Pretreatment of human retinal pigment epithelial cells with sterculic acid forestalls fenretinide-induced apoptosis.

The ratio of saturated to monounsaturated fatty acids, thought to play a critical role in many cellular functions, is regulated by stearoyl-CoA desaturase (SCD), a rate-limiting enzyme in the biosynthesis of monounsaturated fatty acids. Previously, we observed a decrease in both SCD protein and enzymatic activity in apoptosis induced by fenretinide, a synthetic analog of retinoic acid, in the human retinal pigment epithelial (RPE) cell line ARPE-19. Here, we investigated the effect of pretreating ARPE-19 with sterculic acid, a cyclopropenoic fatty acid inhibitor of SCD, on preventing fenretinide-induced apoptosis, given the role of SCD in cell proliferation and apoptosis. We show that sterculic acid pretreatment prevents the effects of fenretinide-induced apoptosis shown by changes in cell morphology, viability, and caspase-3 activation. Analysis of endoplasmic reticulum (ER)-associated proteins shows that sterculic acid pretreatment reduced the fenretinide-induced upregulation of heme oxygenase-1, ATF3 and GADD153 expression that are in response to reactive oxygen species (ROS) generation. Sterculic acid is as effective as allopurinol in inhibition of xanthine oxidase (XDH), and this may play a role in reducing the potential role of XDH in fenretinide-induced ROS generation. Sterculic acid pretreatment also results in a reduction in SOD2 mRNA expression. Dihydroceramide accumulation, compared to ceramide, and ROS generation indicate that a ceramide-independent pathway mediates fenretinide-induced apoptosis, and ROS mediation is borne out by activation of the NF-κBp50 and NF-κBp65 downstream signaling cascade. Its prevention by sterculic acid pretreatment further indicates the latter's antioxidant/anti-inflammatory effect. Taken together, our results suggest that sterculic acid pretreatment can mitigate ROS-mediated fenretinide-induced apoptosis. Thus, sterculic acid may serve as a potential antioxidant and therapeutic agent. These effects may be independent of its effects on SCD activity.

Cell-Free Amniotic Fluid and Regenerative Medicine: Current Applications and Future Opportunities.

Amniotic fluid (AF) provides critical biological and physical support for the developing fetus. While AF is an excellent source of progenitor cells with regenerative properties, recent investigations indicate that cell-free AF (cfAF), which consists of its soluble components and extracellular vesicles, can also stimulate regenerative and reparative activities. This review summarizes published fundamental, translational, and clinical investigations into the biological activity and potential use of cfAF as a therapeutic agent. Recurring themes emerge from these studies, which indicate that cfAF can confer immunomodulatory, anti-inflammatory, and pro-growth characteristics to the target cells/tissue with which they come into contact. Another common observation is that cfAF seems to promote a return of cells/tissue to a homeostatic resting state when applied to a model of cell stress or disease. The precise mechanisms through which these effects are mediated have not been entirely defined, but it is clear that cfAF can safely and effectively treat cutaneous wounds and perhaps orthopedic degenerative conditions. Additional applications are currently being investigated, but require further study to dissect the fundamental mechanisms through which its regenerative effects are mediated. By doing so, rational design can be used to fully unlock its potential in the biotechnology lab and in the clinic.

Comparative Analysis of Co-Cultured Amniotic Cell-Conditioned Media with Cell-Free Amniotic Fluid Reveals Differential Effects on Epithelial-Mesenchymal Transition and Myofibroblast Activation.

Myofibroblast activation is a cellular response elicited by a variety of physiological or pathological insults whereby cells initiate a coordinated response intended to eradicate the insult and then revert back to a basal state. However, an underlying theme in various disease states is persistent myofibroblast activation that fails to resolve. Based on multiple observations, we hypothesized that the secreted factors harvested from co-culturing amniotic stem cells might mimic the anti-inflammatory state that cell-free amniotic fluid (AF) elicits. We optimized an amnion epithelial and amniotic fluid cell co-culture system, and tested this hypothesis in the context of myofibroblast activation. However, we discovered that co-cultured amniotic cell conditioned media (coACCM) and AF have opposing effects on myofibroblast activation: coACCM activates the epithelial-mesenchymal transition (EMT) and stimulates gene expression patterns associated with myofibroblast activation, while AF does the opposite. Intriguingly, extracellular vesicles (EVs) purified from AF are necessary and sufficient to activate EMT and inflammatory gene expression patterns, while the EV-depleted AF potently represses these responses. In summary, these data indicate that coACCM stimulates myofibroblast activation, while AF represses it. We interpret these findings to suggest that coACCM, AF, and fractionated AF represent unique biologics that elicit different cellular responses that are correlated with a wide variety of pathological states, and therefore could have broad utility in the clinic and the lab.

