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In a time when the entire world is going through a pandemic which can be curbed or substantially avoided and reduced by maintaining distance, it is crucial for every individual to follow the social distancing norms. However, it can be a challenging and daunting task to ensure safe distance all the time. Thus a Sixth Sense Jacket is a great way to keep the user mindful and alert of their surrounding without actually being alert all the time. It can not only alert the user but can also keep daily essentials such as keys and cards clean using dry sanitization. The jacket comes with a bunch of special features such as Pulse Oximeter which can be used to check vital health stats such as heart rate and blood saturation (SpO2) levels. The prime purpose of the Sixth Sense Jacket is to keep the user passively mindful of it's surrounding all the time and thus can be used in any field where it is crucial to stay alert in a given surrounding. The various domains this jacket can be used are women safety, battlefield and to add an extra sense to visually impaired people. The jacket can be a useful means to pre-empt any given danger that might befall the user and hence the name "Sixth Sense" Jacket.

Economic production quantity (EPQ) model has been analyzed for trended demand, and units in inventory are subject to constant rate. The system allows rework of imperfect units, and preventive maintenance time is random. A search method is used to study the model. The proposed methodology is validated by a numerical example. The sensitivity analysis is carried out to determine the critical model parameters. It is observed that the rate of change of demand, and the deterioration rate have a significant impact on the decision variables and the total cost of an inventory system. The model is highly sensitive to the production and demand rate.

An isocratic stability indicating reversed-phase liquid chromatographic determination was developed for the quantitative determination of lacosamide in the pharmaceutical dosage form. A Hypersil C-18, 4.5μm column with mobile phase containing acetonitrile-water (20:80, v/v) was used. The flow rate was 1.0 mL min-1 and effluents were monitored at 258 nm. The retention time of lacosamide was 8.9 min. The method was found to be linear in the concentration range of 5-100 μg/ml and the recovery was found to be in the range of 99.15 - 100.09 %. The limit of detection and limit of quantification were found to be 2 μg/ml and 5 μg/ml, respectively. Lacosamide stock solutions were subjected to acid and alkali hydrolysis, chemical oxidation and dry heat degradation. The drug was found to be stable to the dry heat and acidic condition attempted. The proposed method was validated and successfully applied to the estimation of lacosamide in tablet dosage forms.

A successful renal transplantation service was started in Nepal at the Tribhuvan Univer-sity Teaching Hospital in August 2008, and a continuing regular service is being provided currently to needy people. We report here our experience in thirty five end stage renal disease patients who re-ceived kidneys from close relatives during a one year period. The mean age of donors was 46.7 years. Seventeen (49%) donations were from parents, 13 (37%) from spouses, four (11%) between siblings and one (3%) between mother and daughter in law. Although the left kidney was given preference, right sided donor nephrectomy was needed in five (14%) cases. Six (17%) donors had minor post-operative problems. The mean age of recipients was 33.2 years, four (11%) of whom had pre-emptive renal transplantation. Recipients were immunosuppressed with dacluzimab, prednisolone, mycophena-late, and cyclosporine or tacrolimus. The average time taken for graft implantation was 137 minutes. The mean cold ischemia time and second warm ischemia time were 133 and 36 minutes respectively. Four (11%) patients developed urinary tract infection, three (9%) had significant hematuria, one (3%) developed a peri-transplant abscess, and one (3%) had ureteric ischemia and urine leak which required re-exploration in the early post-operative period. Four patients (11%) developed acute rejection of which three were cell- mediated rejection and one was antibody-mediated. There were two (6%) deaths, one due to transplant-related sepsis and the other due to subarachnoid hemorrhage following rupture of a posterior communicating artery aneurysm. No kidney has been lost otherwise.

Abstract Background Radiofrequency catheter ablation of excitation foci inside pulmonary veins (PV) generates stenoses that can become quite severe during or after the follow-up period. Since severe PV stenoses have most often disastrous consequences, it would be important to know the underlying mechanism of this temporal evolution. The present study proposes a potential explanation based on mechanical considerations. Methods we have used a mathematical-physical model to examine the cyclic increase in axial wall stress induced in the proximal (= upstream), non-stenosed segment of a stenosed pulmonary vein during the forward flow phases. In a representative example, the value of this increase at peak flow was calculated for diameter stenoses (DS) ranging from 1 to 99%. Results The increase becomes appreciable at a DS of roughly 30% and rise then strongly with further increasing DS value. At high DS values (e.g. > 90%) the increase is approximately twice the value of the axial stress present in the PV during the zero-flow phase. Conclusion Since abnormal wall stresses are known to induce damages and abnormal biological processes (e.g., endothelium tears, elastic membrane fragmentations, matrix secretion, myofibroblast generation, etc) in the vessel wall, it seems plausible that the supplementary axial stress experienced cyclically by the stenotic and the proximal segments of the PV is responsible for the often observed progressive reduction of the vessel lumen after healing of the ablation injury. In the light of this model, the only potentially effective therapy in these cases would be to reduce the DS as strongly as possible. This implies most probably stenting or surgery.

