Influence of fly ash on thermo-mechanical and mechanical behavior of injection molded polypropylene matrix composites.

Polypropylene composites find widespread application in industries, including packaging, plastic parts, automotive, textiles, and specialized devices like living hinges known for their remarkable flexibility. This study focuses on the manufacturing of polypropylene composite specimens by incorporating varying weight percentages of fly ash particles with polypropylene using a twin-screw extruder and injection molding machine. The composites were comprehensively tested, evaluating tensile, compressive, and flexural strength, solid-state and polymer melt properties, modulus, damping, and thermal response. The findings reveal that the compressive strength of polypropylene increases up to 2 wt% of added fly ash particles and subsequently exhibits a slight decline. Tensile strength demonstrates an increase up to 1 wt% of fly ash, followed by a decrease with a 2 wt% addition, and then a subsequent increase. Flexural strength shows improvement up to 3 wt% fly ash addition before declining. The storage modulus curve is categorized into three regions: the glassy region (up to 0 °C), the glass transition region (0-50 °C), and the glass transition region of polypropylene (>50 °C), each corresponding to different molecular motions. Weight loss curves exhibit similar trends, indicating uniform pyrolysis behavior attributed to consistent chemical bonds. Plastic degradation commences around 440 °C and concludes near 550 °C. Additionally, elemental mapping of fly ash composition identified various elements such as O, Si, K, Mg, Ca, Cl, Na, P, Al, Fe, S, Cu, Ti, and Ni. These findings offer valuable insights into the mechanical and thermal properties of polypropylene composites reinforced with fly ash, rendering them suitable for a wide range of industrial applications necessitating strength and durability across temperature variations.

NOD1 activation in 3T3-L1 adipocytes confers lipid accumulation in HepG2 cells.

AIMS: Activation of specific innate immune receptors has been characterized to modulate nutrient metabolism in individual metabolic tissue directly or indirectly via secretory molecules. Activation of the nucleotide-binding oligomerization domain-containing protein 1 (NOD1) in adipocytes has been reported to induce lipolysis linked with insulin resistance and inflammatory response. These cues are positioned to modulate metabolic action in distal organs through paracrine/endocrine signaling. Here, we assessed the role of NOD1-mediated lipolysis and inflammatory response in adipocytes to affect lipid metabolism in hepatocytes. MAIN METHODS: Human hepatoma cells (HepG2) were exposed to conditioned medium obtained from 3 T3-L1 adipocytes pretreated with NOD1 ligand (iE-DAP) and the effects on lipid accumulation, inflammation and insulin response were assessed. Activation of mechanisms leading to hepatic lipid accumulation was investigated by gene expression analysis. KEY FINDINGS: The conditioned medium from NOD1-activated 3 T3-L1 adipocytes (CM-DAP) induced lipid accumulation in HepG2 cells, driven by both lipolysis and inflammatory responses. The CM-DAP-induced lipid accumulation was independent to de novo lipogenesis and resulted from the enhanced transport of fatty acids inside and consequent increase in rate of triglycerides synthesis in hepatocytes. Moreover, CM-DAP-induced lipid accumulation instigated the expression of the markers of fatty acid oxidation and VLDL assembly for the export of triglycerides from hepatocyte. Furthermore, CM-DAP-induced lipid accumulation was associated with induction of inflammatory response and impairment of insulin signaling in HepG2 cells. SIGNIFICANCE: Beyond showing liver-specific mechanisms to adipocytes-derived factors, our findings support the involvement of adipose tissue as a mediator in NOD1-mediated biological responses to modulate hepatic metabolism.

Moringa oleifera impedes protein glycation and exerts reno-protective effects in streptozotocin-induced diabetic rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Moringa oleifera is a valued plant with wide distribution in tropical and subtropical regions of the world. It is traditionally used for the treatment of fever, infections, rheumatism, cancer, improving cardiac, renal and hepatic functions, and regulating blood glucose level. The plant has been scientifically reported for the anti-inflammatory, antioxidant, renoprotective, and anti-diabetic properties. Diabetic patients are prone to develop end-stage renal diseases due to incidence of diabetes-induced renal dysfunctions. Given that, increased production and accumulation of advanced glycation end-products (AGEs) play a conspicuous role in the development of diabetes-linked renal dysfunctions, nature-based interventions with AGEs inhibitory activity can prevent renal dysfunctions leading to renoprotection. AIM OF THE STUDY: The study aimed to demonstrate the preventive effects of the ethanolic extract of the leaves of Moringa oleifera (EEMO) on protein glycation and its further assessment for the renoprotective effect in diabetic rats. MATERIALS AND METHODS: Antiglycation activity of EEMO was assessed in vitro using bovine serum albumin. For reno-protective activity assessment, streptozotocin (STZ)-induced diabetic rats were orally treated with EEMO (100 mg/kg) or standard antiglycation agent aminoguanidine (100 mg/kg) for consecutive 8 weeks. The effects on glucose homeostasis, renal functions, and renal morphology were assessed by clinical biochemistry, molecular and histological examination. RESULTS: Presence of EEMO efficiently prevented glucose-, fructose- or methylglyoxal-mediated glycation of protein. Under in vivo set-up, compared to diabetic control rats, EEMO treatment effectively improved the glucose tolerance and body weight, and reduced the serum levels of triglycerides and total cholesterol. Additionally, EEMO administration significantly ameliorated renal dysfunctions in diabetic rats characterized by improved levels of creatinine, urea nitrogen, and uric acid in serum, and total protein level in urine, accompanied by improved kidney morphology. The diabetes-associated pro-inflammatory response characterized by upregulated expression of the inducible nitric oxide synthase (iNos), activation of nuclear factor kappa B (NF-κB) and the raised levels of inflammatory factors, interleukin-1 beta (IL-1ß) and interleukin-6 (IL-6) in renal tissue was significantly attenuated in EEMO-treated rats. Moreover, EEMO treatment diminished renal reactive oxygen species (ROS) levels in diabetic animals. CONCLUSIONS: Our study demonstrated that EEMO prevented AGEs formation and ameliorated renal dysfunctions in diabetic rats by blocking inflammatory/oxidative pathways. Our observations justify M. oleifera as a potential source of therapeutic interventions for diabetic nephropathy management.

