Circulating small extracellular vesicles in Alzheimer's disease: a case-control study of neuro-inflammation and synaptic dysfunction.

BACKGROUND: Alzheimer's disease (AD) is a neurodegenerative disease characterized by Aß plaques and neurofibrillary tangles. Chronic inflammation and synaptic dysfunction lead to disease progression and cognitive decline. Small extracellular vesicles (sEVs) are implicated in AD progression by facilitating the spread of pathological proteins and inflammatory cytokines. This study investigates synaptic dysfunction and neuroinflammation protein markers in plasma-derived sEVs (PsEVs), their association with Amyloid-ß and tau pathologies, and their correlation with AD progression. METHODS: A total of 90 [AD = 35, mild cognitive impairment (MCI) = 25, and healthy age-matched controls (AMC) = 30] participants were recruited. PsEVs were isolated using a chemical precipitation method, and their morphology was characterized by transmission electron microscopy. Using nanoparticle tracking analysis, the size and concentration of PsEVs were determined. Antibody-based validation of PsEVs was done using CD63, CD81, TSG101, and L1CAM antibodies. Synaptic dysfunction and neuroinflammation were evaluated with synaptophysin, TNF-α, IL-1ß, and GFAP antibodies. AD-specific markers, amyloid-ß (1-42), and p-Tau were examined within PsEVs using Western blot and ELISA. RESULTS: Our findings reveal higher concentrations of PsEVs in AD and MCI compared to AMC (p < 0.0001). Amyloid-ß (1-42) expression within PsEVs is significantly elevated in MCI and AD compared to AMC. We could also differentiate between the amyloid-ß (1-42) expression in AD and MCI. Similarly, PsEVs-derived p-Tau exhibited elevated expression in MCI compared with AMC, which is further increased in AD. Synaptophysin exhibited downregulated expression in PsEVs from MCI to AD (p = 0.047) compared to AMC, whereas IL-1ß, TNF-α, and GFAP showed increased expression in MCI and AD compared to AMC. The correlation between the neuropsychological tests and PsEVs-derived proteins (which included markers for synaptic integrity, neuroinflammation, and disease pathology) was also performed in our study. The increased number of PsEVs correlates with disease pathological markers, synaptic dysfunction, and neuroinflammation. CONCLUSIONS: Elevated PsEVs, upregulated amyloid-ß (1-42), and p-Tau expression show high diagnostic accuracy in AD. The downregulated synaptophysin expression and upregulated neuroinflammatory markers in AD and MCI patients suggest potential synaptic degeneration and neuroinflammation. These findings support the potential of PsEV-associated biomarkers for AD diagnosis and highlight synaptic dysfunction and neuroinflammation in disease progression.

The Role of Osteogenic Effect and Vascular Function in Bone Health in Hypertensive Rats: A Study of Anti-hypertensive and Hemorheologic Drugs.

Vascular dysfunction contributes to the development of osteopenia in hypertensive patients, as decreased blood supply to bones results in tissue damage and dysfunction. The effect of anti-hypertensive medicines on bone mass in hypertensive individuals is inconclusive because of the varied mechanism of their action, and suggests that reducing blood pressure (BP) alone is insufficient to enhance bone mass in hypertension. Pentoxifylline (PTX), a hemorheological drug, improves blood flow by reducing blood viscosity and angiogenesis, also has an osteogenic effect. We hypothesized that improving vascular function is critical to increasing bone mass in hypertension. To test this, we screened various anti-hypertensive drugs for their in vitro osteogenic effect, from which timolol and hydralazine were selected. In adult female spontaneously hypertensive rats (SHRs), timolol and hydralazine did not improve vascular function and bone mass, but PTX improved both. In female SHR animals, PTX restored bone mass, strength and mineralization, up to the level of normotensive control rats. In addition, we observed lower blood vasculature in the femur of adult SHR animals, and PTX restored them. PTX also restored the bone vascular and angiogenesis parameters that had been impaired in OVX SHR compared to sham SHR. This study demonstrates the importance of vascular function in addition to increased bone mass for improving bone health as achieved by PTX without affecting BP, and suggests a promising treatment option for osteoporosis in hypertensive patients, particularly at-risk postmenopausal women.

