New perspectives of nanoneuroprotection, nanoneuropharmacology and nanoneurotoxicity: modulatory role of amino acid neurotransmitters, stress, trauma, and co-morbidity factors in nanomedicine

Recent advancement in nanomedicine suggests that nanodrug delivery using nanoformulation of drugs or use of nanoparticles for neurodiagnostic and/or neurotherapeutic purposes results in superior effects than the conventional drugs or parent compounds. This indicates a bright future for nanomedicine in treating neurological diseases in clinics. However, the effects of nanoparticles per se in inducing neurotoxicology by altering amino acid neurotransmitters, if any, are still being largely ignored. The main aim of nanomedicine is to enhance the drug availability within the central nervous system (CNS) for greater therapeutic successes. However, once the drug together with nanoparticles enters into the CNS compartments, the fate of nanomaterial within the brain microenvironment is largely remained unknown. Thus, to achieve greater success in nanomedicine, our knowledge in understanding nanoneurotoxicology in detail is utmost important. In addition, how co-morbidity factors associated with neurological disease, e.g., stress, trauma, hypertension or diabetes, may influence the neurotherapeutic potentials of nanomedicine are also necessary to explore the details. Recent research in our laboratory demonstrated that engineered nanoparticles from metals or titanium nanowires used for nanodrug delivery in laboratory animals markedly influenced the CNS functions and alter amino acid neurotransmitters in healthy animals. These adverse reactions of nanoparticles within the CNS are further aggravated in animals with different co-morbidity factors viz., stress, diabetes, trauma or hypertension. This effect, however, depends on the composition and dose of the nanomaterials used. On the other hand, nanodrug delivery by TiO₂ nanowires enhanced the neurotherapeutic potential of the parent compounds in CNS injuries in healthy animals and do not alter amino acids balance. However, in animals with any of the above co-morbidity factors, high dose of nanodrug delivery is needed to achieve some neuroprotection. Taken together, it appears that while exploring new nanodrug formulations for neurotherapeutic purposes, co-morbidly factors and composition of nanoparticles require more attention. Furthermore, neurotoxicity caused by nanoparticles per se following nanodrug delivery may be examined in greater detail with special regards to changes in amino acid balance in the CNS.     

Minocycline,levodopa and MnTMPyP induced changes in the mitochondrial proteome profile of MPTP and maneb and paraquat mice models of Parkinson's disease

Mitochondrial dysfunction is the foremost perpetrator of the nigrostriatal dopaminergic neurodegeneration leading to Parkinson's disease (PD). However, the roles played by majority of the mitochondrial proteins in PD pathogenesis have not yet been deciphered. The present study investigated the effects of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and combined maneb and paraquat on the mitochondrial proteome of the nigrostriatal tissues in the presence or absence of minocycline, levodopa and manganese (III) tetrakis (1-methyl-4-pyridyl) porphyrin (MnTMPyP). The differentially expressed proteins were identified and proteome profiles were correlated with the pathological and biochemical anomalies induced by MPTP and maneb and paraquat. MPTP altered the expression of twelve while combined maneb and paraquat altered the expression of fourteen proteins. Minocycline, levodopa and MnTMPyP, respectively, restored the expression of three, seven and eight proteins in MPTP and seven, eight and eight proteins in maneb- and paraquat-treated groups. Although levodopa and MnTMPyP rescued from MPTP- and maneb- and paraquat-mediated increase in the microglial activation and decrease in manganese-superoxide dismutase expression and complex I activity, dopamine content and number of dopaminergic neurons, minocycline defended mainly against maneb- and paraquat-mediated alterations. The results demonstrate that MPTP and combined maneb and paraquat induce mitochondrial dysfunction and microglial activation and alter the expression of a bunch of mitochondrial proteins leading to the nigrostriatal dopaminergic neurodegeneration and minocycline, levodopa or MnTMPyP variably offset scores of such changes.     

Application of higher order statistics for atrial arrhythmia classification

Atrial fibrillation (AF) and atrial flutter (AFL) are the two common atrial arrhythmia encountered in the clinical practice. In order to diagnose these abnormalities the electrocardiogram (ECG) is widely used. The conventional linear time and frequency domain methods cannot decipher the hidden complexity present in these signals. The ECG is inherently a non-linear, non-stationary and non-Gaussian signal. The non-linear models can provide improved results and capture minute variations present in the time series. Higher order spectra (HOS) is a non-linear dynamical method which is highly rugged to noise. In the present study, the performances of two methods are compared: (i) 3rd order HOS cumulants and (ii) HOS bispectrum. The 3rd order cumulant and bispectrum coefficients are subjected to dimensionality reduction using independent component analysis (ICA) and classified using classification and regression tree (CART), random forest (RF), artificial neural network (ANN) and k-nearest neighbor (KNN) classifiers to select the best classifier. The ICA components of cumulant coefficients have provided the average accuracy, sensitivity, specificity and positive predictive value of 99.50%, 100%, 99.22% and 99.72% respectively using KNN classifier. Similarly, the ICA components of HOS bispectrum coefficients have yielded the average accuracy, sensitivity, specificity and PPV of 97.65%, 98.16%, 98.75% and 99.53% respectively using KNN. So, the ICA performed on the 3rd order HOS cumulants coupled with KNN classifier performed better than the HOS bispectrum method. The proposed methodology is robust and can be used in mass screening of cardiac patients.     

