, 54 from people with other infections and 9 from healthy people. For the standardization of the CIEP test, the size and distance between the wells were evaluated, as well as the concentration of the antigen and the volumes of the reagents used. Results. The optimal concentration of the antigen was 0,64 mg/mL, the distance between the 3 mm wells, the size of the 3 mm wells and the volume of the reagents of 12 μL. The standardized CIEP had a sensitivity of 43,3% and a specificity of 98,4%. Conclusions. The results of the CIEP reveal a low sensitivity of the test, being able to be used as a confirmatory test given its high specificity but cannot be used as a serological screening test due to the low sensitivity referred to.

Crecimiento y metabolismo de Fischerella TB22 en medio de cultivo BG11 / Growth and metabolism of Fischerella TB22 in BG110 culture medium

Los microorganismos fijadores de nitrógeno de vida libre, abarcan una gama morfológica que va desde los organismos unicelulares como las bacterias y algunas cianobacterias, hasta multicelulares, filamentosas, por ello es importante conocer cómo se comportan y se puede saber haciendo una curva de crecimiento microbiano. Para este estudio se prepararon 4 fotobioreactores de columna burbujeada con inoculo de Fischerella TB22, se pusieron en aireación constante con 12 horas luz y 12 horas obscuridad durante 40 días con diferentes tratamientos de ajuste de volumen del medio de cultivo y ajuste del pH. El objetivo de este trabajo fue evaluar el crecimiento en biomasa por peso seco, densidad óptica, pH y amonio de Fischerella sp. en medio de cultivo BG110 durante 12 días. Las variables que se midieron de la curva de crecimiento de las cianobacterias, siguieron el patrón de una curva típica de crecimiento microbiano. (AU)

Free-living nitrogen-fixing microorganisms cover a morphological range that goes from unicellular organisms such as bacteria and some cyanobacteria, to multicellular, filamentous, therefore it is important to know how they behave and can be known by makinga microbial growth curve. For this study, 4 bubbled column photobioreactors with Fischerella TB22inoculum were prepared, they were placed in constant aeration with 12 hours of light and 12 hours of darkness for 40 days with different treatments of volumeadjustment of the culture medium and pH adjustment. The objective of this work was to evaluate the biomass growth by dry weight, optical density, pH, and ammonia of Fischerella sp. in the BG110 culture medium for 12 days. The variables that were measured from the growth curve of cyanobacteria followed the pattern of a typical microbial growth curve. (AU)

Design And Optimization Of Doxorubicin Hcl Proniosomes By-Design Of Experiment

Objective: The present research work was designed to formulate and optimize doxorubicin HCl proniosomes by design of experiment (DoE). Methods: A 4-factor, 3-level Box-Behnken design was used to explain multiple linear regression analysis and contour 3D plot responses. The independent variables selected were tween 20, cholesterol, hydration volume and sonication time; dependent variables percentage entrapment efficiency (PEE), mean vesicle size (MVS). Based on the Box-Behnken design 29 trial runs were studied and optimized for PEE and MVS. Further "Model F-Value" was calculated to confirm the omission of insignificant terms from the full-model equation to derive a multiple linear regression analysis to predict the PEE and MVS of niosomes derived from proniosomes. 3D plots were constructed to show the influence of independent variables on dependent variables. Results: PEE of doxorubicin HCl proniosomes was found to be in the range of 40.21-87.5%. The polynomial equation for PEE exhibited a good correlation coefficient (0.5524) and the "Model F-Value" of 7.41 implies the model is significant. P-values less than 0.0500 indicate model terms are significant. The MVS of doxorubicin HCl proniosomes was found to be in the range of 325.2 nm to 420.25 nm. The mathematical model generated for MVS (R2) was found to be significant with model F-value of 54.22. There is only a 0.01% chance that a "Model F-Value" this large could occur due to noise (P<0.0500) and R2 value of 0.9004. Conclusion: The DoE of Box-Behnken design demonstrated the role of the derived equation, 3D plot in predicting the values of dependent variables for the preparation and optimization of doxorubicin HCl proniosomes. The results suggest that doxorubicin HCl proniosomes can act as a promising carrier.

HPLC-PDA method validated for the determination of hibalactone in Hydrocotyle umbellata subterraneous parts and its ultrasound-assisted extraction optimization

ABSTRACT In this work, we developed and validated a HPLC-PDA method for the quantification of hibalactone in Hydrocotyle umbellata L., Araliaceae, subterraneous parts extracts and optimized its ultrasound-assisted extraction. Chromatographic separations were carried out with an isocratic mobile phase of acetonitrile/methanol/water (10:65:25), a flow of 0.8 ml min−1, detection at 290 nm and C18 column (250 × 4.6 mm, 5 µm). The method validation parameters were determined according to Brazilian legislation. The optimization of the hibalactone ultrasound-assisted extraction was performed using Box-Behnken design and response surface methodology. The HPLC method for hibalactone quantification proved to be selective, linear, precise, accurate and robust, being useful for the analysis of hibalactone in H. umbellata subterraneous parts extracts. The optimal ultrasound-assisted extraction conditions were obtained with solid-to-liquid ratio of 1:5 g ml−1, ethanolic strength of 70% (v/v) and temperature of 65 °C. The results can provide support of the quality control and standardization of raw materials from H. umbellata.

