High sensitivity flow cytometry of membrane vesicles.

Extracellular vesicles (EVs) are attracting attention as vehicles for inter-cellular signaling that may have value as diagnostic or therapeutic targets. EVs are released by many cell types and by different mechanisms, resulting in phenotypic heterogeneity that makes them a challenge to study. Flow cytometry is a popular tool for characterizing heterogeneous mixtures of particles such as cell types within blood, but the small size of EVs makes them difficult to measure using conventional flow cytometry. To address this limitation, a high sensitivity flow cytometer was constructed and EV measurement approaches that allowed them to enumerate and estimate the size of individual EVs, as well as measure the presence of surface markers to identify phenotypic subsets of EVs. Several fluorescent membrane probes were evaluated and it was found that the voltage sensing dye di-8-ANEPPS could produce vesicle fluorescence in proportion to vesicle surface area, allowing for accurate measurements of EV number and size. Fluorescence-labeled annexin V and anti-CD61 antibody was used to measure the abundance of these surface markers on EVs in rat plasma. It was shown that treatment of platelet rich plasma with calcium ionophore resulted in an increase in the fraction of annexin V and CD61-positive EVs. Vesicle flow cytometry using fluorescence-based detection of EVs has the potential to realize the potential of cell-derived membrane vesicles as functional biomarkers for a variety of applications.

Homeostasis of human NK cells is not IL-15 dependent.

IL-15 is a proinflammatory cytokine that plays an important role in the development and activation of NK cells and is a potential target for inflammatory disease therapy. Studies conducted in IL-15- and IL-15R knockout mice identified IL-15 as an important cytokine for NK cell homeostasis. Consistent with this information derived from genetically modified mice, we demonstrated that neutralizing IL-15 with a mouse anti-mouse IL-15 mAb (M96) depletes C57BL/6 mouse NK cells. An mAb directed against macaque IL-15 (Hu714MuXHu) was manufactured and demonstrated to block IL-15-induced activation of nonhuman primate (NHP) NK cells in vitro. Neutralization of macaque IL-15 by parenteral administration of Hu714MuXHu reduces (>95%) circulating NK cell counts in NHPs. A blocking mAb directed against human IL-15 (huIL-15; AMG 714) was manufactured. Unexpectedly, when human subjects were treated with the blocking anti-IL-15 Ab AMG 714 in clinical trials, no reductions in circulating NK cell counts were observed despite achieving significantly higher exposures than the levels of Hu714MuXHu needed to cause NK cell count reductions in NHPs in vivo. Both AMG 714 and Hu714MuXHu are able to block huIL-15 activity in a human T cell blast proliferation and IFN-γ production assay. Both Abs block huIL-15-mediated Stat5 activation and CD69 expression in human NK cells. Collectively, these results demonstrate that NK cell homeostasis is obligatorily dependent upon IL-15 in both mice and NHPs, but that IL-15 is dispensable for maintenance of circulating human NK cells.

Evaluation of Visual Evoked Potential (VEP) in Patients With Chronic Obstructive Pulmonary Disease (COPD).

Chronic Obstructive Pulmonary Disease (COPD), a progressive and partially reversible disease, has drawn world-wide attention for its moderate prevalence rate and causing central and peripheral neuropathy. Considering its severity in causing visual pathway impairment, the present investigation was carried out to find out the functional integrity of the visual pathway through visual evoked potentials (VEP) and to determine the factors influencing the condition in COPD patients. A total of 30 COPD patients of both sexes, classified according to the severity of the disease based on spirometric indices, were subjected to VEP testing and series of wave forms were measured and compared with equal number of control subjects. The latency of N75 and P100 were prolonged (P < 0.01) and the P100 amplitude was reduced significantly (P < 0.01) in COPD patients. Thus, the influence of COPD causing the optic neuropathy is apparent from the significant VEP changes. Non-invasive procedure can possibly be utilized as a routine screening test for COPD patients for better medical care.

Assessment of the performance of three multiplex array panels for the detection of circulating cytokines and chemokines in naive, LPS, and SEB-treated cynomolgus macaques.