Massive Sorghum Collection Genotyped with SSR Markers to Enhance Use of Global Genetic Resources

Large ex situ collections require approaches for sampling manageable amounts of germplasm for in-depth characterization and use. We present here a large diversity survey in sorghum with 3367 accessions and 41 reference nuclear SSR markers. Of 19 alleles on average per locus, the largest numbers of alleles were concentrated in central and eastern Africa. Cultivated sorghum appeared structured according to geographic regions and race within region. A total of 13 groups of variable size were distinguished. The peripheral groups in western Africa, southern Africa and eastern Asia were the most homogeneous and clearly differentiated. Except for Kafir, there was little correspondence between races and marker-based groups. Bicolor, Caudatum, Durra and Guinea types were each dispersed in three groups or more. Races should therefore better be referred to as morphotypes. Wild and weedy accessions were very diverse and scattered among cultivated samples, reinforcing the idea that large gene-flow exists between the different compartments. Our study provides an entry to global sorghum germplasm collections. Our reference marker kit can serve to aggregate additional studies and enhance international collaboration. We propose a core reference set in order to facilitate integrated phenotyping experiments towards refined functional understanding of sorghum diversity.     

Rosiglitazone Treatment of Type 2 Diabetic db/db Mice Attenuates Urinary Albumin and Angiotensin Converting Enzyme 2 Excretion

Alterations within the renal renin angiotensin system play a pivotal role in the development and progression of cardiovascular and renal disease. Angiotensin converting enzyme 2 (ACE2) is highly expressed in renal tubules and has been shown to be renoprotective in diabetes. The protease, a disintegrin and metalloprotease (ADAM) 17, is involved in the ectodomain shedding of several transmembrane proteins including ACE2. Renal ACE2 and ADAM17 were significantly increased in db/db mice compared to controls. We investigated the effect of the insulin sensitizer, rosiglitazone, on albuminuria, renal ADAM17 protein expression and ACE2 shedding in db/db diabetic mice. Rosiglitazone treatment of db/db mice normalized hyperglycemia, attenuated renal injury and decreased urinary ACE2 and renal ADAM17 protein expression. Urinary excreted ACE2 is enzymatically active. Western blot analysis of urinary ACE2 demonstrated two prominent immunoreactive bands at approximately 70 & 90 kDa. The predominant immunoreactive band is approximately 20 kDa shorter than the one demonstrated for kidney lysate, indicating possible ectodomain shedding of active renal ACE2 in the urine. Therefore, it is tempting to speculate that renoprotection of rosiglitazone could be partially mediated via downregulation of renal ADAM17 and ACE2 shedding. In addition, there was a positive correlation between blood glucose, urinary albumin, plasma glucagon, and triglyceride levels with urinary ACE2 excretion. In conclusion, urinary ACE2 could be used as a sensitive biomarker of diabetic nephropathy and for monitoring the effectiveness of renoprotective medication.     

CRYβA3/A1-Crystallin Knockout Develops Nuclear Cataract and Causes Impaired Lysosomal Cargo Clearance and Calpain Activation

βA3/A1-crystallin is an abundant structural protein of the lens that is very critical for lens function. Many different genetic mutations have been shown to associate with different types of cataracts in humans and in animal models. βA3/A1-crystallin has four Greek key-motifs that organize into two crystallin domains. It shown to bind calcium with moderate affinity and has putative calcium-binding site. Other than in the lens, βA3/A1 is also expressed in retinal astrocytes, retinal pigment epithelial (RPE) cells, and retinal ganglion cells. The function of βA3/A1-crystallin in the retinal cell types is well studied; however, a clear understanding of the function of this protein in the lens has not yet been established. In the current study, we generated the βA3/A1-crystallin knockout (KO) mouse and explored the function of βA3/A1-crystallin in lens development. Our results showed that βA3-KO mice develop congenital nuclear cataract and exhibit persistent fetal vasculature condition. At the cellular level KO lenses show defective lysosomal clearance and accumulation of nuclei, mitochondria, and autophagic cargo in the outer cortical region of the lens. In addition, the calcium level and the expression and activity of calpain-3 were increased in KO lenses. Taken together, these results suggest the lack of βA3-crystallin function in lenses, alters calcium homeostasis which in turn causes lysosomal defects and calpain activation. These defects are responsible for the development of nuclear cataract in KO lenses.     