Ferroptosis and its modulators: A raising target for cancer and Alzheimer's disease.

The process of ferroptosis, a recently identified form of regulated cell death (RCD) is associated with the overloading of iron species and lipid-derived ROS accumulation. Ferroptosis is induced by various mechanisms such as inhibiting system Xc, glutathione depletion, targeting excess iron, and directly inhibiting GPX4 enzyme. Also, ferroptosis inhibition is achieved by blocking excessive lipid peroxidation by targeting different pathways. These mechanisms are often related to the pathophysiology and pathogenesis of diseases like cancer and Alzheimer's. Fundamentally distinct from other forms of cell death, such as necrosis and apoptosis, ferroptosis differs in terms of biochemistry, functions, and morphology. The mechanism by which ferroptosis acts as a regulatory factor in many diseases remains elusive. Studying the activation and inhibition of ferroptosis as a means to mitigate the progression of various diseases is a highly intriguing and actively researched topic. It has emerged as a focal point in etiological research and treatment strategies. This review systematically summarizes the different mechanisms involved in the inhibition and induction of ferroptosis. We have extensively explored different agents that can induce or inhibit ferroptosis. This review offers current perspectives on recent developments in ferroptosis research, highlighting the disease's etiology and presenting references to enhance its understanding. It also explores new targets for the treatment of cancer and Alzheimer's disease.

Fluorescence-tagged salivary small extracellular vesicles as a nanotool in early diagnosis of Parkinson's disease.

BACKGROUND: Parkinson's disease is generally asymptomatic at earlier stages. At an early stage, there is an extensive progression in the neuropathological hallmarks, although, at this stage, diagnosis is not possible with currently available diagnostic methods. Therefore, the pressing need is for susceptibility risk biomarkers that can aid in better diagnosis and therapeutics as well can objectively serve to measure the endpoint of disease progression. The role of small extracellular vesicles (sEV) in the progression of neurodegenerative diseases could be potent in playing a revolutionary role in biomarker discovery. METHODS: In our study, the salivary sEV were efficiently isolated by chemical precipitation combined with ultrafiltration from subjects (PD = 70, healthy controls = 26, and prodromal PD = 08), followed by antibody-based validation with CD63, CD9, GAPDH, Flotillin-1, and L1CAM. Morphological characterization of the isolated sEV through transmission electron microscopy. The quantification of sEV was achieved by fluorescence (lipid-binding dye-labeled) nanoparticle tracking analysis and antibody-based (CD63 Alexa fluor 488 tagged sEV) nanoparticle tracking analysis. The total alpha-synuclein (α-synTotal) in salivary sEVs cargo was quantified by ELISA. The disease severity staging confirmation for n = 18 clinically diagnosed Parkinson's disease patients was done by 99mTc-TRODAT-single-photon emission computed tomography. RESULTS: We observed a significant increase in total sEVs concentration in PD patients than in the healthy control (HC), where fluorescence lipid-binding dye-tagged sEV were observed to be higher in PD (p = 0.0001) than in the HC using NTA with a sensitivity of 94.34%. In the prodromal PD cases, the fluorescence lipid-binding dye-tagged sEV concentration was found to be higher (p = 0.008) than in HC. This result was validated through anti-CD63 tagged sEV (p = 0.0006) with similar sensitivity of 94.12%. We further validated our findings with the ELISA based on α-synTotal concentration in sEV, where it was observed to be higher in PD (p = 0.004) with a sensitivity of 88.24%. The caudate binding ratios in 99mTc-TRODAT-SPECT represent a positive correlation with sEV concentration (r = 0.8117 with p = 0.0112). CONCLUSIONS: In this study, for the first time, we have found that the fluorescence-tagged sEV has the potential to screen the progression of disease with clinically acceptable sensitivity and can be a potent early detection method for PD.

Anthropogenic disturbances influence mineral and elemental constituents of freshwater lake sediments.

