Rapid and visual detection of Group B streptococcus using recombinase polymerase amplification combined with lateral flow strips

Conventional culture method for detecting Group B streptococcus (GBS), a common pathogen of neonatal meningitis and sepsis, is time-consuming and unsensitive. Even though real-time fluorescence PCR-based molecular method is more accurate, it need special instrument and elaborate protocol. Here, we established a novel molecular method combining recombinase polymerase amplification with lateral flow strips for detecting GBS. The cAMP factor (cfb) gene is a highly specific and sensitive biomarker to identify GBS and is detectable by using 100 genomic copies as the amplification template. Clinical performance of this assay was evaluated by testing 130 samples, in comparison with culture method and real-time fluorescence PCR, and the results achieved 100% accuracy, which were the same with those of real-time fluorescence PCR, and were better than those of culture method with false-negative detection. This study provides a rapid and visual method, with clinical potential, for the detection of GBS infection of patients.     

Fabrication of biocompatible magneto-fluorescence nanoparticles as a platform for fluorescent sensor and magnetic hyperthermia applications

Multifunctional nanoparticles with special magnetic and optical properties have been attracting a great deal of attention due to their important applications in the bioanalytical and biomedical fields. In this study, we report the fabrication of biocompatible magneto-fluorescence nanoparticles consisting of carbon dots (CDots) and silica-coated cobalt–manganese nanoferrites (Co_0.5Mn_0.5Fe₂O₄) (CoMnF@Si@CDots) (MagSiCDots) by a facile hydrothermal method. The as-prepared MagSiCDots have a particle size of 100–120 nm and show a negative zeta potential of −35.50 mV at a neutral pH. The fluorescence spectrum of the MagSiCDots nanoparticles consists of sharp excitation at 365 nm and broad blue light emission with a maximum wavelength of 442.5 nm and the MagSiCDots exhibit superparamagnetic behaviour with a saturation magnetization of 11.6 emu g⁻¹. The potential of MagSiCDots as a fluorescent sensor and be used for magnetic hyperthermia applications. It is seen that the fluorescent intensity of a colloidal solution (a hydrogen sulfide (H₂S) solution containing MagSiCDots nanoparticles) has a linear relationship with the H₂S concentration range of 0.2–2 μM. The limit of detection (LOD) of H₂S by our MagSiCDots particles is 0.26 μM and they remain stable for at least 90 min. To test the suitability of the MagSiCDots nanoparticles for use in hyperthermia application, induction heating using an AMF was done. It was observed that these nanoparticles had a specific absorption rate (SAR) of 28.25 W g⁻¹. The in vitro and in vivo cytotoxicity of MagSiCDots were tested on HeLa cells lines. The results show a cell viability of about 85% when exposed to 100 μg mL⁻¹ concentration of the particles. The in vivo cytotoxicity using zebrafish assay also confirmed the non-toxicity and biocompatibility of the nanoparticles to living cells. The reported data demonstrate that by combining CoMnF@Si and fluorescent CDots into a single system, not only nontoxic multifunctional nanomaterials but also multimodal nanoparticles for several applications, such as hazard gas detection and acting as a biocompatible heat source for therapeutic treatment of cancer, are provided.

Multifunctional nanoparticles with special magnetic and optical properties have been attracting a great deal of attention due to their important applications in the bioanalytical and biomedical fields.     

Analytical and clinical evaluation of a point-of-care molecular diagnostic system and its influenza A/B assay for rapid molecular detection of the influenza virus

Recently, rapid molecular detection systems have been used for point-of-care testing for the diagnosis of influenza worldwide. Here, we evaluated the performance of the cobas Liat system and the cobas Influenza A/B assay (Liat) using fresh nasopharyngeal samples collected from a Japanese population between December 2017 and February 2018. The performance of the examination was compared with that of antigen testing and a conventional polymerase chain reaction (nested-PCR) method. A total of 159 patients were included in this study, and 77 tested positive using Liat. The concordance rate between Liat and nested PCR was 97.5%. The median time between the ordering of testing and completion of molecular analyses using Liat was 30 min (interquartile range: 28–35 min). The overall sensitivity and specificity of antigen testing were 57.1% and 100%, respectively. The duration from symptom onset to examination did not alter antigen testing sensitivity. The current study demonstrates the high performance of Liat for the rapid molecular identification of the influenza virus.     

