Razvoj i prosudba protočne metode za dijagnosticiranje ehinokokoze u goveda

A novel in vitro flow through technique was developed and evaluated for immunodiagnosis of cystic echinococcosis in cattle using hydatid specific non-cross reactive 8 kDa protein. The 8 kDa protein was prepared from hydatid cyst fluid by DEAE-Sepharose fast flow anion exchange chromatography. In this flow through technique, the 8 kDa antigen was coated on the nitrocellulose membrane of the flow through device. Protein A Colloidal gold was used as detector. The evaluation of the technique was performed by comparing 150 known positive hydatid serum and known negative serum collected from cattle. The test was shown to have high sensitivity and specificity that were closely correlated with those of Enzyme-linked immunoelctrotransfer blot. Furthermore, the immunofiltration based assay is rapid and easy to perform with no requirement of special skill, reagent or instrumentation. This suggests the flow through technique is an acceptable alternative for use in clinical laboratories lacking specialized equipment as well as large scale screening of cystic echinococcosis in the field with both animal and human populations.U radu je opisana nova in vitro protočna tehnika prikladna za imunodijagnostiku ehinokokoze u goveda upotrebom visoko specifične bjelančevine od 8 kDa. Ta je bjelančevina bila pripravljena od hidatidne tekućine metodom DEAE-Sefaroze brzo protočne kromatografije s izmjenom aniona, a zatim naslojena na nitroceluloznu opnu u protočnom sustavu. Kao detektor rabljena je bjelančevina A obilježena koloidalnim zlatom. Metoda je vrednovana na temelju rezultata dobivenih pretraživanjem 150 pozitivnih i negativnih uzoraka seruma goveda. Istraživanje je pokazalo da je riječ o visoko osjetljivom i specifičnom testu vrlo sličnom metodi Western Blotting. Kako je opisani test brz i jednostavan i ne iziskuje posebne vještine, reagencije i opremu, smatra se da je osobito prikladan za opsežne probirne testove za dijagnosticiranje ehinokokoze u ljudi i životinja

Razvoj i prosudba protočne metode za dijagnosticiranje ehinokokoze u goveda

A novel in vitro flow through technique was developed and evaluated for immunodiagnosis of cystic echinococcosis in cattle using hydatid specific non-cross reactive 8 kDa protein. The 8 kDa protein was prepared from hydatid cyst fluid by DEAE-Sepharose fast flow anion exchange chromatography. In this flow through technique, the 8 kDa antigen was coated on the nitrocellulose membrane of the flow through device. Protein A Colloidal gold was used as detector. The evaluation of the technique was performed by comparing 150 known positive hydatid serum and known negative serum collected from cattle. The test was shown to have high sensitivity and specificity that were closely correlated with those of Enzyme-linked immunoelctrotransfer blot. Furthermore, the immunofiltration based assay is rapid and easy to perform with no requirement of special skill, reagent or instrumentation. This suggests the flow through technique is an acceptable alternative for use in clinical laboratories lacking specialized equipment as well as large scale screening of cystic echinococcosis in the field with both animal and human populations.U radu je opisana nova in vitro protočna tehnika prikladna za imunodijagnostiku ehinokokoze u goveda upotrebom visoko specifične bjelančevine od 8 kDa. Ta je bjelančevina bila pripravljena od hidatidne tekućine metodom DEAE-Sefaroze brzo protočne kromatografije s izmjenom aniona, a zatim naslojena na nitroceluloznu opnu u protočnom sustavu. Kao detektor rabljena je bjelančevina A obilježena koloidalnim zlatom. Metoda je vrednovana na temelju rezultata dobivenih pretraživanjem 150 pozitivnih i negativnih uzoraka seruma goveda. Istraživanje je pokazalo da je riječ o visoko osjetljivom i specifičnom testu vrlo sličnom metodi Western Blotting. Kako je opisani test brz i jednostavan i ne iziskuje posebne vještine, reagencije i opremu, smatra se da je osobito prikladan za opsežne probirne testove za dijagnosticiranje ehinokokoze u ljudi i životinja

Učinak 1-okten-3-ol kairomona preživača na ličinke krpelja iz porodice Ixodidae.