Traumas and chronic damages can hamper the regenerative power of nervous, muscle, and connective tissues. Tissue engineering approaches are promising therapeutic tools, aiming to develop reliable, reproducible, and economically affordable synthetic scaffolds which could provide sufficient biomimetic cues to promote the desired cell behaviour without triggering graft rejection and transplant failure. Here, we used 3D-printing to develop 3D-printed scaffolds based on either PLA or graphene@PLA with a defined pattern. Multiple regeneration strategies require a specific orientation of implanted and recruited cells to perform their function correctly. We tested our scaffolds with induced pluripotent stem cells (iPSC), neuronal-like cells, immortalised fibroblasts and myoblasts. Our results demonstrated that the specific "lines and ridges" 100 µm-scaffold topography is sufficient to promote myoblast and fibroblast cell alignment and orient neurites along with the scaffolds line pattern. Conversely, graphene is critical to promote cells differentiation, as seen by the iPSC commitment to neuroectoderm, and myoblast fusions into multinuclear myotubes achieved by the 100 µm scaffolds containing graphene. This work shows the development of a reliable and economical 3D-printed scaffold with the potential of being used in multiple tissue engineering applications and elucidates how scaffold micro-topography and graphene properties synergistically control cell differentiation.

Hematopoietic stem progenitor cells (HSPCs) have distinct metabolic plasticity, which allows them to transition from their quiescent state to a differentiation state to sustain demands of the blood formation. However, it has been difficult to analyze the metabolic status (mitochondrial respiration and glycolysis) of HSPCs due to their limited numbers and lack of optimized protocols for non-adherent, fragile HSPCs. Here, we provide a set of clear, step-by-step instructions to measure metabolic respiration (oxygen consumption rate; OCR) and glycolysis (extracellular acidification rate; ECAR) of murine bone marrow-Lineage neg Sca1 + c-Kit + (LSK) HSPCs. This protocol provides a higher amount of LSK HSPCs from murine bone marrow, improves the viability of HSPCs during incubation, facilitates extracellular flux analyses of non-adherent HSPCs, and provides optimized injection protocols (concentration and time) for drugs targeting oxidative phosphorylation and glycolytic pathways. This method enables the prediction of the metabolic status and the health of HSPCs during blood development and diseases.

Directed collective cell migration governs cell orientation during tissue morphogenesis, wound healing, and tumor metastasis. Unfortunately, current methods for initiating collective cell migration, such as scratching, laser ablation, and stencils, either introduce uncontrolled cell-injury, involve multiple fabrication processes, or have utility limited to cells with strong cell-cell junctions. Using three-dimensional (3D) bioprinted gelatin methacryloyl (GelMA) micropatterns on temperature-responsive poly( N -isopropylacrylamide) (PNIPAm) coated interfaces, we demonstrate that directed injury-free collective cell migration could occur in parallel and perpendicular directions. After seeding cells, we created cell-free spaces between two 3D bioprinted GelMA micropatterns by lowering the temperature of PNIPAm interfaces to promote the cell detachment. Unlike conventional collective cell migration methods initiated by stencils, we observed well-organized cell migration in parallel and perpendicular to 3D bioprinted micropatterns as a function of the distance between 3D bioprinted micropatterns. We further established the utility of controlled collective cell migration for directed functional myotube formation using 3D bioprinted fingerprintlike micropatterns as well as iris musclelike concentric circular patterns. Our platform is unique for myoblast alignment and myotube formation because it does not require anisotropic guidance cues. Together, our findings establish how to achieve controlled collective cell migration, even at the macroscale, for tissue engineering and regeneration.

