Multiple exposures to unloading decrease bone's responsivity but compound skeletal losses in C57BL/6 mice

A single exposure to mechanical unloading can result in significant bone loss, but the consequences of multiple exposures are largely unknown. Within a 18-wk period, adult C57BL/6 male mice were exposed to 2 wk of hindlimb unloading (HLU) followed by 4 wk of reambulation (RA) once (1x-HLU), twice (2x-HLU), or three times (3x-HLU), or served as ambulatory age-matched controls. In vivo μCT longitudinally tracked changes in trabecular and cortical compartments of the femur. Normally ambulating control mice experienced significant age-related loss in trabecular bone volume fraction throughout the course of the experiment. This loss was compounded by HLU with 2x- and 3x-HLU mice experiencing a 27% and 24% greater reduction in trabecular bone and a 60% and 63% inhibition of age-related trabecular thickening. The recovery of cortical bone was also incomplete during each 4-wk RA period and, at completion of the experiment, cortical area in 3x-HLU mice was 5% smaller than in control and 1x-HLU. When eliminating age as a confounding variable, comparison between individual HLU/RA cycles showed that the magnitude of the response diminished during subsequent exposures. The extent of trabecular thinning in mice unloaded for the first time was 1.6-fold greater than the second time and nearly twofold greater than the third time. Similarly, the increase in trabecular thickness during the first RA cycle was twofold greater than during the second and third RA cycle. Together, our data demonstrate that even though multiple exposures to mechanical unloading are more detrimental than a single unloading period, bone's mechanosensitivity is reduced with consecutive unloading/reambulation cycles.     

Simple Microfluidic Devices for in vivo Imaging of C. elegans,Drosophila and Zebrafish

Micro fabricated fluidic devices provide an accessible micro-environment for in vivo studies on small organisms. Simple fabrication processes are available for microfluidic devices using soft lithography techniques ^1-3. Microfluidic devices have been used for sub-cellular imaging ^4,5, in vivo laser microsurgery ^2,6 and cellular imaging ^4,7. In vivo imaging requires immobilization of organisms. This has been achieved using suction ^5,8, tapered channels ^6,7,9, deformable membranes ^2-4,10, suction with additional cooling ⁵, anesthetic gas ¹¹, temperature sensitive gels ¹², cyanoacrylate glue ¹³ and anesthetics such as levamisole ^14,15. Commonly used anesthetics influence synaptic transmission ^16,17 and are known to have detrimental effects on sub-cellular neuronal transport ⁴. In this study we demonstrate a membrane based poly-dimethyl-siloxane (PDMS) device that allows anesthetic free immobilization of intact genetic model organisms such as Caenorhabditis elegans (C. elegans), Drosophila larvae and zebrafish larvae. These model organisms are suitable for in vivo studies in microfluidic devices because of their small diameters and optically transparent or translucent bodies. Body diameters range from ~10 μm to ~800 μm for early larval stages of C. elegans and zebrafish larvae and require microfluidic devices of different sizes to achieve complete immobilization for high resolution time-lapse imaging. These organisms are immobilized using pressure applied by compressed nitrogen gas through a liquid column and imaged using an inverted microscope. Animals released from the trap return to normal locomotion within 10 min.We demonstrate four applications of time-lapse imaging in C. elegans namely, imaging mitochondrial transport in neurons, pre-synaptic vesicle transport in a transport-defective mutant, glutamate receptor transport and Q neuroblast cell division. Data obtained from such movies show that microfluidic immobilization is a useful and accurate means of acquiring in vivo data of cellular and sub-cellular events when compared to anesthetized animals (Figure 1J and 3C-F⁴).Device dimensions were altered to allow time-lapse imaging of different stages of C. elegans, first instar Drosophila larvae and zebrafish larvae. Transport of vesicles marked with synaptotagmin tagged with GFP (syt.eGFP) in sensory neurons shows directed motion of synaptic vesicle markers expressed in cholinergic sensory neurons in intact first instar Drosophila larvae. A similar device has been used to carry out time-lapse imaging of heartbeat in ~30 hr post fertilization (hpf) zebrafish larvae. These data show that the simple devices we have developed can be applied to a variety of model systems to study several cell biological and developmental phenomena in vivo.     