The biopsychosocial model and its potential for a new theory of homeopathy

Since the nineteenth century the theory of conventional medicine has been developed in close alignment with the mechanistic paradigm of natural sciences. Only in the twentieth century occasional attempts were made to (re)introduce the 'subject' into medical theory, as by Thure von Uexküll (1908-2004) who elaborated the so-called biopsychosocial model of the human being, trying to understand the patient as a unit of organic, mental, and social dimensions of life. Although widely neglected by conventional medicine, it is one of the most coherent, significant, and up-to-date models of medicine at present. Being torn between strict adherence to Hahnemann's original conceptualization and alienation caused by contemporary scientific criticism, homeopathy today still lacks a generally accepted, consistent, and definitive theory which would explain in scientific terms its strength, peculiarity, and principles without relapsing into biomedical reductionism. The biopsychosocial model of the human being implies great potential for a new theory of homeopathy, as may be demonstrated with some typical examples.     

Oleogelation using pulse protein-stabilized foams and their potential as a baking ingredient

Structuring liquid oil into a self-standing semisolid material without trans and saturated fat has become a challenge for the food industry after the recent ban of trans fat by the US Food and Drug Administration and Health Canada. Lately, the use of hydrocolloids such as animal proteins and modified cellulose for oleogel preparation has gained more attention. However, plant proteins have never been explored for the development of oleogels. The present study explored the use of freeze-dried foams prepared using protein concentrates and isolates of pea and faba bean with xanthan gum at different pH values for oil adsorption and subsequent oleogelation. Compared to protein isolate stabilized foams, protein concentrate-stabilized foams displayed (i) higher oil binding capacity (OBC) due to a higher number of smaller pore size; and (ii) lower storage modulus and firmness due to the higher oil content. At all pH values, there was no significant difference between the OBC of different protein isolates, but among the concentrates, pea displayed higher OBC than faba bean at pH 5 and faba bean displayed higher OBC than pea at pH 9. Results showed that such oleogels could be used as a shortening alternative. Cakes prepared using the pea protein-based oleogel at pH 9 displayed a similar specific volume as that of shortening-based cake, although with higher hardness and chewiness.

Canola oil was structured into oleogels using freeze-dried foam made with pea or faba bean protein concentrates or isolates and xanthan gum at pH 5, 7 and 9. The oleogels were used to bake cakes and compared with conventional shortening-based cakes.     

A new chromogenic endotoxin-specific assay using recombined limulus coagulation enzymes and its clinical applications

A conventional limulus test is not specific to endotoxin because of the presence in amebocyte lysate of a (1----3)-beta-D-glucan-sensitive factor. By fractionating coagulation enzymes in the lysate and recombining only those factors involved in endotoxin-induced coagulation, we have developed a new test specific to endotoxin. The recombined enzymes reacted only with endotoxin, and not with fungal polysaccharides. Conventional amebocyte lysate, on the other hand, reacted with both of them. A good linearity was obtained with this method between endotoxin concentration and absorbance with a sensitivity of 1 pg/ml of Escherichia coli 0111:B4 endotoxin. The regression lines for different types of endotoxins were parallel to one another. For the correct diagnosis of endotoxemia, this new test has a definite advantage over the one using whole amebocyte lysate.     

A novel assay for cobalt-albumin binding and its potential as a marker for myocardial ischemia-a preliminary report

We initially observed a phenomenon of reduced in vitro binding of exogenous cobalt [Co(II)] to the N-terminus of human serum albumin (HSA) in emergency chest pain patients with early onset unstable angina and myocardial infarction. We then developed a colorimetric assay to measure cobalt-HSA binding and record the results in absorbance units (ABSU). In a preliminary clinical study of 139 emergency patients with acute chest pain, 99 patients with evidence of myocardial ischemia (Group 1) had elevated assay levels (mean ABSU ± SD; 0.519 ± 0.086) compared to 40 patients (Group 2) with no evidence of ischemia (0.316 ± 0.092) (p < 0.00001). In Group 1, 95 of 99 (96.0%) patients had levels higher than a decision threshold of 0.400 ABSU and in Group 2, 37 of 40 (92.5%) samples had higher cobalt binding capacity (ABSU ≤ 0.400). Further studies are warranted to determine if an assay measuring altered cobalt-HSA binding is a clinically useful diagnostic test to rule out myocardial ischemia.     