Lake sediments can provide valuable insights into anthropogenic disturbances such as intensive aquaculture and land use changes. These disturbances often manifest as elevated levels of nutrients and elements within the sediments. This paper uses several analytical techniques, i.e., FTIR (Fourier-transform infrared spectroscopy), XRD (X-ray diffraction), EDS (energy-dispersive X-ray spectroscopy), and SEM (scanning electron microscopy), to examine the elemental constituents of lake sediments, along with their relative mineral abundances and surface morphology. The selected freshwater lakes are from the Central Gangetic Plain. The analysis provides a "fingerprint" of geogenic and biogenic mineral constituents of the sediments. Physicochemical, mineralogical, and elemental analysis shows that intensive aquaculture activities in lake alter the sediment chemistry as evidenced by the increase in pH, organic carbon, organic matter, and total phosphorus which is not observed in the lake where aquaculture is prohibited. Freshwater lake sediment is characterized by a high content of biogenic silica and carbonate minerals. The variations in sediment nutrients and mineral fluxes of the selected lakes are mainly attributed to diverse anthropogenic pressures, differences in lake productivity, and the overall ecological condition of the lakes. In the selected three lakes, major variation was reported in the autochthonous sediments in comparison to the allochthonous sediments. The study concludes that catchment and biotic deposit variations in the lakes cannot be evened out by in-lake mixing mechanisms due to variations in the terrigenous and pelagic deposits of the lake. The results highlight the importance of studying annual fluctuations and spatial variations in geogenic and biogenic mineral particle fluxes in lakes. Such investigations provide valuable insights into the annual dynamics of minerals within lakes, contributing to a more comprehensive understanding of their behavior and distribution.

Interspecific competition and their impacts on the growth of macrophytes and pollutants removal within constructed wetland microcosms treating domestic wastewater.

Eight free water surface constructed wetland microcosm (CWM) units are designed with single as well as mixed planting of Pistia stratiotes, Phragmites karka, and Typha latifolia with control to assess their competitive value (CV), relative growth rates (RGR), and pollutants removal efficiency. Further, the total dry biomass production and other growth parameters such as number of macrophytes, above-ground biomass, below-ground biomass, and root length were also measured to understand the dominant characteristics of the macrophytes. The CWM units with species mixture out-performed species monocultures. Removal of BOD, TP, SRP, NH4+-N, NO3--N, and NO2--N by mixed planting of P. stratiotes and P. karka was higher at most of the time. Typha latifolia was the superior competitor against both P. stratiotes and P. karka due to its aggressive characteristics that inhibits the growth of neighboring macrophytes. However, P. karka was the superior competitor against P. stratiotes. The RGR of T. latifolia in all experimental units was almost two times more than that of P. karka. Novelty Statement The CWM units with species mixture out-performed species monocultures. CWMs with more than one macrophytic species are less vulnerable to seasonal fluctuations and more effective in contaminants removal as compared to single macrophyte wetlands. Removal of BOD, TP, SRP, NH4+-N, NO3--N, and NO2--N by mixed planting of P. stratiotes and P. karka was higher at most of the time. The CWMs with P. stratiotes and P. karka are superior choice due to their higher wastewater nutrients removal capacity. The application of these three macrophytes in mixed cultures in free water surface constructed wetland is rare. The results are useful in designing large-scale multi-species wetlands which are less susceptible to seasonal variation and more effective in pollutants removal than single-species wetlands.

Constructed wetlands for the removal of pharmaceuticals and personal care products (PPCPs) from wastewater: origin, impacts, treatment methods, and SWOT analysis.