<Superscript>1</Superscript>H, <Superscript>15</Superscript>N,and <Superscript>13</Superscript>C resonance assignments of <Emphasis Type="Italic">Staphylococcus aureus</Emphasis> extracellular adherence protein domain 4

The pathogenic bacterium Staphylococcus aureus has evolved to actively evade many aspects of the human innate immune system by expressing a series of secreted inhibitory proteins. Among these, the extracellular adherence protein (Eap) has been shown to inhibit the classical and lectin pathways of the complement system. By binding to complement component C4b, Eap is able to inhibit formation of the CP/LP C3 pro-convertase. Secreted full-length, mature Eap consists of four ~98 residue domains, all of which adopt a similar beta-grasp fold, and are connected through a short linker region. Through multiple biochemical approaches, it has been determined that the third and fourth domains of Eap are responsible for C4b binding. Here we report the backbone and side-chain resonance assignments of the 11.3 kDa fourth domain of Eap. The assignment data has been deposited in the BMRB database under the accession number 26726.     

Mechanism of Zinc absorption in plants: uptake,transport, translocation and accumulation

Zinc (Zn) is an essential micronutrient for plants and animals. Unfortunately, deficiency of Zn in humans has increased on a global scale. The main reason of this micronutrient deficiency is dietary intakes of food with low Zn levels. Adoption of biofortification approaches would result in Zn enrichment of target tissue to a considerable extent. However, there is a basic need to understand Zn absorption mechanisms in plants prior to exploitation of such practical approaches. Zn absorption is a complex physiological trait which is mainly governed by Zn transporters and metal chelators of plant system. Plant growth stage, edaphic factors, season etc. also influence Zn efficiency of particular species. Molecular studies in Zn hyperaccumulators have already demonstrated the participation of specific Zn transporters, vacuolar sequestration and detoxification mechanisms in maintenance of Zn homeostasis. These have been described in detail in present review and provide opportunities for utilization in biofortification programmes. However, issues such as lesser bioavailability of Zn in target organ, uptake of toxic divalent cations (Cd, Ni, Pb, As etc.) along with Zn, sink activity and dilution in Zn concentration in response to sink number etc. in biofortified crops need further investigation. In order to design novel strategy in biofortification programmes, future researches should focus on physiological performance and yield penalties in concerned crop, metabolic load in term of organic acid production and crosstalk of Zn with other mineral nutrients under low and high Zn conditions.     

Understanding and modeling retention of mammalian cells in fluidized bed centrifuges

Within the last decade, fully disposable centrifuge technologies, fluidized‐bed centrifuges (FBC), have been introduced to the biologics industry. The FBC has found a niche in cell therapy where it is used to collect, concentrate, and then wash mammalian cell product while continuously discarding centrate. The goal of this research was to determine optimum FBC conditions for recovery of live cells, and to develop a mathematical model that can assist with process scaleup. Cell losses can occur during bed formation via flow channels within the bed. Experimental results with the kSep400 centrifuge indicate that, for a given volume processed: the bed height (a bed compactness indicator) is affected by RPM and flowrate, and dead cells are selectively removed during operation. To explain these results, two modeling approaches were used: (i) equating the centrifugal and inertial forces on the cells (i.e., a force balance model or FBM) and (ii) a two‐phase computational fluid dynamics (CFD) model to predict liquid flow patterns and cell retention in the bowl. Both models predicted bed height vs. time reasonably well, though the CFD model proved more accurate. The flow patterns predicted by CFD indicate a Coriolis‐driven flow that enhances uniformity of cells in the bed and may lead to cell losses in the outflow over time. The CFD‐predicted loss of viable cells and selective removal of the dead cells generally agreed with experimental trends, but did over‐predict dead cell loss by up to 3‐fold for some of the conditions. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:1520–1530, 2016     

DrRecQ regulates guanine quadruplex DNA structure dynamics and its impact on radioresistance in Deinococcus radiodurans

The GC‐rich genome of Deinococcus radiodurans contains a very high density of putative guanine quadruplex (G4) DNA motifs and its RecQ (drRecQ) was earlier characterized as a 3′→5′ dsDNA helicase. We saw that N‐Methyl mesoporphyrin IX (NMM), a G4 DNA binding drug affected normal growth as well as the gamma radiation resistance of the wild‐type bacterium. Interestingly, NMM treatment and recQ deletion showed additive effect on normal growth but there was no effect of NMM on gamma radiation resistance of recQ mutant. The recombinant drRecQ showed ~400 times higher affinity to G4 DNA (K_d = 11.74 ± 1.77 nM) as compared to dsDNA (K_d = 4.88 ± 1.30 µM). drRecQ showed ATP independent helicase function on G4 DNA, which was higher than ATP‐dependent helicase activity on dsDNA. Unlike wild‐type cells that sparingly stained for G4 structure with Thioflavin T (ThT), recQ mutant showed very high‐density of ThT fluorescence foci on DNA indicating an important role of drRecQ in regulation of G4 DNA structure dynamics in vivo. These results together suggested that drRecQ is an ATP independent G4 DNA helicase that plays an important role in the regulation of G4 DNA structure dynamics and its impact on radioresistance in D. radiodurans.