Capsaicin causes a burning or spicy sensation when this vanilloid compound comes in contact with trigeminal neurons of the tongue. This compound has low solubility in water, which presents difficulties in examining the psychophysical properties of capsaicin by standard aqueous chemosensory tests. This report describes a new approach that utilizes edible strips for delivering precise amounts of capsaicin to the human oral cavity for examining threshold and suprathreshold amounts of this irritant. When incorporated into pullulan-based edible strips, recognition thresholds for capsaicin occurred over a narrow range, with a mean value near 1 nmol. When incorporated into edible strips at suprathreshold amounts, capsaicin yielded robust intensity values that were readily measured in our subject population. Maximal capsaicin intensity was observed 20 s after strips dissolved on the tongue surface, and then decreased in intensity. Suprathreshold studies showed that complete blockage of nasal airflow diminished capsaicin perception in the oral cavity. Oral rinses with vanillin-linoleic acid emulsions decreased mean intensity values for capsaicin by approximately 75%, but only modestly affected recognition threshold values. Also, oral rinses with isointense amounts of aqueous sucrose and sucralose solutions decreased mean intensity values for capsaicin by approximately 50%. In addition, this decrease in capsaicin intensity following an oral rinse with sucrose was partially reversed by the sweet taste inhibitor lactisole. These results suggest that blockage of nasal airflow, vanillin, sucrose, and sucralose modulate capsaicin perception in the human oral cavity. The results further suggest a chemosensory link between receptor cells that detect sweet taste stimuli and trigeminal neurons that detect capsaicin., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Purpose of Review: The hierarchical nature of the hematopoietic system provides an ideal model system to illustrate the features of lineage tracing. We have outlined the utility of lineage tracing methods in establishing the origin and development of hematopoietic cells., Recent Findings: Methods such as CRISPR/Cas9, Polylox barcoding, and single-cell RNA-sequencing have improved our understanding of hematopoiesis., Summary: This review chronicles the fate of the hematopoietic cells emerging from the mesoderm that subsequently develops into the adult blood lineages. Specifically, we explain classic techniques utilized in lineage tracing for the hematopoietic system, as well as novel state-of-the-art methods to elucidate clonal hematopoiesis and cell fate mapping at a single-cell level., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Transparency of the cornea is indispensable for optimal vision. Ocular trauma is a leading cause of corneal opacity, leading to 25 million cases of blindness annually. Recently, mesenchymal stem cells (MSCs) have gained prominence due to their inflammation-suppressing and tissue repair functions. Here, we investigate the potential of MSCs to restore corneal transparency following ocular injury. Using an in vivo mouse model of ocular injury, we report that MSCs have the capacity to restore corneal transparency by secreting high levels of hepatocyte growth factor (HGF). Interestingly, our data also show that HGF alone can restore corneal transparency, an observation that has translational implications for the development of HGF-based therapy., (Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.)

Surgical parabiosis of two animals of different genetic backgrounds creates a unique scenario to study cell-intrinsic versus cell-extrinsic roles for candidate genes of interest, migratory behaviors of cells, and secreted signals in distinct genetic settings. Because parabiotic animals share a common circulation, any blood or blood-borne factor from one animal will be exchanged with its partner and vice versa. Thus, cells and molecular factors derived from one genetic background can be studied in the context of a second genetic background. Parabiosis of adult mice has been  used extensively to research aging, cancer, diabetes, obesity, and brain development. More recently, parabiosis of zebrafish embryos has been used to study the developmental biology of hematopoiesis. In contrast to mice, the transparent nature of zebrafish embryos permits the direct visualization of cells in the parabiotic context, making it a uniquely powerful method for investigating fundamental cellular and molecular mechanisms. The utility of this technique, however, is limited by a steep learning curve for generating the parabiotic zebrafish embryos. This protocol provides a step-by-step method on how to surgically fuse the blastulae of two zebrafish embryos of different genetic backgrounds to investigate the role of candidate genes of interest. In addition, the parabiotic zebrafish embryos are tolerant to heat shock, making temporal control of gene expression possible. This method does not require a sophisticated set-up and has broad applications for studying cell migration, fate specification, and differentiation in vivo during embryonic development., Competing Interests: The authors declare that they have no conflict of interest.

Rare endothelial cells in the aorta-gonad-mesonephros (AGM) transition into hematopoietic stem cells (HSCs) during embryonic development. Lineage tracing experiments indicate that HSCs emerge from cadherin 5 (Cdh5; vascular endothelial-cadherin)(+) endothelial precursors, and isolated populations of Cdh5(+) cells from mouse embryos and embryonic stem cells can be differentiated into hematopoietic cells. Cdh5 has also been widely implicated as a marker of AGM-derived hemogenic endothelial cells. Because Cdh5(-/-) mice embryos die before the first HSCs emerge, it is unknown whether Cdh5 has a direct role in HSC emergence. Our previous genetic screen yielded malbec (mlb(bw306)), a zebrafish mutant for cdh5, with normal embryonic and definitive blood. Using time-lapse confocal imaging, parabiotic surgical pairing of zebrafish embryos, and blastula transplantation assays, we show that HSCs emerge, migrate, engraft, and differentiate in the absence of cdh5 expression. By tracing Cdh5(-/-)green fluorescent protein (GFP)(+/+) cells in chimeric mice, we demonstrated that Cdh5(-/-)GFP(+/+) HSCs emerging from embryonic day 10.5 and 11.5 (E10.5 and E11.5) AGM or derived from E13.5 fetal liver not only differentiate into hematopoietic colonies but also engraft and reconstitute multilineage adult blood. We also developed a conditional mouse Cdh5 knockout (Cdh5(flox/flox):Scl-Cre-ER(T)) and demonstrated that multipotent hematopoietic colonies form despite the absence of Cdh5. These data establish that Cdh5, a marker of hemogenic endothelium in the AGM, is dispensable for the transition of hemogenic endothelium to HSCs., (© 2015 by The American Society of Hematology.)