To assess relative sensitivity for detection of cytokines and chemokines in cynomolgus serum samples, we tested three commercially available multiplex array kits using the Luminex® platform with sera from animals exposed by intravenous injection to 150 µg/kg staphylococcal enterotoxin B (SEB) or 20 µg/kg lipopolysaccharide (LPS). Each of these kits detected similar patterns of changes in circulating cytokines/chemokines in response to SEB or LPS stimulation, especially the induction of high amounts of interleukin (IL)-2 and interferon-gamma (IFN-γ) in response to SEB but not LPS. However, there were clear differences in sensitivity for particular analytes, especially for IL-10. Additional experiments that focused on one multiplex array kit demonstrated very low or undetectable levels of cytokines in naive cynomolgus macaques, except for highly variable background levels of IL-8, monocyte chemoattractant protein-1, and macrophage inflammatory protein-1ß. Therefore, multiplex array analysis of circulating cytokine/chemokine patterns was capable of detection of systemic activation of diverse immune cell subsets.

A Critical Review on Therapeutic Potential of Benzimidazole Derivatives: A Privileged Scaffold.

Benzimidazole nucleus is a predominant heterocycle displaying a wide spectrum of pharmacological activities. The privileged nature of the benzimidazole scaffold has been revealed by its presence in most small molecule drugs and in its ability to bind multiple receptors with high affinity. A literature review of the scaffold reveals several instances where structural modifications of the benzimidazole core have resulted in high-affinity lead compounds against a variety of biological targets. Hence, this structural moiety offers opportunities to discover novel, better, safe and highly potent biological agents. The goal of the present review is to compile the medicinal properties of benzimidazole derivatives with a focus on SAR (Structure-Activity Relationships).

Clinical effects of Bifidobacterium as a probiotic on oral health: A systematic review.

Background: The burden of oral diseases is increasing, which constitute a major public health problem. The use of probiotics as an adjuvant, along with routine dental care practice by an individual, can produce additional benefits in the maintenance of one's oral health. The study aimed to investigate the effect of Bifidobacterium as a probiotic on oral health. Material and Methods: Six databases and registers were searched from the start of the database to December 2021 without any restrictions. Randomized controlled trials (RCTs) evaluating the clinical effects of Bifidobacterium as a probiotic on oral health were included in the study. The Preferred Reporting Items for Systematic Reviews and Meta-analyses (PRISMA) guidelines were followed to conduct this systematic review. The included studies were analyzed for the risk of bias using the Cochrane risk-of-bias tool for randomized trial (RoB 2) tool as well as quality of available evidence using GRADE criteria. Results: From the 22 qualified studies, four studies showed non-significant results. There was a high risk of bias in 13 studies and some concerns of bias in nine studies. No adverse effects were reported, and the quality of available evidence was moderate. Conclusion: The effect of Bifidobacterium on oral health is questionable. Further high-quality RCTs are required on the clinical effects of bifidobacteria and also the optimum level of probiotic needed, and ideal mode of administration to provide oral health benefits. Furthermore, synergistic effects of the combined use of various strains of probiotics need to be studied.

Determination of absolute counts of circulating regulatory T cells in cynomolgus macaques using an optimized flow cytometric method.

Regulatory T cells (Tregs) are a rare subset of lymphocytes that inhibit the activation and effector functions of T cells and are important regulators of immune responses. Although Tregs are well characterized in humans and rodents, little is known about their immunophenotyping (IP) profile in cynomolgus macaques (Macaca fascicularis), which is an important species for pharmacological and toxicological evaluation of potential immune modulators because of their similar physiologic, genetic, and metabolic response patterns to humans. The authors have developed an immunophenotyping panel using a high-throughput 96-well microtiter plate-based assay to detect circulating Tregs (CD3(+)CD4(+)CD25(hi)FoxP3(+)) and have determined the normal range for the number of Tregs in naive healthy cynomolgus macaques to be 56.4 to 179.7 cells/µL (mean ± SEM = 113.6 ± 5.1 cells/µL; n = 25). Furthermore, the authors compared the resulting FoxP3(+) Treg profiles with a CD127(lo) cell-surface panel (CD3(+)CD4(+)CD25(hi) CD127(lo)) and found a close correlation between the absolute numbers of CD3(+)CD4(+)CD25(hi)FoxP3(+) and CD3(+)CD4(+)CD25(hi)CD127(lo) cells (mean ± SD = 120 ± 8.0 cells/µL). Quantification of circulating Tregs in cynomolgus macaques in this high-throughput assay may help to identify drug candidates that affect this rare, but critical, immunoregulatory cell population.

