Receptor dependent immobilization of spermatozoa by sperm immobilization factor isolated from Escherichia coli: Proof of evidence

Objective: To examine the receptor‐ligand interaction between E. coli and spermatozoa resulting in sperm immobilization. Methods: Sperm immobilization factor (SIF) was isolated and purified from filtrate of E. coli suspension. Receptor on human spermatozoa for SIF was made to isolated and purified by salt extraction, gel permeation and ion exchange chromatography. Receptor dependent immobilization of spermatozoa by SIF was confirmed by competitive inhibition by addition of the purified receptor and binding to sperm receptor by Fluorescin Isothiocynate (FITC) labelled SIF using fluorescence microscopy. Results: Heat labile, non‐dialyzable, sperm immobilization factor of ～56 kDa was isolated and purified from E. coli. Using SIF as a tool, receptor of 113 kDa could be isolated and purified from human spermatozoa. Addition of purified receptor completely inhibited sperm immobilization and death induced by SIF suggesting receptor‐dependent immobilization of spermatozoa. Further incubation of sperm with FITC labelled SIF resulted in staining of whole sperm. Conclusion: The study provides evidence of receptor‐ligand interaction between E. coli and spermatozoa.     

Cataract: A major secondary complication of diabetes,its epidemiology and an overview on major medicinal plants screened for anticataract activity

Cataract is a visual impairment caused due to opacification or optical dysfunction of crystallin lens affecting more than 17 million people around the world. Even though the incidences of cataract are increasing day by day among the elderly persons but, still except surgery no other ways of treatment have been successfully developed so far. Thus, the aim of writing the present review is to provide an insight over the pathophysiological and etiological aspects of cataract along with discussing the remedies available for the disorder. The review also describes different experimental models with their relevant mechanism and significance such as galactose-induced, naphthalene-induced and selenite-induced cataract models which are mainly used for evaluating the anticataract activity of a particular drug (mainly of natural origin). The review includes list of plants and their phytoconstituents which have been so far evaluated pharmacologicaly for the treatment of cataract. From the survey, it was confirmed that the antioxidant property of plants phytoconstituents are basically responsible for their effective anticataract activity. Thus, the valuable information provided in the present review will help researches in developing an alternative method rather than surgery for the treatment of cataract which will minimize the rate of blindness due to cataract thus, benefiting and extending protective aspects of eyes, an integral part of human body.     

Absence of Mutation at the 5′-Upstream Promoter Region of the <Emphasis Type="Italic">TPM4</Emphasis> Gene From Cardiac Mutant Axolotl (<Emphasis Type="Italic">Ambystoma mexicanum</Emphasis>)

Tropomyosins are a family of actin-binding proteins that show cell-specific diversity by a combination of multiple genes and alternative RNA splicing. Of the 4 different tropomyosin genes, TPM4 plays a pivotal role in myofibrillogenesis as well as cardiac contractility in amphibians. In this study, we amplified and sequenced the upstream regulatory region of the TPM4 gene from both normal and mutant axolotl hearts. To identify the cis-elements that are essential for the expression of the TPM4, we created various deletion mutants of the TPM4 promoter DNA, inserted the deleted segments into PGL3 vector, and performed promoter–reporter assay using luciferase as the reporter gene. Comparison of sequences of the promoter region of the TPM4 gene from normal and mutant axolotl revealed no mutations in the promoter sequence of the mutant TPM4 gene. CArG box elements that are generally involved in controlling the expression of several other muscle-specific gene promoters were not found in the upstream regulatory region of the TPM4 gene. In deletion experiments, loss of activity of the reporter gene was noted upon deletion which was then restored upon further deletion suggesting the presence of both positive and negative cis-elements in the upstream regulatory region of the TPM4 gene. We believe that this is the first axolotl promoter that has ever been cloned and studied with clear evidence that it functions in mammalian cell lines. Although striated muscle-specific cis-acting elements are absent from the promoter region of TPM4 gene, our results suggest the presence of positive and negative cis-elements in the promoter region, which in conjunction with positive and negative trans-elements may be involved in regulating the expression of TPM4 gene in a tissue-specific manner.     

Probing mucin-type O-linked glycosylation in living animals

Changes in O-linked protein glycosylation are known to correlate with disease states but are difficult to monitor in a physiological setting because of a lack of experimental tools. Here, we report a technique for rapid profiling of O-linked glycoproteins in living animals by metabolic labeling with N-azidoacetylgalactosamine (GalNAz) followed by Staudinger ligation with phosphine probes. After injection of mice with a peracetylated form of GalNAz, azide-labeled glycoproteins were observed in a variety of tissues, including liver, kidney, and heart, in serum, and on isolated splenocytes. B cell glycoproteins were robustly labeled with GalNAz but T cell glycoproteins were not, suggesting fundamental differences in glycosylation machinery or metabolism. Furthermore, GalNAz-labeled B cells could be selectively targeted with a phosphine probe by Staudinger ligation within the living animal. Metabolic labeling with GalNAz followed by Staudinger ligation provides a means for proteomic analysis of this posttranslational modification and for identifying O-linked glycoprotein fingerprints associated with disease.     