In vitro evaluations were made to evaluate the efficacy of 1-octen-3-ol on five ixodid larval stages using Petridish bioassay. The species of the ruminant ticks which are common in Indian sub continent including Rhipicephalus (Boophilus) microplus, Haemaphysalis bispinosa, Rhipicephalus haemaphysaloides, Hyalomma marginatum and dog tick Rhipicephalus sanguineus were made use in the current study. Amongst the ruminant tick larvae tested with 1-octen-3-ol, maximum attraction of 72 per cent and 71 per cent was recorded for H. marginatum and R. microplus respectively while H. bispinosa and R. haemaphysaloides showed 52 and 50 per cent attraction respectively. 1-octen-3-ol being a compound found characteristically in bovine odour failed to evoke any response (questing / attraction) in the brown dog tick.U radu je istražena in vitro učinkovitost 1-okten-3-ola, sastojka karakterističnog mirisa goveda, na ličinke pet različitih vrsta krpelja preživača koji se javljaju u Indiji: Rhipicephalus (Boophilus) microplus, Haemaphysalisž bispinosa, Rhipicephalus haemaphysaloides, Hyalomma marginatum, Rhipicephalus sanguineus. Među pretraženim krpeljima privlačnost u iznosu od 72% bila je dokazana za krpelja H. marginatum te 71% za krpelja R. microplus. Manja privlačnost zabilježena je za H. bispinosa i R. haemaphysaloides (52 i 50%). Isti sastojak uopće ne privlači krpelja R. sanguineus

Abnormal Complement Activation and Inflammation in the Pathogenesis of Retinopathy of Prematurity

Retinopathy of prematurity (ROP) is a neurovascular complication in preterm babies, leading to severe visual impairment, but the underlying mechanisms are yet unclear. The present study aimed at unraveling the molecular mechanisms underlying the pathogenesis of ROP. A comprehensive screening of candidate genes in preterms with ROP (n = 189) and no-ROP (n = 167) was undertaken to identify variants conferring disease susceptibility. Allele and genotype frequencies, linkage disequilibrium and haplotypes were analyzed to identify the ROP-associated variants. Variants in CFH (p = 2.94 x 10(-7)), CFB (p = 1.71 x 10(-5)), FBLN5 (p = 9.2 x 10(-4)), CETP (p = 2.99 x 10(-5)), and CXCR4 (p = 1.32 x 10(-8)) genes exhibited significant associations with ROP. Further, a quantitative assessment of 27 candidate proteins and cytokines in the vitreous and tear samples of babies with severe ROP (n = 30) and congenital cataract (n = 30) was undertaken by multiplex bead arrays and further validated by western blotting and zymography. Significant elevation and activation of MMP9 (p = 0.038), CFH (p = 2.24 x 10(-5)), C3 (p = 0.05), C4 (p = 0.001), IL-1ra (p = 0.0019), vascular endothelial growth factor (VEGF) (p = 0.0027), and G-CSF (p = 0.0099) proteins were observed in the vitreous of ROP babies suggesting an increased inflammation under hypoxic condition. Along with inflammatory markers, activated macrophage/microglia were also detected in the vitreous of ROP babies that secreted complement component C3, VEGF, IL-1ra, and MMP-9 under hypoxic stress in a cell culture model. Increased expression of the inflammatory markers like the IL-1ra (p = 0.014), MMP2 (p = 0.0085), and MMP-9 (p = 0.03) in the tears of babies at different stages of ROP further demonstrated their potential role in disease progression. Based on these findings, we conclude that increased complement activation in the retina/vitreous in turn activated microglia leading to increased inflammation. A quantitative assessment of inflammatory markers in tears could help in early prediction of ROP progression and facilitate effective management of the disease, thereby preventing visual impairment

Association between full service and fast food restaurant density, dietary intake and overweight/obesity among adults in Delhi, India