The continuous exposure to pharmaceuticals and personal care products can lead to a series of individual antagonistic and synergistic effects and long-lasting toxicity to humans and aquatic lives. This may also lead to developing antibiotic resistance, teratogenic, carcinogenic, and endocrine-disrupting effects. However, several PPCPs are also considered biologically active for non-target aquatic organisms, such as mosquito fish, goldfish, and the algae Pseudokirchneriella subcapitata. Various physicochemical methods such as ozonation, photolysis, and membrane separation are recognized for the effective removal of PPCPs. However, the high operation and maintenance costs and associated ecological impacts have limited their further use. Constructed wetlands are considered eco-friendly and sustainable for the removal of pharmaceuticals and personal care products together with antibiotic resistance genes. Several mechanisms such as sorption, biodegradation, oxidation, photodegradation, volatilization, and hydrolysis are occurring during the phytoremediation of PPCPs. During these processes, more than 50% of PPCPs can be eliminated through constructed wetlands. They also offer several additional benefits as obtained macrophytic biomass may be used as raw material in pulp and paper industries and a source for second-generation biofuel production. In this study, we have discussed the origin and impacts of PPCPs together with their treatment methods. We have also investigated the strengths, weaknesses, opportunities, and threats associated with constructed wetlands during the treatment of wastewater laden with pharmaceutical and personal care products.

MMP-7 derived peptides with MHC class-I binding motifs from canine mammary tumor tissue elicit strong antigen-specific T-cell responses in BALB/c mice.

Matrix Metalloproteinases (MMPs)-induced altered proteolysis of extracellular matrix proteins and basement membrane holds the key for tumor progression and metastasis. Matrix metalloproteinases-7 (Matrilysin), the smallest member of the MMP family also performs quite alike; thus serves as a potential candidate for anti-tumor immunotherapy. Conversely, being an endogenous tumor-associated antigen (TAA), targeting MMP-7 for immunization is challenging. But MMP-7-based xenovaccine can surmount the obstacle of poor immunogenicity and immunological tolerance, often encountered in TAA-based conventional vaccine for anti-tumor immunotherapy. This paves the way for investigating the potential of MMP-7-derived major histocompatibility complex (MHC)-binding peptides to elicit precise epitope-specific T-cell responses towards their possible inclusion in anti-tumor vaccine formulations. Perhaps it also ushers the path of achieving multiple epitope-based broad and universal cellular immunity. In current experiment, an immunoinformatics approach has been employed to identify the putative canine matrix matelloproteinases-7 (cMMP-7)-derived peptides with MHC class-I-binding motifs which can elicit potent antigen-specific immune responses in BALB/c mice. Immunization with the cMMP-7 DNA vaccine induced a strong CD8+ cytotoxic T lymphocytes (CTLs) and Th1- type response, with high level of gamma interferon (IFN-γ) production in BALB/c mice. The two identified putative MHC-I-binding nonameric peptides (Peptide32-40 and Peptide175-183) from cMMP-7 induced significant lymphocyte proliferation along with the production of IFN-γ from CD8+ T-cells in mice immunized with cMMP-7 DNA vaccine. The current observation has depicted the immunogenic potential of the two cMMP-7-derived nonapeptides for their possible exploitation in xenovaccine-mediated anti-tumor immunotherapy in mouse model.

The Evolving Landscape of Exosomes in Neurodegenerative Diseases: Exosomes Characteristics and a Promising Role in Early Diagnosis.

Neurodegenerative diseases (ND) remains to be one of the biggest burdens on healthcare systems and serves as a leading cause of disability and death. Alzheimer's disease (AD) is among the most common of such disorders, followed by Parkinson's disease (PD). The basic molecular details of disease initiation and pathology are still under research. Only recently, the role of exosomes has been linked to the initiation and progression of these neurodegenerative diseases. Exosomes are small bilipid layer enclosed extracellular vesicles, which were once considered as a cellular waste and functionless. These nano-vesicles of 30-150 nm in diameter carry specific proteins, lipids, functional mRNAs, and high amounts of non-coding RNAs (miRNAs, lncRNAs, and circRNAs). As the exosomes content is known to vary as per their originating and recipient cells, these vesicles can be utilized as a diagnostic biomarker for early disease detection. Here we review exosomes, their biogenesis, composition, and role in neurodegenerative diseases. We have also provided details for their characterization through an array of available techniques. Their updated role in neurodegenerative disease pathology is also discussed. Finally, we have shed light on a novel field of salivary exosomes as a potential candidate for early diagnosis in neurodegenerative diseases and compared the biomarkers of salivary exosomes with other blood/cerebrospinal fluid (CSF) based exosomes within these neurological ailments.

Drug-induced reactive oxygen species-mediated inhibitory effect on growth of Trypanosoma evansi in axenic culture system.