The transport and intracellular trafficking of heme biosynthesis intermediates are crucial for hemoglobin production, which is a critical process in developing red cells. Here, we profiled gene expression in terminally differentiating murine fetal liver-derived erythroid cells to identify regulators of heme metabolism. We determined that TMEM14C, an inner mitochondrial membrane protein that is enriched in vertebrate hematopoietic tissues, is essential for erythropoiesis and heme synthesis in vivo and in cultured erythroid cells. In mice, TMEM14C deficiency resulted in porphyrin accumulation in the fetal liver, erythroid maturation arrest, and embryonic lethality due to profound anemia. Protoporphyrin IX synthesis in TMEM14C-deficient erythroid cells was blocked, leading to an accumulation of porphyrin precursors. The heme synthesis defect in TMEM14C-deficient cells was ameliorated with a protoporphyrin IX analog, indicating that TMEM14C primarily functions in the terminal steps of the heme synthesis pathway. Together, our data demonstrate that TMEM14C facilitates the import of protoporphyrinogen IX into the mitochondrial matrix for heme synthesis and subsequent hemoglobin production. Furthermore, the identification of TMEM14C as a protoporphyrinogen IX importer provides a genetic tool for further exploring erythropoiesis and congenital anemias.

We used exome sequencing to identify mutations in sideroflexin 4 (SFXN4) in two children with mitochondrial disease (the more severe case also presented with macrocytic anemia). SFXN4 is an uncharacterized mitochondrial protein that localizes to the mitochondrial inner membrane. sfxn4 knockdown in zebrafish recapitulated the mitochondrial respiratory defect observed in both individuals and the macrocytic anemia with megaloblastic features of the more severe case. In vitro and in vivo complementation studies with fibroblasts from the affected individuals and zebrafish demonstrated the requirement of SFXN4 for mitochondrial respiratory homeostasis and erythropoiesis. Our findings establish mutations in SFXN4 as a cause of mitochondriopathy and macrocytic anemia., (Copyright © 2013 The American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.)