Leukocyte depletion for safe blood transfusion

Leukocytes have ability to distinguish between self cells (body own cells) and foreign (allogenic) cells on the basis of human leukocyte antigen (HLA) proteins that are present on the cell membrane and are effectively unique to a person. During allogenic blood transfusion a person receives large number of allogenic donor leukocytes and these are recognized as foreign cells by the recipient immune system which leads to several adverse reactions. To avoid such leukocyte-mediated adverse reactions leukodepleted blood transfusion is required. Leukocytes can be separated on the basis of size, dielectric properties, by affinity separation, freeze-thawing and centrifugation but all these methods are time consuming and costly. Filtration is another method for leukocyte depletion that is comparatively less expensive and more efficient as it gives more than 90% leukodepletion of blood along with minimal cell loss. However, present filtration procedures also have some limitations as they work efficiently with blood components but not with whole blood and show non-specific adhesion of large number of platelets and red blood cells along with leukocytes. All the currently available filters are costly, which has been a major reason for their limited application. Therefore, demand for a more efficient and cost-effective filter is high in medical community and scientists are attenpting to improve the efficiency of currently available filters. The present review gives an overview of the significance of leukodepleted blood transfusion and focuses on different methods for leukocyte depletion and challenges involved in all these technologies.     

Hepatoprotective effect of Hibiscus hispidissimus Griffith, ethanolic extract in paracetamol and CCl4 induced hepatotoxicity in Wistar rats

Hibiscus hispidissimus Griff. is used in tribal medicine of Kerala, the southern most state of India, to treat liver diseases. In the present study, the effect of the ethanolic extract of Hibiscus hispidissimus whole plant on paracetamol (PCM)-induced and carbon tetrachloride (CCl4)-induced liver damage in healthy Wistar albino rats was studied. The results showed that significant hepatoprotective effects were obtained against liver damage induced by PCM and CCl4 as evidenced by decreased levels of serum enzymes, glutamate oxaloacetate transaminase (SGOT), glutamate pyruvate transaminase (SGPT), serum alkaline phosphatase (SAKP), serum bilirubin (SB) and an almost normal histological architecture of the liver of the treated groups compared to the toxin controls. The extract also showed significant antilipid peroxidant effects in vitro, besides exhibiting significant activity in quenching 1, 1-diphenyl-2-picryl hydrazyl (DPPH) radical, indicating its potent antioxidant effects.     

Modification of Antioxidant Status of Host Cell in Response to Bougainvillea Antiviral Proteins

Bougainvillea xbuttiana antiviral proteins (AVPs) exhibited high antioxidant activity as measured by ferric reducing / antioxidant (FRAP) power assay. These AVPs were also found to modify activities of antioxidant enzymes like superoxide dismutase, peroxidase and catalase. The activities of superoxide dismutase and peroxidase increased, while the activity of catalase decreased in Tobacco mosaic virus (TMV) infected tobacco leaves. The trend was reversed when the leaves were treated with AVP alone. However, in TMV + AVP treated leaves, the activities of all the three enzymes were found to be midway between the activities obtained with other two treatments. It is therefore, suggested that Bougainvillea AVPs might be controlling viral diseases by scavenging reactive oxygen species as well as by altering host plant cell metabolism to maintain its antioxidant status.     

Oxygen deprivation induced cell death: an update

Mammalian cells have multiple responses to low or zero oxygen concentrations. In the complete absence of oxygen, cells undergo cell death through apoptosis, and not necrosis. Apoptotic signaling during oxygen deprivation occurs through the release of cytochrome c and apaf-1 mediated caspase-9 activation. The upstream regulators of cytochrome c release are the Bcl-2 family members. Pro-apoptotic Bcl-2 family members such as bax or bak are clearly required to initiate cytochrome c/apaf-1/caspase-9 mediated cell death during oxygen deprivation. Here we review what is currently known oxygen deprivation induced cell death and speculate about initiating mechanisms resulting in the activation of pro-apoptotic Bcl-2 family members.