Impairment of the hemostatic potential of platelets during storage as evaluated by flow cytometry, thrombin generation, and thrombelastography under conditions promoting formation of coated platelets

BACKGROUND: The increasing demand for platelet (PLT) transfusions has focused attention on appropriate use. Coated PLTs are a subpopulation of highly procoagulant PLTs formed by simultaneous stimulation by the agonist's collagen and thrombin hypothesized to drive clot formation at the site of vascular injury. Prolonged storage of PLTs may reduce their ability to support optimal hemostasis upon transfusion. STUDY DESIGN AND METHODS: PLT concentrates (PCs) stored for 1, 4, 6, and 8 days were costimulated with thrombin and the collagen glycoprotein VI (GPVI) receptor agonist convulxin, and their ability to form coated PLTs was determined by flow cytometry. Further, a plasma-based thrombin generation assay and thrombelastography were used to evaluate the aged PCs' capacity to support thrombin generation and clot formation, respectively. The stored PCs were additionally tested by standard quality control methods. RESULTS: PLT quality as measured by standard analyses was acceptable according to current practice. The hemostatic potential, however, was impaired with increasing storage time. The formation of coated PLTs decreased significantly from approximately 85 to 55 percent with increasing storage time (p<0.05). The velocity of clot formation was significantly increased from Day 4 (p<0.05). The velocity of thrombin generation and resistance against fibrinolysis were significantly reduced on Day 8 compared to Day 1 of storage (p<0.05). CONCLUSION: Data in the present study suggest that storage significantly reduced the stored PLTs' ability to respond to conditions expected to exist at the site of vascular injury and that storage-induced reduction in PLT activation sensitivity correlated with a loss of hemostatic potential.     

Thermometric enzyme linked immunosorbent assay in continuous flow system: optimization and evaluation using human serum albumin as a model system.

Thermometric enzyme-linked immunosorbent assay (TELISA) is described. After the procedure of optimization, human serum albumin was assayed using anti-human serum albumin bound to Sepharose CL 4-B in the enzyme thermistor unit and catalase as label on the free antigen. The model system was used for assays down to 10(-13)M and the preparation of immobilized antibodies was used repeatedly up to 100 times. Comparative studies of the TELISA technique with bromocresol green, immunoturbidimetric and rocket immunoelectrophoretic methods were carried out and showed that TELISA could be used as an alternative method.     

In vitro characterization of A-315675, a highly potent inhibitor of A and B strain influenza virus neuraminidases and influenza virus replication

A-315675 is a novel, pyrrolidine-based compound that was evaluated in this study for its ability to inhibit A and B strain influenza virus neuraminidases in enzyme assays and influenza virus replication in cell culture. A-315675 effectively inhibited influenza A N1, N2, and N9 and B strain neuraminidases with inhibitor constant (K(i)) values between 0.024 and 0.31 nM. These values were comparable to or lower than the K(i) values measured for oseltamivir carboxylate (GS4071), zanamivir, and BCX-1812, except for the N1 enzymes that were found to be the most sensitive to BCX-1812. The time-dependent inhibition of neuraminidase catalytic activity observed with A-315675 is likely due to its very low rate of dissociation from the active site of neuraminidase. The half times for dissociation of A-315675 from B/Memphis/3/89 and A/Tokyo/3/67 (H3N2) influenza virus neuraminidases of 10 to 12 h are significantly slower than the half times measured for oseltamivir carboxylate (33 to 60 min). A-315675 inhibited the replication of several laboratory strains of influenza virus in cell culture with potencies that were comparable or superior to those for oseltamivir carboxylate and BCX-1812, except for the A/H1N1 viruses that were found to be two- to fourfold more susceptible to BCX-1812. A-315675 and oseltamivir carboxylate exhibited comparable potencies against a panel of A/H1N1 and A/H3N2 influenza virus clinical isolates, but A-315675 was found to be significantly more potent than oseltamivir carboxylate against the B strain isolates. The favorable in vitro results relative to other clinically effective agents provide strong support for the further investigation of A-315675 as a potential therapy for influenza virus infections.     