High insecticide resistances levels in Anopheles gambiaes s.l. in northern Uganda and its relevance for future malaria control.

OBJECTIVE: The aim of the study was to determine the level of insecticide resistance and diversity in Anopheles mosquitoes in northern Uganda. Standard WHO insecticide susceptibility test assays were used to test for susceptibility to 0.5% malathion, 0.1% bendiocarb, 0.05% deltamethrin and 0.75% permethrin on 3-5 day old generation one progeny. We also screened for species diversity and knockdown resistance using PCR assay. RESULTS: Anopheles gambiae s.s. is the predominant malaria vector in northern Uganda followed by An. arabiensis. An. gambiae s.s. was susceptible to malathion and bendiocarb with the observed mortality rate of 100% and 98-100% observed respectively while very high resistance was observed with deltamethrin and permethrin. Minimal KDR-eastern variant homozygous forms of 8.3% in An. gambiae s.s. were detected in Oyam district. In conclusion, this study confirms that An. gambiae s.s. females are susceptible to malathion and bendiocarb while high intensity of resistance was observed with deltamethrin and permethrin in the same area. Use of carbamate and organophosphate insecticides bendiocarb and malathion for indoor residual spraying activities in northern Uganda is highly recommended since high levels of pyrethroids resistance (deltamethrin and permethrin) was detected in the area.

Web-Based Virtual Microscopy of Digitized Blood Slides for Malaria Diagnosis: An Effective Tool for Skills Assessment in Different Countries and Environments.

BACKGROUND: Morphological examination of blood films remains the reference standard for malaria diagnosis. Supporting the skills required to make an accurate morphological diagnosis is therefore essential. However, providing support across different countries and environments is a substantial challenge. OBJECTIVE: This paper reports a scheme supplying digital slides of malaria-infected blood within an Internet-based virtual microscope environment to users with different access to training and computing facilities. The feasibility of the approach was established, allowing users to test, record, and compare their own performance with that of other users. METHODS: From Giemsa stained thick and thin blood films, 56 large high-resolution digital slides were prepared, using high-quality image capture and 63x oil-immersion objective lens. The individual images were combined using the photomerge function of Adobe Photoshop and then adjusted to ensure resolution and reproduction of essential diagnostic features. Web delivery employed the Digital Slidebox platform allowing digital microscope viewing facilities and image annotation with data gathering from participants. RESULTS: Engagement was high with images viewed by 38 participants in five countries in a range of environments and a mean completion rate of 42/56 cases. The rate of parasite detection was 78% and accuracy of species identification was 53%, which was comparable with results of similar studies using glass slides. Data collection allowed users to compare performance with other users over time or for each individual case. CONCLUSIONS: Overall, these results demonstrate that users worldwide can effectively engage with the system in a range of environments, with the potential to enhance personal performance through education, external quality assessment, and personal professional development, especially in regions where educational resources are difficult to access.

Ovicidal, repellent, adulticidal and field evaluations of plant extract against dengue, malaria and filarial vectors.

Mosquitoes are insect vectors responsible for the transmission of parasitic and viral infections to millions of people worldwide, with substantial morbidity and mortality. Infections transmitted by mosquitoes include malaria, yellow fever, chikungunya, filariasis and other arboviruses. Insecticides of botanical origin may serve as suitable alternative biocontrol techniques in the future. The adulticidal activities of crude hexane, benzene, ethyl acetate, acetone and methanol leaf extracts of Acalypha alnifolia were assayed for their toxicity against three important vector mosquitoes, viz., Aedes aegypti, Anopheles stephensi and Culex quinquefasciatus. The adult mortality was observed after 24 h of exposure. All extracts showed moderate adulticide effects; however, the highest adult mortality was found in methanol extract were observed. The LC(50) values of A. alnifolia leaf extracts against adulticidal activity of (hexane, benzene, ethyl acetate, acetone and methanol) A. aegypti, A. stephensi and C. quinquefasciatus were the following: A. aegypti values were 371.87, 342.97, 320.17, 300.86 and 279.75 ppm; A. stephensi values were 358.35, 336.64, 306.10, 293.01 and 274.76 ppm; C. quinquefasciatus values were 383.59, 354.13, 327.74, 314.33 and 291.71 ppm. The results of the repellent activity of hexane, benzene, ethyl acetate, acetone and methanol extract of A. alnifolia plant at three different concentrations of 1.0, 3.0, and 5.0 mg/cm(2) were applied on skin of forearm in man and exposed against adult female mosquitoes. In this observation, this plant crude extracts gave protection against mosquito bites without any allergic reaction to the test person, and also, the repellent activity is dependent on the strength of the plant extracts. Mean percent hatchability of the ovicidal activity was observed 48 h post-treatment. The percent hatchability was inversely proportional to the concentration of extract and directly proportional to the eggs. Mortality of 100 % with methanol extract of A. alnifolia was exerted at 125 and 300 ppm. The larval density was decreased after the treatment of plant extracts at the breeding sites (water bodies system) of vector mosquitoes, and hence, these plant extracts are suitable alternatives of synthetic insecticides for mosquito vector management.These results suggest that the leaf solvent plant extracts have the potential to be used as an ideal eco-friendly approach for the control of mosquitoes. This study provides first report on the mosquito ovicidal, repellent and adulticidal activities of these plant extracts against mosquito vector species from India.