Relationship between dental anxiety with dental caries and periodontal disease among army recruits in Bangalore city - A cross sectional study.

Introduction: Dental anxiety can affect the dental health status of an individual leading to avoidance behavior, poor oral hygiene and periodontal health and delay in seeking necessary treatment. Objectives: This study aimed to assess the prevalence of dental anxiety in army recruits in Bangalore city and to explore the relationship of dental anxiety with dental caries and periodontal disease among army recruits in Bangalore city. Methodology: The total population of the army recruits in Army Service Corps Center and College was around 1000. General information included name, age, sex educational qualification, address, number of dental visits and age of first visit. The closed ended multiple-choice questionnaire consisting of 4 questions based on previous dental experience given by Corah was used to assess the anxiety level of the individual. Clinical examination of dental and periodontal health was noted using Decayed, Missing, Filled and Surface (DMFS) Index and Community Periodontal Index and Treatment Needs (CPITN) Index. Results: The mean Dental Anxiety Scale score was 9.64 with a standard deviation of 3.6. Of the 836 participants in the study, 444 (53.1%) reported no dental anxiety at all. Two hundred and sixty-three (31.5%) were moderately dental anxious (scoring 9-12), 10.4% were highly anxious (scoring 13-14) and 5% were severely anxious (scoring 15-20). Anxious individuals had poorer periodontal health necessitating treatment compared to nonanxious individuals. The mean DMFS of the study population was highest among those having high and severe dental anxiety (mean DMFS = 11.0 and 12.59, respectively) which was statistically significant (P < 0.001). Conclusion: The present study showed a prevalence of dental anxiety of 47%. Impact of dental anxiety on oral health was found to be obvious as higher DMFT/DMFS was evident among dentally anxious subjects; dental anxiety also showed a strong association with higher CPITN scores.

Assessment of Oral Health Care-Related Expenditure among People of Kerala: A Cross-Sectional Study.

Background: Oral health is an effective unit of general health. Dental caries and periodontal diseases are considered to be the major diseases present in modern times. Over the decades, the idea of maintaining effective oral health and interest in oral health prevention measures has diminished; thus increasing the cost of health care. Aims and Objectives: This study aims to estimate the household expenditure on oral health care among people residing in Kerala, India. Materials and Methods: Five hundred participants were surveyed who were residents of Kerala. A self-administered questionnaire that was tested and validated was used to assess the cost of oral health care. Data collected were analyzed using software for IBM SPSS version 23 for Windows (New York, USA). Results: Of 500 people, 37% are men and 63% are women. 100% of people reported using a toothbrush as an oral hygiene aid. 65% of households reported changing toothbrush for 3 months or more, while 35% of families changed for 1 month or less. Significant statistically weak correlation was observed when education, employment, and income were compared to the annual expenditure on dental care. Furthermore, there has been a moderately strong correlation observed between the socio-economic status of families and the annual cost of dental care. Conclusion: People should be aware of preventive oral hygiene aids, and appropriate policies should be formulated which will ultimately result in decreased expenditure on a curative aspect of the dental disease.

In situ multi-satellite detection of coherent vortices as a manifestation of Alfvénic turbulence.

Turbulence in fluids and plasmas is a ubiquitous phenomenon driven by a variety of sources-currents, sheared flows, gradients in density and temperature, and so on. Turbulence involves fluctuations of physical properties on many different scales, which interact nonlinearly to produce self-organized structures in the form of vortices. Vortex motion in fluids and magnetized plasmas is typically governed by nonlinear equations, examples of which include the Navier-Stokes equation, the Charney-Hasegawa-Mima equations and their numerous generalizations. These nonlinear equations admit solutions in the form of different types of vortices that are frequently observed in a variety of contexts: in atmospheres, in oceans and planetary systems, in the heliosphere, in the Earth's ionosphere and magnetosphere, and in laboratory plasma experiments. Here we report the discovery by the Cluster satellites of a distinct class of vortex motion-short-scale drift-kinetic Alfvén (DKA) vortices-in the Earth's magnetospheric cusp region. As is the case for the larger Kelvin-Helmholtz vortices observed previously, these dynamic structures should provide a channel for transporting plasma particles and energy through the magnetospheric boundary layers.