Sorting of endocytic ligands and receptors is critical for diverse cellular processes. The physiological significance of endosomal sorting proteins in vertebrates, however, remains largely unknown. Here we report that sorting nexin 3 (Snx3) facilitates the recycling of transferrin receptor (Tfrc) and thus is required for the proper delivery of iron to erythroid progenitors. Snx3 is highly expressed in vertebrate hematopoietic tissues. Silencing of Snx3 results in anemia and hemoglobin defects in vertebrates due to impaired transferrin (Tf)-mediated iron uptake and its accumulation in early endosomes. This impaired iron assimilation can be complemented with non-Tf iron chelates. We show that Snx3 and Vps35, a component of the retromer, interact with Tfrc to sort it to the recycling endosomes. Our findings uncover a role of Snx3 in regulating Tfrc recycling, iron homeostasis, and erythropoiesis. Thus, the identification of Snx3 provides a genetic tool for exploring erythropoiesis and disorders of iron metabolism., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Growth Factor Independence (Gfi) transcription factors play essential roles in hematopoiesis, differentially activating and repressing transcriptional programs required for hematopoietic stem/progenitor cell (HSPC) development and lineage specification. In mammals, Gfi1a regulates hematopoietic stem cells (HSC), myeloid and lymphoid populations, while its paralog, Gfi1b, regulates HSC, megakaryocyte and erythroid development. In zebrafish, gfi1aa is essential for primitive hematopoiesis; however, little is known about the role of gfi1aa in definitive hematopoiesis or about additional gfi factors in zebrafish. Here, we report the isolation and characterization of an additional hematopoietic gfi factor, gfi1b. We show that gfi1aa and gfi1b are expressed in the primitive and definitive sites of hematopoiesis in zebrafish. Our functional analyses demonstrate that gfi1aa and gfi1b have distinct roles in regulating primitive and definitive hematopoietic progenitors, respectively. Loss of gfi1aa silences markers of early primitive progenitors, scl and gata1. Conversely, loss of gfi1b silences runx-1, c-myb, ikaros and cd41, indicating that gfi1b is required for definitive hematopoiesis. We determine the epistatic relationships between the gfi factors and key hematopoietic transcription factors, demonstrating that gfi1aa and gfi1b join lmo2, scl, runx-1 and c-myb as critical regulators of teleost HSPC. Our studies establish a comparative paradigm for the regulation of hematopoietic lineages by gfi transcription factors., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Defects in the availability of haem substrates or the catalytic activity of the terminal enzyme in haem biosynthesis, ferrochelatase (Fech), impair haem synthesis and thus cause human congenital anaemias. The interdependent functions of regulators of mitochondrial homeostasis and enzymes responsible for haem synthesis are largely unknown. To investigate this we used zebrafish genetic screens and cloned mitochondrial ATPase inhibitory factor 1 (atpif1) from a zebrafish mutant with profound anaemia, pinotage (pnt (tq209)). Here we describe a direct mechanism establishing that Atpif1 regulates the catalytic efficiency of vertebrate Fech to synthesize haem. The loss of Atpif1 impairs haemoglobin synthesis in zebrafish, mouse and human haematopoietic models as a consequence of diminished Fech activity and elevated mitochondrial pH. To understand the relationship between mitochondrial pH, redox potential, [2Fe-2S] clusters and Fech activity, we used genetic complementation studies of Fech constructs with or without [2Fe-2S] clusters in pnt, as well as pharmacological agents modulating mitochondrial pH and redox potential. The presence of [2Fe-2S] cluster renders vertebrate Fech vulnerable to perturbations in Atpif1-regulated mitochondrial pH and redox potential. Therefore, Atpif1 deficiency reduces the efficiency of vertebrate Fech to synthesize haem, resulting in anaemia. The identification of mitochondrial Atpif1 as a regulator of haem synthesis advances our understanding of the mechanisms regulating mitochondrial haem homeostasis and red blood cell development. An ATPIF1 deficiency may contribute to important human diseases, such as congenital sideroblastic anaemias and mitochondriopathies.

The nuclear protein FOG-1 binds transcription factor GATA-1 to facilitate erythroid and megakaryocytic maturation. However, little is known about the function of FOG-1 during myeloid and lymphoid development or how FOG-1 expression is regulated in any tissue. We used in situ hybridization, gain- and loss-of-function studies in zebrafish to address these problems. Zebrafish FOG-1 is expressed in early hematopoietic cells, as well as heart, viscera, and paraspinal neurons, suggesting that it has multifaceted functions in organogenesis. We found that FOG-1 is dispensable for endoderm specification but is required for endoderm patterning affecting the expression of late-stage T-cell markers, independent of GATA-1. The suppression of FOG-1, in the presence of normal GATA-1 levels, induces severe anemia and thrombocytopenia and expands myeloid-progenitor cells, indicating that FOG-1 is required during erythroid/myeloid commitment. To functionally interrogate whether GATA-1 regulates FOG-1 in vivo, we used bioinformatics combined with transgenic assays. Thus, we identified 2 cis-regulatory elements that control the tissue-specific gene expression of FOG-1. One of these enhancers contains functional GATA-binding sites, indicating the potential for a regulatory loop in which GATA factors control the expression of their partner protein FOG-1.

A series of 5-alkylsulfamoyl benzimidazole derivatives have been designed and synthesized as novel angiotensin II (Ang II) receptor antagonists. The compounds have been evaluated for in vitro Ang II antagonism and for in vivo antihypertensive activity on isolated rat aortic ring and desoxycortisone acetate induced hypertensive rats, respectively. The activity is found related to size of alkyl group. The maximum activity is observed with a compact and bulky alkyl group like tert-butyl and cyclohexyl. The compounds 4g and 4h have shown promising both in vitro and in vivo activities. A receptor binding model is also proposed on the basis on the basis of structure-activity relationship in this study.

A series of 5-(alkyl and aryl)carboxamido benzimidazole derivatives had been designed, synthesized and evaluated for in vitro angiotensin II--AT1 receptor antagonism and in vivo antihypertensive activities. The pharmacological activities were inversely related to the size of alkyl and aryl substituents. It can be suggested that compounds with lower alkyl groups at 5-position of benzimidazole nucleus demonstrated potent antihypertensive activity.