Sonication of antibiotic loaded cement spacers: A valuable technique for detection of infection persistence in two-stage revision for infected joint arthroplasty

We evaluated the diagnostic utility of sonication of antibiotic loaded cement spacers comparing with periprosthetic tissue cultures for the detection of persisting infection in 14 patients undergoing staged procedures. Sonication improved microbial detection of intraoperative cultures from 14.2% to 28.5% (P = 0.481). Routine sonication of spacers is recommended.

Detection of periprosthetic joint infection pathogens by metagenomic next-generation sequencin / 中华骨科杂志

Objective@#To investigate and compare the capability of metagenomic next-generation sequencing (mNGS) in detecting pathogens and diagnosing of periprosthetic joint infection (PJI) from synovial fluid and sonicate fluid of patients who underwent revision arthroplasty.@*Methods@#Thirty-five consecutive patients who underwent revision arthroplasty from May 2018 to November 2018 were included prospectively. There were 22 males and 13 females, 11 hip revisions and 24 knee revisions. All the patients were divided into the PJI group and aseptic loosening (AL) group. Synovial fluid and sonicate fluid of the explanted prostheses were obtained for microbiological culture and mNGS tests. Periprosthetic tissues were only collected for culture. Synovial fluid of three patients undergoing primary arthroplasty been treated by sonication as the negative control group concurrently. Comparisons of microbiological results and diagnostic value from mNGS and culture tests were performed.@*Results@#In the 13 culture-positive PJI patients, mNGS results of synovial fluid were positive in 12 cases, while culture and mNGS results were completely consistent at species level in 7 cases, consistent at the genus level in 1 case. mNGS results of sonicate fluid were positive in 13 cases, while culture and mNGS results were completely consistent at species level in 9 cases, consistent at the genus level in 1 case. In 7 culture-negative PJI patients, 6 cases had consistent mNGS results at species level both from synovial fluid and sonicate fluid, however, one case had positive mNGS result only from sonicate fluid. All culture results and mNGS results of synovial fluid were negative in all 15 AL patients, however, mNGS results of sonicate fluid was positive in 1 AL case. Cultures and mNGS results were negative in all three pairs of negative-control samples. In all 70 samples, mNGS detected 24 pathogens in sonicate fluids and 22 pathogens in synovial fluids. There was no significant difference in number of raw reads and human reads ratio between mNGS of sonicate fluid and synovial fluid. mNGS of sonicate fluid generated significantly higher number of microbial reads and of stringently mapped reads of pathogen in species-level than that of synovial fluids. There was no significant difference in diagnostic sensitivity of PJI between mNGS of sonicate fluids and synovial fluids (90.0% vs 100.0%). Both of them were significantly higher than that of culture of synovial fluid, periprosthetic tissues. Diagnostic sensitivity of sonicate fluid mNGS was not significantly higher than that in culture of sonicate fluid (65%). The specificities were similar among various microbiological testing methods.@*Conclusion@#mNGS of either synovial fluid or sonicate fluid from patients who underwent revision arthroplasty can be used to detect the presence of pathogens effectively and diagnose PJI accurately. mNGS can identify more pathogens and generate a higher number of pathogenic reads from sonicate fluids than synovial fluid. mNGS of synovial fluids has met the clinical diagnostic demands for most PJI patients. mNGS of sonicate fluid could be applied in some cases.

Effects of ultrasound combined with microbubbles on intracellular Ca 2+ homeostasis in carboplatin-treated microenvironment of A549 cell line / 中华超声影像学杂志