Surfactant-stabilized contrast agent on the nanoscale for diagnostic ultrasound imaging

Ultrasound contrast agents (CA) are generally micron-sized stabilized gas bubbles, injected IV. However, to penetrate beyond the vasculature and accumulate in targets such as tumors, CA must be an order of magnitude smaller. We describe a method of achieving nanometer-sized, surfactant-stabilized CA by differential centrifugation. High g force was shown to destroy bubble integrity. Optimal conditions (300 rpm for 3 min) produced an agent with a mean diameter of 450 nm, which gave 25.5 dB enhancement in vitro at a dose of 10 microL/mL, with a 13 min half-life. In vivo, the CA produced excellent power Doppler and grey-scale pulse inversion harmonic images at low acoustic power when administered. In vivo dose-response curves obtained in three rabbits showed enhancement between 20 and 25 dB for dosages above 0.025 mL/kg. These results encourage further investigation of the possible diagnostic and therapeutic benefits of using nanoparticles as CA, including passive targeting and accumulation in tumors.     

The association of depression and anxiety with obesity and unhealthy behaviors among community-dwelling US adults

OBJECTIVE: The aim of this study was to examine the extent to which depression and anxiety are associated with smoking, obesity, physical inactivity and alcohol consumption in the US population using the Patient Health Questionnaire 8 (PHQ-8) and two questions on lifetime diagnosis of anxiety and depression. METHODS: Data were analyzed in 38 states, the District of Columbia and two territories using the 2006 Behavioral Risk Factor Surveillance System (n=217,379), a large state-based telephone survey. RESULTS: Overall, adults with current depression or a lifetime diagnosis of depression or anxiety were significantly more likely than those without each diagnosis to smoke, to be obese, to be physically inactive, to binge drink and drink heavily. There was a dose-response relationship between depression severity and the prevalence of smoking, obesity and physical inactivity and between history of depression (never depressed, previously depressed, currently depressed) and the prevalence of smoking, obesity, physical inactivity, binge drinking and heavy drinking. Lifetime diagnosis of depression and anxiety had an additive association with smoking prevalence. CONCLUSION: The associations between depression, anxiety, obesity and unhealthy behaviors among US adults suggest the need for a multidimensional and integrative approach to health care.     

The design and evaluation of placebo material for crude herbals: Artemisia afra herb as a model

Herbal materials are known to present significant challenges with regard to designing credible placebos. This study intended to demonstrate the possibility of designing placebo material for crude herbals and used Artemisia afra, a popular traditional herbal medicine in South Africa, as a model. To produce the placebo, step-wise solvent extractions were conducted on the plant leaves and the process was monitored spectrophotometrically and using high performance liquid chromatography (HPLC) with diode array detection. The odour and taste between the placebo and A. afra was matched by inclusion of linalool and sodium saccharin, respectively. The muscle relaxant activity of the placebo was evaluated using an isolated guinea-pig tracheal muscle preparation. The UV absorbance of the extracts and the HPLC chromatograms, showed that most of the phytochemical constituents had been removed and the placebo closely resembled the A. afra leaves. The EC(50) of the placebo and the leaves were 4846.00 and 68.49 mg/mL, respectively, which showed that not only did the A. afra leaves possess muscle relaxant activity, but that the placebo did not possess any significant activity compared with the A. afra leaves (p value 0.0001). These results demonstrated that it is possible to design credible, pharmacologically inert placebo material for crude herbals.     

Identification of Leishmania donovani antigens stimulating cellular immune responses in exposed immune individuals

Human visceral leishmaniasis (VL), also known as kala azar (KA) in India, is a systemic progressive disease caused by Leishmania donovani. In VL, Th1 responses correlate with recovery from and resistance to disease and resolution of infection results in lifelong immunity against the disease. However, recent data suggest an important role for interleukin (IL)-10 in maintaining the resistant state. We evaluated whole cell extract (WE) and 11 antigenic fractions [F1-F11, molecular weight (MW) range of 139-24.2 kDa] from L. donovani (2001 strain, a fresh field isolate from Bihar), for their ability to induce in vitro T cell proliferation and production of interferon (IFN)-gamma, interleukin (IL)-12, IL-10 and IL-4 by peripheral blood mononuclear cells (PBMCs) of exposed immune individuals (14 patients with history of VL, 10 household endemic contacts) and 20 non-endemic healthy controls. Twenty-one of 24 exposed individuals and no healthy controls showed proliferative response to WE. Whole-extract activated IFN-gamma, IL-12, IL-10 levels were higher in the exposed group than in controls; IL-4 was not detectable in any of the samples. Among 21 responders to WE, frequent proliferative responses were seen to fractions F1-F4 (MW > 64.2 kDa) and none to fractions F5-F11; fractions F1-F11 stimulated comparable levels of IFN-gamma and IL-12 while IL-10 levels were higher in response to F5-F11 compared to F1-F4. These data demonstrate the presence of immunostimulatory antigens in the high MW fractions of whole L. donovani antigen. However, these fractions do not stimulate a Th1 response and produce variable amounts of IFN-gamma and the regulatory cytokine, IL-10. Hence, these high MW immunostimulatory fractions need to be evaluated in greater depth for their possible role as protective antigens.