Dental Caries, Body Mass Index, and Socioeconomic Status among Preschoolers in Private Preschools and Anganwadi Centers in Bengaluru City: A Comparative Study.

AIM: Preschool is the time when deleterious oral habits, caries pattern, and risk factor are established and is the time to intervene and establish healthy trends which can have a lifelong influence. Individuals living in various socioeconomic conditions have an assortment of hazard factors that impact oral well-being. This work was conducted to study the prevalence of dental caries, body mass index (BMI), and socioeconomic status (SES) among preschoolers in private preschools and Anganwadi centers within Bengaluru. MATERIALS AND METHODS: This cross-sectional study was conducted in Anganwadis and preschools in Bengaluru city. About 800 students were selected by stratified cluster sampling technique. Written informed consent was obtained before the start of the study. Study proforma was used for the recording of sociodemographic details, anthropometric measurements, decayed, missing and filled teeth (dmft), and decayed, missing and filled surfaces (dmfs) index scoring. Statistical analysis was done using SPSS version 22.0. RESULTS: The mean dmf (t) score in Anganwadi children was 1.60, whereas in private preschool children mean dmf (t) was 1.16. The mean dmf (s) score in Anganwadi children was 3.05, whereas in private preschool children mean dmf (s) was 1.76. There was a statistically significant difference between the two groups with respect to dmf (s) score (p = 0.01). CONCLUSION: It was found that there is an association between dental caries, BMI, and SES among Anganwadi children when a linear correlation was done. CLINICAL SIGNIFICANCE: Dental caries can cause serious physical and mental problems in children which affects their quality of life. These problems have their root cause from childhood and are related to their BMI and SES. This can be prevented by giving proper health education and dietary advice to parents. HOW TO CITE THIS ARTICLE: Sukumaran N, Sharma V, Bhat PK. Dental Caries, Body Mass Index, and Socioeconomic Status among Preschoolers in Private Preschools and Anganwadi Centers in Bengaluru City: A Comparative Study. Int J Clin Pediatr Dent 2020;13(6):630-634.

Awareness about oral cancer among nonhealth professional students - A cross-sectional study in Bengaluru city.

BACKGROUND AND AIM: Oral cancer is one of the most life-threatening conditions, early diagnosis of which greatly increases the probability of cure and survival rates. Knowledge regarding risk factors and early signs among the nonhealth professionals which help in early detection prevention and of the disease. Therefore, the aim of this study is to assess the awareness about tobacco use among nonhealth professional students in Bangalore city. METHODOLOGY: A cross-sectional descriptive questionnaire study was conducted to assess the knowledge and awareness about oral cancer among 800 nonhealth professional students in Bengaluru city. A self-administered questionnaire containing 26 questions pertaining to awareness, signs, symptoms and risk factors of tobacco use was administered to the students. RESULTS: The results revealed that the majority of the students (55.5%) were aware about oral cancer but have less knowledge in terms of its signs and symptoms of oral cancer. The main source for information pertaining to oral cancer was obtained from mass media, i.e., TV, radio and social media (21.5%). CONCLUSION: The study reveals that there is a need for education and raise awareness about oral cancer among nonhealth professional students.

Cytogenetic and cytokine profile in elderly patients with cytopenia.

In the elderly with cytopenia, the diagnosis of myelodysplastic syndrome (MDS) may be missed. Cytokine levels contribute to the pathology of MDS. Hence, the objectives were to evaluate cytogenetic profile as a prognostic indicator in risk stratification and cytokine levels as a screening tool in patients with cytopenia for diagnosis. Over 2 years (2016-2018), 150 elderly patients were screened. MDS diagnosis was confirmed by morphology. Interleukin-2 (IL-2) and IL-6 levels were assessed in 50 patients, and karyotyping was performed in 20 confirmed cases of MDS. Age-matched healthy controls were used for comparison of cytokine levels. Among 150 patients, 88.6% had anemia, including nutritional anemia (51.2%). MDS diagnosis was confirmed in 35 patients. In 15 patients, unexplained cytopenia (UC) was present. Karyotyping in 20 MDS patients was normal in 15 (75%) patients and revealed a complex karyotype in four (20%) patients and double chromosomal abnormality in one (5%) patient. The Revised International Prognostic Scoring System (IPSS-R) scored 91% in the low-risk group and 9% (n = 3) in the high-risk group; the latter three developed acute myeloid leukemia (AML) and two of them had a 7q deletion. Among the 15 cases of UC, one patient died from refractory anemia. No significant difference in levels of IL-2 and IL-6 were found between MDS and UC patients when compared with healthy controls, as well as between different risk groups and karyotypes. A significant difference in IL-2 levels was found in MDS patients with disease progression and with stable disease. On the basis of the findings, it is suggested that IL-2 levels will help in predicting disease progression.