Background: Activation of Akt stimulates phosphorylation of eNOS, production of nitric oxide and reduces oxidative stress. The study has been designed to investigate the effect of DAQ B1, an activator of Akt, in hypertension associated vascular endothelial dysfunction., Methods: Rats were uninephroctomized and DOCA (40 mg kg(-1), s.c.) was administered to rats to produce hypertension (MABP>140 mm Hg). Vascular endothelial dysfunction was assessed using isolated aortic ring preparation, electron microscopy of thoracic aorta and serum concentration of nitrite/nitrate. The expression of messenger RNA for p22phox and eNOS was assessed by reverse transcription-polymerase chain reaction. Serum TBARS and aortic superoxide anion were estimated to assess oxidative stress., Results: DAQ B1 (5 mg kg(-1), p.o.) or atorvastatin (30 mg kg(-1), p.o.) markedly improved acetylcholine induced endothelium dependent relaxation, vascular endothelial lining, expression of mRNA for eNOS and p22phox, serum nitrite/nitrate concentration and serum TBARS in hypertensive rats. However, this ameliorative effect of DAQ B1 has been prevented by L-NAME (25 mg kg(-1), i.p.), an inhibitor of eNOS., Conclusion: Therefore, it may be concluded that DAQ B1 induced activation of Akt may activate eNOS and consequently reduce oxidative stress to improve hypertension associated vascular endothelial dysfunction.

The study has been designed to investigate the effect of demethylasterroquinone B1 (DAQ B1), an activator of Akt, in diabetes mellitus (DM) and hyperhomocysteinemia (HHcy)-induced vascular endothelial dysfunction. Streptozotocin (55 mg kg(-1), i.v.) and methionine (1.7% w/w, p.o., 4 weeks) were administered to rats to produce DM (serum glucose >140 mg dl(-1)) and HHcy (serum homocysteine >10 microM), respectively. Vascular endothelial dysfunction was assessed using isolated aortic ring preparation, electron microscopy of thoracic aorta and serum concentration of nitrite/nitrate. The expression of messenger RNA for p22phox and eNOS was assessed by reverse transcription-polymerase chain reaction. Serum thiobarbituric acid reactive substances (TBARS) and aortic superoxide anion were estimated to assess oxidative stress. DAQ B1 (5 mg kg(-1), p.o.) or atorvastatin (30 mg kg(-1), p.o.) in diabetic and hyperhomocysteinemic rats significantly reduced serum glucose and homocysteine concentration. DAQ B1 or atorvastatin markedly improved acetylcholine-induced endothelium-dependent relaxation, vascular endothelial lining, serum nitrite/nitrate concentration and serum TBARS in diabetic and hyperhomocysteinemic rats. However, this ameliorative effect of DAQ B1 has been prevented by L-NAME (25 mg kg(-1), i.p.), an inhibitor of eNOS. Therefore, it may be concluded that DAQ B1-induced activation of Akt may activate eNOS and consequently reduce oxidative stress to improve vascular endothelial dysfunction.

The study has been designed to investigate the effect of 8-Br-cAMP, an activator of protein kinase A, in hypertension-induced vascular endothelial dysfunction. Rats were uninephroctomized and desoxycortisone acetate (DOCA) (40 mg/kg, s.c.) was administered to rats to produce hypertension (mean arterial blood pressure > 140 mmHg). Vascular endothelial dysfunction was assessed using isolated aortic ring preparation, electron microscopy of thoracic aorta and serum concentration of nitrite/nitrate. The expression of mRNA for p22phox and eNOS was assessed by using reverse transcriptase-polymerase chain reaction. Serum thiobarbituric acid reactive substances concentration and aortic superoxide anion concentration were estimated to assess oxidative stress. 8-Br-cAMP (5 mg/kg, i.p.) or atorvastatin (30 mg/kg, p.o.) prevented hypertension-induced attenuation of acetylcholine-induced endothelium-dependent relaxation, impairment of vascular endothelial lining, decrease in expression of mRNA for endothelial nitric oxide synthase (eNOS), serum nitrite/nitrate concentration and increase in expression of mRNA for p22phox, superoxide anion and serum TBARS. The ameliorative effect of 8-Br-cAMP was prevented by N-nitro-L-arginine methyl ester (25 mg/kg, i.p.) and glibenclamide (30 mg/kg, i.p.). It may be concluded that 8-Br-cAMP may stimulate expression and activity of eNOS and suppress expression of p22phox subunit of NADPH oxidase to reduce oxidative stress and subsequently improve vascular endothelial dysfunction.