Trypanosoma evansi, an extracellular haemoflagellate, has a wide range of hosts receptive and susceptible to infection, in which it revealed highly inconsistent clinical effects. Drugs used for the treatment of trypanosomosis have been utilized for more than five decades and have several problems like local and systemic toxicity. In the present investigation, imatinib and sorafenib were selected as drugs as they are reported to have the potential to cause reactive oxygen species (ROS)-mediated effect in cancer cells. Both have also been reported to have potential against T. brucei, T. cruzi and Leishmania donovani. To date, imatinib and sorafenib have not evaluated for their growth inhibitory effect against T. evansi. Imatinib and sorafenib showed significant (p < 0.001) inhibition on parasite growth and multiplication with IC50 (50% inhibitory concentration) values 6.12 µM and 0.33 µM respectively against T. evansi. Both the drug molecules demonstrated for the generation of ROS in T. evansi and were found up to 65% increased level of ROS as compared with negative control in the axenic culture system. Furthermore, different concentrations of imatinib and sorafenib were found non-toxic on horse peripheral blood mononuclear cells and Vero cell lines. Also, in conclusion, our results demonstrated that imatinib- and sorafenib-induced generation of ROS contributed inhibitory effect on the growth of Trypanosoma evansi in an axenic culture system.

An aptasensor for rapid and sensitive detection of estrogen receptor alpha in human breast cancer.

The analysis of estrogen receptor (ER) expression in breast carcinomas plays a crucial role in determining the endocrine responsiveness of tumors for systemic adjuvant therapy. Conventionally, the ER levels in breast carcinomas had been detected using the dextran-coated charcoal assay and radioimmunoassay, which are now substituted with safer and economic antibody-based assays such as immunohistochemistry (IHC) and enzyme-linked immunosorbent assay (ELISA). Despite a gold (Au) standard method, the IHC has been criticized for factors such as tissue fixation, antibody selection, and threshold staining for result interpretation that could falsify test accuracy and reproducibility. The quest for alternative methods of ER quantification in tissue samples paved the way for aptamer-based diagnostics. Previously, we have isolated a DNA aptamer against human ER alpha (ERα) using an in vitro evolution system. In this study, we developed an electrochemical sensor using the 76-nucleotide DNA ERα- aptamer for rapid, precise, and cost-effective detection of ERα expression in human breast cancer patients. The aptasensor was constructed by covalently immobilizing the thiolated ERα- aptamer onto a screen-printed Au electrode. Construction of aptasensors was confirmed through atomic force microscopy and differential pulse voltammetry measurements. A detection limit of 0.001 ng/ml was calculated for full-length ERα (66.2 kDa) in a detection time of 10 min. Analysis of the cancerous breast tissue samples using the ELISA and aptasensor methods enabled distinctive classification of samples into the categories of ER -ve, weak ER +ve, and strong ER +ve samples. The current change of this aptasensor lies within 5% after a storage of 60 days at 4°C. Further studies on a reasonably large sample size are required to realize the clinical potential of the sensor.

Tele-Emergency Services in the Himalayas.

Introduction:Nonavailability of emergency healthcare services in mountainous, isolated, and sparsely populated regions is a universal problem. In a first of its kind initiative, Tele-emergency services (TES) was provided in Keylong and Kaza in Himachal Pradesh in Northern India, at an altitude of 3,353 meters with temperatures of -30°C during winter months.Methods:Existing rooms in regional hospital (Keylong) and community health center (Kaza) were converted into tele-emergency centers by connecting them, to a state-of-the-art emergency department at the Joint Commission International-accredited Apollo Main Hospital at Chennai, 2,925 km away. Training was carried out at both ends. Average turnaround time for an emergency teleconsult was less than 12 minutes. Tele-ECG, Spirometry, and Point-of-Care Diagnostics for blood biochemistry were made available.Results:In the first 35 months, 753 teleconsults were given in the 24/7 TES, out of a total of 10,213 teleconsults constituting 7.4%. Out of a total of 6,442 telelaboratory tests, 431 tests were done in an emergency setting constituting 6.7%. Of the 16 cases of myocardial infarction remotely diagnosed, 4 were thrombolysed through telementoring. Of seven patients with Supra Ventricular Tachycardia, six patients were stabilized through electrical cardioversion and one through chemical cardioversion through telementoring. Ten deaths were documented, of which one occurred at the site. One hundred ninety-six were stabilized and transferred to higher centers. Thirteen required helicopter evacuations. Detailed analysis revealed that the total average cost for a single emergency teleconsult during this period was US$208.Conclusions:Preliminary analysis confirms that delivering TES in inhospitable terrains in a Public Private Partnership mode is doable and is welcomed by the community.