Abstract Background The food environment has been implicated as an underlying contributor to the global obesity epidemic. However, few studies have evaluated the relationship between the food environment, dietary intake, and overweight/obesity in low- and middle-income countries (LMICs). The aim of this study was to assess the association of full service and fast food restaurant density with dietary intake and overweight/obesity in Delhi, India. Methods Data are from a cross-sectional, population-based study conducted in Delhi. Using multilevel cluster random sampling, 5364 participants were selected from 134 census enumeration blocks (CEBs). Geographic information system data were available for 131 CEBs (n = 5264) from a field survey conducted using hand-held global positioning system devices. The number of full service and fast food restaurants within a 1-km buffer of CEBs was recorded by trained staff using ArcGIS software, and participants were assigned to tertiles of full service and fast food restaurant density based on their resident CEB. Height and weight were measured using standardized procedures and overweight/obesity was defined as a BMI ≥25 kg/m2. Results The most common full service and fast food restaurants were Indian savory restaurants (57.2%) and Indian sweet shops (25.8%). Only 14.1% of full service and fast food restaurants were Western style. After adjustment for age, household income, education, and tobacco and alcohol use, participants in the highest tertile of full service and fast food restaurant density were less likely to consume fruit and more likely to consume refined grains compared to participants in the lowest tertile (both p < 0.05). In unadjusted logistic regression models, participants in the highest versus lowest tertile of full service and fast food restaurant density were significantly more likely to be overweight/obese: odds ratio (95% confidence interval), 1.44 (1.24, 1.67). After adjustment for age, household income, and education, the effect was attenuated: 1.08 (0.92, 1.26). Results were consistent with further adjustment for tobacco and alcohol use, moderate physical activity, and owning a bicycle or motorized vehicle. Conclusions Most full service and fast food restaurants were Indian, suggesting that the nutrition transition in this megacity may be better characterized by the large number of unhealthy Indian food outlets rather than the Western food outlets. Full service and fast food restaurant density in the residence area of adults in Delhi, India, was associated with poor dietary intake. It was also positively associated with overweight/obesity, but this was largely explained by socioeconomic status. Further research is needed exploring these associations prospectively and in other LMICs

A Robust Model System for Retinal Hypoxia: Live Imaging of Calcium Dynamics and Gene Expression Studies in Primary Human Mixed Retinal Culture

The detailed mechanisms underlying oxidative stress that leads to neuroinflammation and neurodegeneration in retinal vascular conditions, including diabetic retinopathy, retinopathy of prematurity etc., remain largely unexplored mainly due to a lack of suitable disease models that can simulate the inherent neuron–glia interactions in human retina. Specifically, establishment of a mixed retinal culture (MRC) containing both neuron and glial cell types remains a challenge due to different conditions required for their optimal growth and differentiation. Here, we establish a novel primary MRC model system containing neurons, astrocytes, Müller glia, and microglia from human donor retina that can be used to study the neuromodulatory effects of glial cells under the stress. The cell characterization based on immunostaining with individual cell type–specific markers and their presence in close vicinity to each other further underscores their utility for studying their cross talk. To the best of our knowledge, this is the first instance of an in vitro model obtained from human donor retina containing four major cell types. Next, we induce hypoxic stress to MRC to investigate if hypoxia activated neuroglia modulates altered gene expression for inflammatory, apoptotic, and angiogenic markers and Ca2+ transients by live cell imaging. Further, we performed k-means clustering of the Ca2+ responses to identify the modification of clustering pattern in stressed condition. Finally, we provide the evidence that the altered Ca2+ transient correlates to differential expression of genes shown to be involved in neuroinflammation, angiogenesis, and neurodegeneration under the hypoxic conditions as seen earlier in human cell lines and animal models of diabetic retinopathy. The major features of the hypoxic conditions in the proposed human MRC model included: increase in microglia activity, chemokine and cytokine expression, and percentage of cells having higher amplitude and frequency of Ca2+ transients. Thus, the proposed experimental system can potentially serve as an ideal in vitro model for studying the neuroinflammatory and neurodegenerative changes in the retina and identifying newer drug targets