Introduction: A new use of methylene blue as an ulcerogenic agent and the mechanisms involved were identified with an objective to exploit methylene blue as a pharmacological tool to study investigational antiulcer agents., Methods: Ulcerogenic potential was assessed using electron microscopy and measurement of an ulcer index after administering methylene blue (5-125 mg kg(-1), p.o.) or absolute ethanol (99%v/v, 2 ml, p.o.) to fasted rats. Estimation of thiobarbituric acid reactive substances, superoxide dismutase, reduced glutathione and catalase was used to assess oxidative stress. H(+)/K(+) ATPase activity, gastric mucosal blood flow and gastric acid secretion were measured to study the mechanism of methylene induced gastric ulcer., Results: Methylene blue (100 mg kg(-1), p.o.) produced marked ulceration of the gastric mucosa due to increased levels of thiobarbituric acid reactive substances, activities of the H(+)/K(+) ATPase and superoxide dismutase, and decreased blood flow to the gastric mucosa, activity of catalase combined with reduced glutathione levels., Discussion: It may be concluded that methylene blue activates the H(+)/K(+) ATPase to increase gastric acid secretion and reduces blood supply to gastric mucosa to produce oxidative stress that subsequently causes ulceration of gastric mucosa. Methylene blue can be used as an ulcerogenic agent to study mechanisms of investigational antiulcer agents.

The present study has been designed to investigate the effect of fasudil (Rho-kinase inhibitor) in hypercholesterolemia- and hypertension-induced endothelial dysfunction. High fat diet (8 weeks) and desoxycortisone acetate (DOCA) (40 mg.kg-1) were administered (s.c.) to rats to produce hypercholesterolemia and hypertension (mean arterial blood pressure > 120 mmHg), respectively. Endothelial dysfunction was assessed using isolated aortic ring, electron microscopy of thoracic aorta, and serum concentration of nitrite/nitrate. The expression of mRNA for p22phox and eNOS was assessed by using RT-PCR. Serum thiobarbituric acid reactive substances concentration and aortic superoxide anion concentration were estimated to assess oxidative stress. Fasudil (30 mg.kg-1, p.o.) and atorvastatin (30 mg.kg-1, p.o.) treatments markedly prevented hypercholesterolemia- and hypertension-evoked attenuation of acetylcholine-induced endothelium-dependent relaxation, impairment of vascular endothelial lining, decrease in expression of mRNA for eNOS and serum nitrite/nitrate concentration, and an increase in expression of mRNA for p22phox, superoxide anion, and serum thiobarbituric acid reactive substances. The ameliorative effect of fasudil was prevented by L-NAME. In conclusion, fasudil-induced inhibition of Rho-kinase may improve hypercholesterolemia- and hypertension-induced endothelial dysfunction.

The study has been designed to investigate the effect of 8-Br-cAMP, an activator of protein kinase A (PKA), in diabetes mellitus- and hyperhomocysteinemia-induced vascular endothelial dysfunction. Streptozotocin (55 mg kg-1, i.v.) and methionine (1.7% w/w, p.o., 4 weeks) were administered to rats to produce diabetes mellitus (serum glucose >200 mg dL-1) and hyperhomocysteinemia (serum homocysteine >10 microM), respectively. Vascular endothelial dysfunction was assessed using isolated aortic ring preparation, electron microscopy of thoracic aorta, and serum concentration of nitrite/nitrate. The expression of mRNA for p22phox and endothelial nitric oxide synthase (eNOS) was assessed by using reverse transcriptase-polymerase chain reaction (TBARS) (RT-PCR). Serum thiobarbituric acid-reactive substances (TBARS) concentration and aortic superoxide anion concentration were estimated to assess oxidative stress. 8-Br-cAMP (5 mg kg-1, i.p.) or atorvastatin (30 mg kg-1, p.o.) prevented diabetes mellitus- and hyperhomocysteinemia-induced attenuation of acetylcholine-induced endothelium-dependent relaxation, impairment of vascular endothelial lining, decrease in expression of mRNA for eNOS, serum nitrite/nitrate concentration, and increase in expression of mRNA for p22phox, superoxide anion, and serum TBARS. The ameliorative effect of 8-Br-cAMP was prevented by N omega-nitro-L-arginine methyl ester (L-NAME) (25 mg kg-1, i.p.) and glibenclamide (5 mg kg-1, i.p.). Therefore, it may be concluded that 8-Br-cAMP-induced activation of PKA may improve vascular endothelial dysfunction.