Novel Insights into Regulation of Human Teeth Biomineralization: Deciphering the Role of Post-Translational Modifications in a Tooth Protein Extract.

The importance of whole protein extracts from different types of human teeth in modulating the process of teeth biomineralization is reported. There are two crucial features in protein molecules that result in efficient teeth biomineralization. Firstly, the unique secondary structure characteristics within these proteins i.e. the exclusive presence of a large amount of intrinsic disorder and secondly, the presence of post-translational modifications (PTM) like phosphorylation and glycosylation within these protein molecules. The present study accesses the structural implications of PTMs in the tooth proteins through scanning electron microscopy and transmission electron microscopy. The deglycosylated/dephosphorylated protein extracts failed to form higher-order mineralization assemblies. Furthermore, through nanoparticle tracking analysis (NTA) we have shown that dephosphorylation and deglycosylation significantly impact the biomineralization abilities of the protein extract and resulted in smaller sized clusters. Hence, we propose these post-translational modifications are indispensable for the process of teeth biomineralization. In addition to basic science, this study would be worth consideration while designing of biomimetics architecture for an efficient peptide-based teeth remineralization strategy.

Moderate reagent mixing on an orbital shaker reduces the incubation time of enzyme-linked immunosorbent assay.

Rapid diagnostic tests can be developed using ELISA for detection of diseases in emergency conditions. Conventional ELISA takes 1-2 days, making it unsuitable for rapid diagnostics. Here, we report the effect of reagents mixing via shaking or vortexing on the assay timing of ELISA. A 48-min protocol of ELISA involving 12-min incubations with reagent mixing at 750 rpm for every step was optimized. Contrary to this, time-optimized control ELISA performed without mixing produced similar results in 8 h, leaving a time gain of 7 h using the developed protocol. Collectively, the findings suggest the development of ELISA-based rapid diagnostics.

Evaluating Changes in Cell-Wall Components Associated with Clubroot Resistance Using Fourier Transform Infrared Spectroscopy and RT-PCR.

Clubroot disease is a serious threat to canola production in western Canada and many parts of the world. Rcr1 is a clubroot resistance (CR) gene identified recently and its molecular mechanisms in mediating CR have been studied using several omics approaches. The current study aimed to characterize the biochemical changes in the cell wall of canola roots connecting to key molecular mechanisms of this CR gene identified in prior studies using Fourier transform infrared (FTIR) spectroscopy. The expression of nine genes involved in phenylpropanoid metabolism was also studied using qPCR. Between susceptible (S) and resistance (R) samples, the most notable biochemical changes were related to an increased biosynthesis of lignin and phenolics. These results were supported by the transcription data on higher expression of BrPAL1. The up-regulation of PAL is indicative of an inducible defence response conferred by Rcr1; the activation of this basal defence gene via the phenylpropanoid pathway may contribute to clubroot resistance conferred by Rcr1. The data indicate that several cell-wall components, including lignin and pectin, may play a role in defence responses against clubroot. Principal components analysis of FTIR data separated non-inoculated samples from inoculated samples, but not so much between inoculated S and inoculated R samples. It is also shown that FTIR spectroscopy can be a useful tool in studying plant-pathogen interaction at cellular levels.

Myosin-Induced Gliding Patterns at Varied [MgATP] Unveil a Dynamic Actin Filament.