彩色多普勒超声评价肾缺血再灌注后模型兔肾动脉血流动力学变化及与肾功能的关系

背景:既往对肾缺血再灌注损伤诊断主要依靠实验室生化检查及病理检查,彩色多普勒超声仪能无创快速动态显示肾缺血再灌注后肾血流的变化.目的:运用彩色多普勒超声评价兔肾缺血再灌注后肾动脉血流动力学变化,并探讨其与肾功能指标血尿素氮和血肌酐变化之间的相关性.方法:随机数字表法将大白兔分为对照组、假手术组、缺血再灌注组.缺血再灌注组肾缺血再灌注模型;假手术组仅行右肾切除;对照组不作手术处理.前两组按照不同时间点再分为3个亚组进行观察.结果与结论:随着兔肾缺血再灌注损伤加重,各级肾动脉收缩期峰值流速、搏动指数、阻力指数及血清尿素氮和血肌酐逐渐增大,24 h达高峰,叶间动脉最先出现改变血流动力学改变,其中阻力指数是反映肾缺血再灌注损伤程度最敏感指标.提示彩色多普勒超声是一种评价肾缺血再灌注损伤程度的有效方法.     

Metabolomics-driven identification of perturbations in amino acid and sphingolipid metabolism as therapeutic targets in a rat model of anorexia nervosa disease using chemometric analysis and a multivariate analysis platform

It is important to explore novel therapeutic targets and develop an effective strategy for the treatment of anorexia nervosa. In this work, serum samples were analyzed using ultra-performance liquid chromatography coupled with quadrupole time-of flight mass spectrometry (UPLC/Q-TOF MS) coupled with chemometric analysis and multivariate analysis to obtain the metabolites and their corresponding pathways. In addition, knock-in and knock-down of the key enzyme in vivo was performed to verify the reliability of the obtained metabolic pathway, which is closely associated with the anorexia nervosa pathomechanism and the potential targets. There were significant differences in the biochemical parameters between the model group and the control group. A total of 26 potential biomarkers were identified to resolve the difference between the control and model rats, which were closely related to amino acid metabolism, sphingolipid metabolism, arachidonic acid metabolism, the citrate cycle, and so forth. According to the ingenuity pathway analysis, we further elucidated the relationship between the gene, protein, and metabolite alteration in anorexia nervosa, which are involved in cellular compromise, lipid metabolism, small molecule biochemistry, cell signaling, molecular transport, nucleic acid metabolism, cell morphology, cellular function and maintenance. Arginosuccinate synthetase (ASS) deficiency was accompanied by a significant downregulation of the β-endorphin and ghrelin in the animal models. The metabolites and pathways obtained using the metabolomics strategy may provide valuable information for the early treatment for anorexia nervosa.

It is important to explore novel therapeutic targets and develop an effective strategy for the treatment of anorexia nervosa.     

In vivo toxicity evaluation of two polyoxotungstates with potential antidiabetic activity using Wistar rats as a model system

In this study, the in vivo hypoglycemic effect of a donut-shaped polyanion salt (NH₄)₁₄[Na@P₅W₃₀O₁₁₀]·31H₂O {NaP₅W₃₀} and its Ag-containing derivative K₁₄[Ag@P₅W₃₀O₁₁₀]·22H₂O·6KCl {AgP₅W₃₀}, as well as their hepatotoxicity and nephrotoxicity, was evaluated. In the screening hypoglycemic study, Wistar albino rats with streptozotocin induced diabetes were treated intraperitoneally with three single doses (5, 10, and 20 mg per kg per b.w.) of both investigated polyoxotungstates. The blood glucose levels, measured before and after 2, 4 and 6 h polyoxotungstate application, showed that both studied compounds induced the most pronounced and time dependent glucose lowering effects at the doses of 20 mg kg⁻¹. Thus, daily doses of 20 mg kg⁻¹ were administered to Wistar albino rats orally for 14 days in further toxicity examinations. The serum glucose concentration and biochemical parameters of kidney and liver function, as well as a histopathological analysis of kidney and liver tissues were evaluated 14 days after the polyoxotungstate administration. Both investigated compounds did not induce statistically significant alterations of the serum glucose and uric acid concentrations, as well as some of the liver function markers (serum alanine and aspartate aminotransferases, and alkaline phosphatase activities). However, the significant decrease in serum total protein and albumin concentrations and the increase in biochemical parameters of renal function – serum urea (up to 63.1%) and creatinine concentrations (up to 23.3%) were observed for both polyoxotungstates. In addition, the detected biochemical changes were in accordance with kidney and liver histhopathological analysis. Accordingly, the hepatotoxic and nephrotoxic effects of these potential antidiabetic polyoxotungstates could be considered as mild.