Adulticidal and repellent properties of Cassia tora Linn. (Family: Caesalpinaceae) against Culex quinquefasciatus, Aedes aegypti, and Anopheles stephensi.

Mosquitoes have developed resistance to various synthetic insecticides, making its control increasingly difficult. Insecticides of botanical origin may serve as suitable alternative biocontrol techniques in the future. The adulticidal and repellent activities of crude hexane, chloroform, benzene, acetone, and methanol extracts of the leaf of Cassia tora were assayed for their toxicity against three important vector mosquitoes, viz., Culex quinquefasciatus, Aedes aegypti, and Anopheles stephensi. The adult mortality was observed after 24 h of exposure. All extracts showed moderate adulticidal effects; however, the highest adult mortality observed was found in methanol extract. The LC(50) and LC(90) values of C. tora leaf extracts against adulticidal activity of (hexane, chloroform benzene, acetone, and methanol) C. quinquefasciatus, A. aegypti, and A. stephensi were the following: C. quinquefasciatus LC(50) values were 338.81, 315.73, 296.13, 279.23, and 261.03 ppm and LC(90) values were 575.77, 539.31, 513.99, 497.06, and 476.03 ppm; A. aegypti LC(50) values were 329.82, 307.31, 287.15, 269.57, and 252.03 ppm and LC(90) values were 563.24, 528.33, 496.92, 477.61, and 448.05 ppm; and A. stephensi LC(50) values were 317.28, 300.30, 277.51, 263.35, and 251.43 ppm and LC(90) values were 538.22, 512.90, 483.78, 461.08, and 430.70 ppm, respectively. The results of the repellent activity of hexane, chloroform, benzene, acetone, and methanol extracts of C. tora plant at three different concentrations of 1.0, 2.5, and 5.0 mg/cm(2) were applied on skin of forearm in man and exposed against adult female mosquitoes. In this observation, this plant crude extracts gave protection against mosquito bites without any allergic reaction to the test person, and also, the repellent activity is dependent on the strength of the plant extracts. These results suggest that the leaf solvent plant extracts have the potential to be used as an ideal eco-friendly approach for the control of mosquitoes. This is the first report on mosquito adulticidal and repellent activities of the reported C. tora against mosquito vectors from Southern India.

Perinatal outcomes of fetal echogenic bowel.

OBJECTIVE: To investigate perinatal outcomes of fetal echogenic bowel (FEB). METHOD: This is a retrospective observational study of FEB cases from Jan 2005-Dec 2010. Data from ultrasound and fetal medicine investigations, uterine artery Doppler (UAD), intra-partum care and neonatal outcome were obtained from Fetal Medicine, Obstetric and Neonatal Databases. RESULTS: There were 139 cases presenting at 21(+5) (15(+1) -35(+5) ) weeks gestation. Overall, 106/139 (76.2%) were live born (LB), 8/139 (5.8%) were complicated by intra-uterine deaths (IUD), 11/139 (7.9%) had termination of pregnancy (TOP) and 14/139 (10.1%) were lost to follow-up after 28 weeks gestation. Six had chromosomal/genetic abnormalities, two had congenital cytomegalovirus, none had cystic fibrosis.Uterine artery Doppler was normal in 106/130 (81.5%) cases. In this group, there were no cases of fetal growth restriction (FGR), 95/106 (89.6%) were LB, 1/106 (0.94%) had an IUD. In the abnormal UAD group, 17/24 (70.1%) developed FGR, 11/24 (45.8%) were LB, 4/24 (16.7%) had TOP, 7/24 (29.2%) had IUD.In total, 20/106 (18.9%) live births were admitted for specialist neonatal care, 12/20 (60%) for prematurity. Only one had primary bowel pathology. CONCLUSION: Pregnancies with FEB and screen positive UAD are at risk of adverse perinatal outcome. Primary bowel pathology is rare following the finding of FEB.