Actin filaments have key roles in cell motility but are generally claimed to be passive interaction partners in actin-myosin-based motion generation. Here, we present evidence against this static view based on an altered myosin-induced actin filament gliding pattern in an in vitro motility assay at varied [MgATP]. The statistics that characterize the degree of meandering of the actin filament paths suggest that for [MgATP] ≥ 0.25 mM, the flexural rigidity of heavy meromyosin (HMM)-propelled actin filaments is similar (without phalloidin) or slightly lower (with phalloidin) than that of HMM-free filaments observed in solution without surface tethering. When [MgATP] was reduced to ≤0.1 mM, the actin filament paths in the in vitro motility assay became appreciably more winding in both the presence and absence of phalloidin. This effect of lowered [MgATP] was qualitatively different from that seen when HMM was mixed with ATP-insensitive, N-ethylmaleimide-treated HMM (NEM-HMM; 25-30%). In particular, the addition of NEM-HMM increased a non-Gaussian tail in the path curvature distribution as well as the number of events in which different parts of an actin filament followed different paths. These effects were the opposite of those observed with reduced [MgATP]. Theoretical modeling suggests a 30-40% lowered flexural rigidity of the actin filaments at [MgATP] ≤ 0.1 mM and local bending of the filament front upon each myosin head attachment. Overall, the results fit with appreciable structural changes in the actin filament during actomyosin-based motion generation, and modulation of the actin filament mechanical properties by the dominating chemomechanical actomyosin state.

A novel multi-modal platform to image molecular and elemental alterations in ischemic stroke.

Stroke is a major global health problem, with the prevalence and economic burden predicted to increase due to aging populations in western society. Following stroke, numerous biochemical alterations occur and damage can spread to nearby tissue. This zone of "at risk" tissue is termed the peri-infarct zone (PIZ). As the PIZ contains tissue not initially damaged by the stroke, it is considered by many as salvageable tissue. For this reason, much research effort has been undertaken to improve the identification of the PIZ and to elucidate the biochemical mechanisms that drive tissue damage in the PIZ in the hope of identify new therapeutic targets. Despite this effort, few therapies have evolved, attributed in part, to an incomplete understanding of the biochemical mechanisms driving tissue damage in the PIZ. Magnetic resonance imaging (MRI) has long been the gold standard to study alterations in gross brain structure, and is frequently used to study the PIZ following stroke. Unfortunately, MRI does not have sufficient spatial resolution to study individual cells within the brain, and reveals little information on the biochemical mechanisms driving tissue damage. MRI results may be complemented with histology or immuno-histochemistry to provide information at the cellular or sub-cellular level, but are limited to studying biochemical markers that can be successfully "tagged" with a stain or antigen. However, many important biochemical markers cannot be studied with traditional MRI or histology/histochemical methods. Therefore, we have developed and applied a multi-modal imaging platform to reveal elemental and molecular alterations that could not previously be imaged by other traditional methods. Our imaging platform incorporates a suite of spectroscopic imaging techniques; Fourier transform infrared imaging, Raman spectroscopic imaging, Coherent anti-stoke Raman spectroscopic imaging and X-ray fluorescence imaging. This approach does not preclude the use of traditional imaging techniques, and rather it should be use to complement traditional methods such as MRI or histology and immunohistochemistry, to gain a greater insight into disease mechanisms. We demonstrate the potential of this approach by characterizing biochemical alterations within the PIZ 24h after the induction of photothrombotic stroke in mice. Substantial molecular and elemental alterations were identified in the PIZ 24h after stroke that are consistent with tissue swelling and edema, but not oxidative stress. This reveals important mechanistic information, that could not previously be obtained, which should be considered in future studies aimed at developing therapeutic intervention from this model.

Sensing protein antigen and microvesicle analytes using high-capacity biopolymer nano-carriers.