Objective To investigate the effects of ultrasound combined with microbubbles on intracellular Ca2+ homeostasis in carboplatin ( CBP )-treated A549 cell and its possible mechanisms of inhibiting A549 cell line activity . Methods According to whether SonoVue was used or not ,and the different dose of CBP ,the groups A-F were arranged as the ultrasound(US) group(group A) ,the ultrasound combined with microbubbles ( USMB) group( group B) ,the low dose CBP ( 100 μg/ml) + US group( group C) ,the low dose CBP+USMB group( group D) ,the high dose CBP ( 200 μg/ml)+ US group ( group E) and the high dose CBP+USMB group( group F) .A549 cells were bathed and washed by a calcium-free buffer , loaded with Ca2+ indicator fluo-4 AM . Real-time images were acquired using laser confocal microscopy .The fluorescence intensity of intracellular calcium ion concentration ([Ca2+ ]i) in individual living cell was observed and the calcium overload was analyzed . Results After ultrasound irradiation ,the normalized fluorescence intensity of [Ca2+ ]i increased rapidly ,then returned to a new homeostasis (selected cells in groups A ,B ,E ,F) or experienced a second calcium oscillation ( some cells in group C and D) . All the selected cells in group B and some cells in group C and D exhibited superimposed oscillations . The calcium overloading time in group D was longer than those of any other groups . Four cells in group A experienced delayed calcium oscillations . Compared with group A ,the selected cells in other groups exhibited a larger amplitude of calcium oscillation( all P < 0 .05) and the selected cells in group B and D exhibited calcium oscillation for a longer period of time( all P <0 .05) . Conclusions In the calcium-free buffer ,US ,USMB , CBP+ US ,CBP + USMB are direct stimuli of calcium overload in A 549 cells . SonoVue ,CBP ,CBP +SonoVue are all synergistic stimuli of calcium overload in A 549 cells irradiated by ultrasound .US ,USMB and CBP may synergistically induce calcium release from intracellular store sites in A 549 cells . Calcium overload is a possible mechanism of ultrasound combined with microbubbles in assisting CBP chemotherapy .

Development of carrier-free nanocrystals of poorly water-soluble drugs by exploring metastable zone of nucleation

There has been increasing interest in research and development of nanocrystals for the delivery of poorly water-soluble drugs that can be directly produced from solution. Compared with traditional carrier-based or encapsulation designs, drug nanocrystals circumvent possible side-effects due to carrier polymers and poor stability issues associated with encapsulation. The production of carrier-free nanocrystals requires careful control of nucleation and thus a thorough understanding of the relevant solution's metastable zone. A solution may stay supersaturated without forming any nuclei and become metastable. The maximal degree of supersaturation is known as the metastable zone width. When nucleation is triggered directly from the metastable zone, it helps to produce homogeneous nuclei leading to uniform nanocrystals. Herein, we report a study in which the solubility and metastable limit of paclitaxel (PTX) in ethanol aqueous solution were measured at 40 °C. A wide range of metastable compositions were studied to prepare carrier-free PTX nanocrystals with particle size smaller than 250 nm and PDI less than 0.25. Compared with the raw material, dissolution rate of PTX nanocrystals was significantly increased. The study enables production of high-quality drug nanocrystals for treating patients.

Effects of ultrasound combined with microbubbles on intracellular Ca 2+ homeostasis in carboplatin‐treated microenvironment of A549 cell line / 中华超声影像学杂志

To investigate the effects of ultrasound combined with microbubbles on intracellular Ca2+ homeostasis in carboplatin ( CBP )‐treated A549 cell and its possible mechanisms of inhibiting A549 cell line activity . Methods According to whether SonoVue was used or not ,and the different dose of CBP ,the groups A‐F were arranged as the ultrasound( US) group( group A ) ,the ultrasound combined with microbubbles ( USMB) group( group B) ,the low dose CBP ( 100 μg/ml) + US group( group C) ,the low dose CBP+USMB group( group D) ,the high dose CBP ( 200 μg/ml)+ US group ( group E) and the high dose CBP+USMB group( group F) .A549 cells were bathed and washed by a calcium‐free buffer , loaded with Ca2+ indicator fluo‐4 AM . Real‐time images were acquired using laser confocal microscopy . T he fluorescence intensity of intracellular calcium ion concentration ( [ Ca 2+ ] i ) in individual living cell was observed and the calcium overload was analyzed . Results After ultrasound irradiation ,the normalized fluorescence intensity of [ Ca2+ ] i increased rapidly ,then returned to a new homeostasis ( selected cells in groups A ,B ,E ,F ) or experienced a second calcium oscillation ( some cells in group C and D ) . All the selected cells in group B and some cells in group C and D exhibited superimposed oscillations . T he calcium overloading time in group D was longer than those of any other groups . Four cells in group A experienced delayed calcium oscillations . Compared with group A ,the selected cells in other groups exhibited a larger amplitude of calcium oscillation ( all P ＜ 0 .05 ) and the selected cells in group B and D exhibited calcium oscillation for a longer period of time( all P ＜0 .05) . Conclusions In the calcium‐free buffer ,US ,USMB , CBP+ US ,CBP + USMB are direct stimuli of calcium overload in A 549 cells . SonoVue ,CBP ,CBP +SonoVue are all synergistic stimuli of calcium overload in A 549 cells irradiated by ultrasound .US ,USMB and CBP may synergistically induce calcium release from intracellular store sites in A 549 cells . Calcium overload is a possible mechanism of ultrasound combined with microbubbles in assisting CBP chemotherapy .