The study has been designed to investigate the effect of bis(maltolato) oxovanadium (BMOV), a protein tyrosine phosphatase inhibitor, on hypercholesterolemia and hypertension-induced vascular endothelial dysfunction. High fat diet (8 weeks) and deoxycorticosterone acetate (DOCA; 40 mg kg(-1), s.c.) were administered to rats to produce hypercholesterolemia and hypertension (mean arterial blood pressure >120 mmHg) respectively. Vascular endothelial dysfunction was assessed using isolated aortic ring preparation, electron microscopy of thoracic aorta, and serum concentration of nitrite/nitrate. Serum thiobarbituric acid reactive substances (TBARS) were estimated to assess oxidative stress. BMOV (0.2 mg/ml in drinking water) or atorvastatin (30 mg kg(-1), p.o.) markedly improved acetylcholine-evoked endothelium-dependent relaxation, lining of vascular endothelium, serum nitrite/nitrate concentration, and serum TBARS in hypercholesterolemic and hypertensive rats. However, this ameliorative effect of BMOV has been prevented by L-NAME (25 mg kg(-1), i.p.), an inhibitor of NOS, or by glibenclamide (5 mg kg(-1), i.p.), a blocker of ATP-sensitive K(+) channels. It may be concluded that BMOV-induced inhibition of PTPase may improve vascular endothelial dysfunction.

The study has been designed to investigate the effect of Bis-(maltolato) oxovanadium (BMOV), an inhibitor of protein tyrosin phosphatase (PTPase), in diabetes mellitus and hyperhomocysteinemia induced vascular endothelial dysfunction. Streptozotocin (55 mg kg(-1), i.v.) and methionine (1.7% w/w, p.o., 4 weeks) were administered to rats to produce diabetes mellitus (serum glucose >140 mg dl(-1)) and hyperhomocysteinemia (serum homocysteine>10 microM), respectively. Vascular endothelial dysfunction was assessed using isolated aortic ring preparation, electron microscopy of thoracic aorta and serum concentration of nitrite/nitrate. Serum thiobarbituric acid reactive substances (TBARS) were estimated to assess oxidative stress. Atorvastatin has been employed in the present study as standard drug to improve vascular endothelial dysfunction. BMOV (0.2 mg/ml in drinking water) or atorvastatin (30 mg kg(-1), p.o.) in diabetic and hyperhomocysteinemic rats significantly reduced serum glucose and homocysteine concentration. BMOV or atorvastatin markedly improved acetylcholine induced endothelium dependent relaxation, vascular endothelial lining, serum nitrite/nitrate concentration and serum TBARS in diabetic and hyperhomocysteinemic rats. However, this ameliorative effect of BMOV has been prevented by l-NAME (25 mg kg(-1), i.p.), an inhibitor of NOS or by glibenclamide (5 mg kg(-1), i.p.), a blocker of ATP sensitive K(+) channels. Therefore, it may be concluded that BMOV induced inhibition of PTPase may activate eNOS due to opening of ATP sensitive K(+) channels and consequently reduce oxidative stress to improve vascular endothelial dysfunction.

The present study has been designed to investigate the effect of fasudil (Rho-kinase inhibitor) in diabetes mellitus (DM) and hyperhomocyteinemia (HHcy) induced vascular endothelial dysfunction (VED). Streptozotocin (55 mg kg(-1), i.v., once only) and methionine (1.7% w/w, p.o., daily for 4 weeks) were administered to rats to produce DM (serum glucose >140 mg dl(-1)) and HHcy (serum homocysteine >10 microM) respectively. VED was assessed using isolated aortic ring, electron microscopy of thoracic aorta, and serum concentration of nitrite/nitrate. Serum thiobarbituric acid reactive substances (TBARS) concentration was estimated to assess oxidative stress. Atorvastatin has been employed in the present study as standard agent to improve vascular endothelial dysfunction. Fasudil (15 mg kg(-1) and 30 mg kg(-1), p.o., daily) and atorvastatin (30 mg kg(-1), p.o., daily) treatments significantly attenuated increase in serum glucose and homocysteine but their concentrations remained markedly higher than sham control value. Fasudil and atorvastatin treatments markedly prevented DM and HHcy-induced (i) attenuation of acetylcholine induced endothelium-dependent relaxation, (ii) impairment of vascular endothelial lining, (iii) decrease in serum nitrite/nitrate concentration, and (iv) increase in serum TBARS. It may be concluded that fasudil prevented DM and HHcy-induced VED partially by decreasing serum glucose and homocysteine concentration due to inhibition of Rho-kinase. Moreover, inhibition of Rho-kinase by fasudil and consequent prevention of oxidative stress may have directly improved VED in diabetic and hyperhomocysteinemic rats. The Rho-kinase appears to be a pivotal target site involved in DM and HHcy-induced VED.