Effect of Child Development Centre model early stimulation among at risk babies--a randomized controlled trial.

OBJECTIVE: To study the effectiveness of Child Development Centre (CDC) model early stimulation therapy done in the first year of postnatal life, in improving the developmental outcome of at-risk neonates at one and two years of age. DESIGN: Randomized controlled trial. SETTING AND SUBJECTS: The study participants included a consecutive sample of 800 babies discharged alive from the level II nursery of Medical College, Thiruvananthapuram. INTERVENTION: The control group received routine postnatal check-up as per hospital practice. Intervention group in addition received CDC model early stimulation therapy (home-based). RESULTS: The intervention group of babies had a statistically significant higher score for mental developmental index (MDI) and psychomotor developmental index (PDI) at one and two years of age. After adjusting all significant risk factors for development, the babies who had intervention had significantly higher Bayley scores, 5.8 units at one year and 2.8 units at two year, as compared to control babies. CONCLUSION: Early stimulation therapy was effective at one year. The beneficial effect also persisted at two years, without any additional interventions in the second year.

Developmental Evaluation Clinic--CDC experience.

We describe our five year experience of conducting developmental evaluation clinic at CDC. We have also assessed the prevalence of developmental delay (defined as delay in any two areas in Denver Development Screening Test II [DDST II]), documented the possible prenatal, natal, postnatal risk factors for developmental delay and, also identified the pattern of developmental disorders. A total of 2111 children were screened. DDST II results were abnormal for 953 (45.1 %) children. On multivariate analysis, delayed cry at birth, increasing age of the child, presence of feeding problems, assisted delivery, and birth injury were found to be associated with increasingly abnormal DDST II results. Nearly 50 % of referred babies had developmental delay without a specific clinical diagnosis, 13.9 % had speech problems, 9.5 % had neurological problems and 5.2 % had chromosomal anomalies. Birth related events are important risk factors for developmental delay.

Effects of p38 MAP kinase inhibitors on the differentiation and maturation of erythroid progenitors.

In rodents, p38 MAP kinase inhibitors (p38is) induce bone marrow hypocellularity and reduce reticulocyte and erythrocyte counts. To identify target cell populations affected, a differentiating primary liquid erythroid culture system using sca-1(+)cells from mouse bone marrow was developed and challenged with p38is SB-203580, SB-226882, and SB-267030. Drug-related alterations in genes involved at different stages of erythropoiesis, cell-surface antigen expression (CSAE), burst-forming unit erythroid (BFU-E) colony formation, and cellular morphology (CM), growth (CG), and viability were evaluated. CSAE, CM, and decreases in BFU-E formation indicated delayed maturation, while CG and viability were unaffected. Terminal differentiation was delayed until day 14 versus day 7 in controls. CSAE demonstrated higher percentages of sca-1(+)cells after day 2 and reduced percentages of ter119(+) cells after day 7 in all treated cultures. Real-time reverse transcriptase polymerase chain reaction revealed a transient delay in expression of genes involved at early, intermediate, and late stages of erythropoiesis, followed by rebound expression at later time points. Results demonstrate p38is do not irreversibly inhibit erythrogenesis but induce a potency-dependent, transient delay in erythropoietic activity. The delay in activity is suggestive of effects on sca-1(+)bone marrow cells caused by alterations in expression of genes related to erythroid commitment and differentiation resulting in delayed maturation.

Assessment of oral mucosal conditions among beedi workers residing in beedi workers colonies in Karnataka, India.