Lab-on-a-chip systems with molecular motor driven transport of analytes attached to cytoskeletal filament shuttles (actin filaments, microtubules) circumvent challenges with nanoscale liquid transport. However, the filaments have limited cargo-carrying capacity and limitations either in transportation speed (microtubules) or control over motility direction (actin). To overcome these constraints we here report incorporation of covalently attached antibodies into self-propelled actin bundles (nanocarriers) formed by cross-linking antibody conjugated actin filaments via fascin, a natural actin-bundling protein. We demonstrate high maximum antigen binding activity and propulsion by surface adsorbed myosin motors. Analyte transport capacity is tested using both protein antigens and microvesicles, a novel class of diagnostic markers. Increased incubation concentration with protein antigen in the 0.1-100 nM range (1 min) reduces the fraction of motile bundles and their velocity but maximum transportation capacity of >1 antigen per nm of bundle length is feasible. At sub-nanomolar protein analyte concentration, motility is very well preserved opening for orders of magnitude improved limit of detection using motor driven concentration on nanoscale sensors. Microvesicle-complexing to monoclonal antibodies on the nanocarriers compromises motility but nanocarrier aggregation via microvesicles shows unique potential in label-free detection with the aggregates themselves as non-toxic reporter elements.

Exploring the flexible chemistry of 4-fluoro-3-nitrophenyl azide for biomolecule immobilization and bioconjugation.

Bioconjugation and functionalization of polymer surfaces are two major tasks in materials chemistry which are accomplished using a variety of coupling agents. Immobilization of biomolecules onto polymer surfaces and the construction of bioconjugates are essential requirements of many biochemical assays and chemical syntheses. Different linkers with a variety of functional groups are used for these purposes. Among them, the benzophenones, aryldiazirines, and arylazides represent the most commonly used photolinker to produce the desired chemical linkage upon their photo-irradiation. In this review, we describe the versatile applications of 4-fluoro-3-nitrophenyl azide, one of the oldest photolinkers used for photoaffinity labeling in the late 1960s. Surprisingly, this photolinker, historically known as 1-fluoro-2-nitro-4-azidobenzene (FNAB), has remained unexplored for a long time because of apprehension that FNAB forms ring-expanded dehydroazepine as a major product and hence cannot activate an inert polymer. The first evidence of photochemical activation of an inert surface by FNAB through nitrene insertion reaction was reported in 2001, and the FNAB-activated surface was found to conjugate a biomolecule without any catalyst, reagent, or modification. FNAB has distinct advantages over perfluorophenyl azide derivatives, which are contemporary nitrene-generating photolinkers, because of its simple, single-step preparation and ease of thermochemical and photochemical reactions with versatile polymers and biomolecules. Covering these aspects, the present review highlights the flexible chemistry of FNAB and its applications in the field of surface engineering, immobilization of biomolecules such as antibodies, enzymes, cells, carbohydrates, oligonucleotides, and DNA aptamers, and rapid diagnostics. Graphical Abstract An overview of the FNAB-engineered activated polymer surfaces for covalent ligation of versatile biomolecules.

Telemedicine in the Himalayas: Operational Challenges-A Preliminary Report.

INTRODUCTION: Nonavailability of quality healthcare in mountainous, isolated, inaccessible sparsely populated regions is a universal problem. In this project, remote virtual healthcare was provided at Keylong and Kaza in Himachal Pradesh (HP) in North India. This innovative public-private partnership (PPP) provides 24/7 affordable healthcare to an alpine community where people commute 20-50 km for primary and 250 km for secondary healthcare services. Following a need assessment study, an MoU was signed by Apollo Hospitals in January 2015 with the National Health Mission. The government paid for all services delivered, Capital Expenditure (CAPEX) and Operating Expenditure (OPEX). Noncompliance to auditable weekly and monthly program MIS would result in penalties. METHODS: Apollo Telehealth Services customized a turnkey solution, end-to-end, on a program management approach with measurable milestones and monthly reports. Key health issues in the region were identified. Very Small Aperture Terminals were installed amidst landslides and subzero temperatures. In February and March 2015, staff recruited from the community and local government staff were trained in Chennai. A major cultural transformation had to be effected. Urban teleconsultants were sensitized for community interaction, while deploying cutting- edge technology. RESULTS: Case records were audited. In the first 42 weeks, 2,213 teleconsults were provided, including 171 emergencies. Telelaboratory services and telehealth education programs have also been added. CONCLUSIONS: Evaluation confirms that delivering remote healthcare in inhospitable terrains in a PPP mode is effective.