Study of the in vivo hypoglycemic effect, hepatotoxicity and nephrotoxicity of a donut-shaped polyanion salt (NH₄)₁₄[Na@P₅W₃₀O₁₁₀]·31H₂O {NaP₅W₃₀} and its Ag-containing derivative K₁₄[Ag@P₅W₃₀O₁₁₀]·22H₂O·6KCl {AgP₅W₃₀}.     

Generation and characterization of a human bradykinin receptor B1 transgenic rat as a pharmacodynamic model

Antagonists of the B1 bradykinin receptor (B1R) offer the promise of novel therapeutic agents for the treatment of inflammatory and neuropathic pain. However, the in vivo characterization of the pharmacodynamics of B1R antagonists is hindered by the low level of B1R expression in healthy tissue and the profound species selectivity exhibited by many compounds for the human B1R. To circumvent these issues, we generated a transgenic rat expressing the human B1R under the control of the neuron-specific enolase promoter. Membranes prepared from whole brain homogenates of heterozygous transgenic rats indicate a B1R expression level of 30 to 40 fmol/mg; there is no detectable B1R expression in control nontransgenic rats. The pharmacological profile of the B1R expressed in the transgenic rat matches that expected of the human, but not the rat receptor. The mapping of the transgene insertion site to rat chromosome 1 permitted the development of a reliable assay for the identification of homozygous transgenic rats. Significantly, homozygous transgenic rats express 2-fold more B1R than heterozygous animals. Autoradiographic analyses of tissue sections from transgenic rats reveal that the B1R is broadly expressed in both the brain and spinal cord. The human B1R expressed in the transgenic rat functions in an in vitro contractile assay and thus has the potential to elicit a functional response in vivo. Using the humanized B1R transgenic rat, an assay was developed that is suitable for the routine evaluation of a test compound's ability to occupy the human B1R in the central nervous system.     

Biofabrication of a novel leukocyte‐fibrin‐platelet membrane as a cells and growth factors delivery platform for tissue engineering applications

Autologous platelet‐rich hemocomponents have emerged as potential biologic tools for regenerative purpose, but their therapeutic efficacy still remains controversial. This work represents the characterization study of an innovative autologous leukocyte‐fibrin‐platelet membrane (LFPm), which we prepared according to a novel protocol involving multiple cycles of apheresis. The high content in fibrinogen gave to our hemocomponent the appearance of a manipulable and suturable membrane with high elasticity and deformation capacity. Moreover, being highly enriched with platelets, leukocytes, and monocytes/macrophages, the LFPm sustained the local release of bioactive molecules (platelet derived growth factor, vascular endothelial growth factor, interleukin‐10, and tumour necrosis factor alpha). In parallel, the evaluation of stemness potential highlighted also that the LFPm contained cells expressing pluripotency and multipotency markers both at the messenger ribonucleic acid (NANOG, SOX2, THY1, NT5E, and ENG) and surface‐protein level (CD44^high/CD73⁺/CD34⁺/CD117⁺/CD31⁺). Finally, biodegradation analysis interestingly showed a good stability of the membrane for at least 3 weeks in vitro and 1 week in vivo. In both cases, biodegradation was associated with progressive exposure of fibrin scaffold, loss/migration of cellular elements, and release of growth factors. Overall, collected evidence could shed some light on the regenerative effect that LFPms may exert after the autologous implant on a defect site.