BACKGROUND AND OBJECTIVES: Beedi workers are an important part of our society. Due to their occupation of beedi making, they face different types of health hazards. This study was undertaken to assess the oral mucosal conditions among beedi workers residing in beedi workers' colonies in Karnataka. METHODOLOGY: A cross-sectional study was carried out among 700 beedi workers in the age group of 18-60 years residing in five beedi workers' colonies in Karnataka. The information was recorded regarding personal history by personal interview by the investigator. The clinical examination was done using WHO 1997 "Oral Health Assessment Form." Significance is assessed at 5% level of significance. RESULTS: It was found that 26.9% of study population had different oral mucosal lesions. INTERPRETATION AND CONCLUSION: The present study showed that oral mucosal conditions of beedi workers residing in beedi workers' colonies in Karnataka were relatively poor.

Cloning, expression, and sequence analysis of diaminopropionate ammonia lyase gene from a nonvirulent Salmonella typhimurium PU011.

BACKGROUND: Seeds ofLathyrus sativus, a legume plant, contain 3-oxalyl and 2,3-dioxalyl DAP (O-DAP), neurotoxins which when consumed causes Neurolathyrism or Osteolathyrism, in humans, affecting nervous system and bone formation respectively. Some microorganisms viz virulent and non-virulentSalmonella typhimurium, Salmonella typhi and Pseudomonad have been shown to detoxifyL-alpha,beta-diaminopropionate (DAP), the immediate precursor of O-DAP. RESULT: The gene coding for diaminopropionate ammonia lyase (DAPAL) which detoxifies DAP was cloned from nonvirulentS. typhimurium PU011 into Escherichia coli DH5alpha and the nucleotides sequenced (1212 bp). Whereas the specific enzyme activity of DAPAL obtained from recombinantE. coli PU018 was 0.346 U/mg, the specific activity of the enzyme from nonvirulentS. typhimurium PU011 was 0.351 U/mg. The DAPAL corresponding to 43 kDa protein was found both in nonvirulentS. typhimurium PU011 andE. coli PU018. The Km value was found to be 0.740 mM and 0.680 mM forS. typhimurium PU011 and 0.741 mM and 0.683 mM forE. coli PU018 when grown in minimal medium (MM+DAP) andL. sativus seed extracts respectively, indicating that both of them were capable of utilizing the neurotoxins present inL. sativus seeds. The biomass, enzyme production and the effect of pH and temperature on DAPAL enzyme activity from both non-virulentS. typhimurium PU011 andE. coli PU018 were found to be similar. CONCLUSION: The recombinantE. coli PU018 as well as non-virulentS. typhimurium PU011 are as good as pathogenicS. typhimurium in detoxifying DAP, the immediate precursor of O-DAP present inL. sativus seeds.

Non-Lethal Endotoxin Injection: A Rat Model of Hypercoagulability.

Systemic inflammation co-activates coagulation, which unchecked culminates in a lethal syndrome of multi-organ microvascular thrombosis known as disseminated intravascular coagulation (DIC). We studied an endotoxin-induced inflammatory state in rats to identify biomarkers of hemostatic imbalance favoring hypercoagulability. Intraperitoneal injection of LPS at 15 mg/kg body weight resulted in peripheral leukopenia and widespread neutrophilic sequestration characteristic of an acute systemic inflammatory response. Early indicators of hemostatic pathway activation developed within 4 hours, including increased circulating concentrations of procoagulant extracellular vesicles (EVs), EVs expressing endothelial cell and platelet membrane markers, and high concentration of soluble intercellular adhesion molecule-1 (sICAM-1), plasminogen activator inhibitor-1 (PAI-1), and D-dimers. Inflammation persisted throughout the 48-hour observation period; however, increases were found in a subset of serum microRNA (miRNA) that coincided with gradual resolution of hemostatic protein abnormalities and reduction in EV counts. Dose-adjusted LPS treatment in rats provides a time-course model to develop biomarker profiles reflecting procoagulant imbalance and rebalance under inflammatory conditions.

Ultrashort solitons and kinetic effects in nonlinear metamaterials.

We present a stability analysis of a modified nonlinear Schrödinger equation describing the propagation of ultrashort pulses in negative refractive index media. Moreover, using methods of quantum statistics, we derive a kinetic equation for the pulses, making it possible to analyze and describe partial coherence in metamaterials. It is shown that a unique short pulse soliton, which is found analytically, can propagate in the medium.