Bolt: a New Age Peptide Search Engine for Comprehensive MS/MS Sequencing Through Vast Protein Databases in Minutes.

Recent increases in mass spectrometry speed, sensitivity, and resolution now permit comprehensive proteomics coverage. However, the results are often hindered by sub-optimal data processing pipelines. In almost all MS/MS peptide search engines, users must limit their search space to a canonical database due to time constraints and q value considerations, but this typically does not reflect the individual genetic variations of the organism being studied. In addition, engines will nearly always assume the presence of only fully tryptic peptides and limit PTMs to a handful. Even on high-performance servers, these search engines are computationally expensive, and most users decide to dial back their search parameters. We present Bolt, a new cloud-based search engine that can search more than 900,000 protein sequences (canonical, isoform, mutations, and contaminants) with 41 post-translation modifications and N-terminal and C-terminal partial tryptic search in minutes on a standard configuration laptop. Along with increases in speed, Bolt provides an additional benefit of improvement in high-confidence identifications. Sixty-one percent of peptides uniquely identified by Bolt may be validated by strong fragmentation patterns, compared with 13% of peptides uniquely identified by SEQUEST and 6% of peptides uniquely identified by Mascot. Furthermore, 30% of unique Bolt identifications were verified by all three software on the longer gradient analysis, compared with only 20% and 27% for SEQUEST and Mascot identifications respectively. Bolt represents, to the best of our knowledge, the first fully scalable, cloud-based quantitative proteomic solution that can be operated within a user-friendly GUI interface. Data are available via ProteomeXchange with identifier PXD012700.

Detection and verification of 2.3 million cancer mutations in NCI60 cancer cell lines with a cloud search engine.

Today we have unprecedented access to human genomic and proteomic data that appear to be rapidly approaching our current understanding of comprehensive coverage. Combining genomic information with shotgun proteomics remains challenging due to the large increase in proteomics search space. However, making this connection between genomic and proteomic information is critical for cancer studies to vaccine development. Furthermore, as we progress towards personalized medicine, it will be essential for proteomics analysis to identify individual mutations and variants in order to fully understand protein networks and to develop personalized therapies. While these advantages are well-established, only a few studies have demonstrated the successful integration of proteomic data with large genomic input. We present and examine the abilities of Bolt, a new cloud-based proteomics search engine to search for the presence of over 2.3 million known cancer mutations in a matter of minutes while still performing a standard proteomics search that includes 31 post translational modifications. We use previously published proteomics data sets and identify mutations that are verified using genomic studies as well as previous proteomics efforts. Our results also emphasize the need to search for mutations in a comprehensive manner while still searching for both common and rare PTMs. SIGNIFICANCE: We present and examine the abilities of Bolt, a new cloud-based proteomics search engine to search for the presence of over 2.3 million known cancer mutations in a matter of minutes while still performing a standard proteomics search that includes 31 post translational modifications. No other proteomics search software can do so.

PeMtb: A Database of MHC Antigenic Peptide of Mycobacterium tuberculosis.

BACKGROUND: For design of a subunit vaccine for tuberculosis, identification of antigenic Tcell epitope is of utmost importance. Several MHC prediction server are available that can accurately predict antigenic peptide of variable lengths. However, peptides predicted from one server not necessarily are predicted form another server, thus creating a confusing situation for scientists to choose a best epitope. METHOD: Keeping the above problem in mind, we developed a comprehensive database of peptides of Mycobacterial proteins. Each protein was taken from PubMed and was run through different MHC prediction servers, with the results being compiled into one database. RESULTS: For each protein, PeMtb generates a set of three different mers of variable lengths (12 mer or 13-mer) based on their ranking; with each mer being predicted for a plethora of MHC alleles. Researcher can choose the peptide (mers) that gives best binding affinity from most of the servers. CONCLUSION: The database relieves the investigators of the painstaking task of searching various MHC prediction servers for the right epitope (T-cell epitope) for a particular Mycobacterial antigen. We trust and anticipate that PeMtb will be a practical platform for trial and computational analyses of antigenic peptides for Mycobacterium tuberculosis. All the resources and information can be accessed by PeMtb home page www.pemtb-amu.org.

Comparative Proteome Profiling during Cardiac Hypertrophy and Myocardial Infarction Reveals Altered Glucose Oxidation by Differential Activation of Pyruvate Dehydrogenase E1 Component Subunit ß.

Cardiac hypertrophy and myocardial infarction (MI) are two etiologically different disease forms with varied pathological characteristics. However, the precise molecular mechanisms and specific causal proteins associated with these diseases are obscure to date. In this study, a comparative cardiac proteome profiling was performed in Wistar rat models for diseased and control (sham) groups using two-dimensional difference gel electrophoresis followed by matrix-assisted laser desorption/ionization tandem time-of-flight mass spectrometry. Proteins were identified using Protein Pilot™ software (version 4.0) and were subjected to stringent statistical analysis. Alteration of key proteins was validated by Western blot analysis. The differentially expressed protein sets identified in this study were associated with different functional groups, involving various metabolic pathways, stress responses, cytoskeletal organization, apoptotic signaling and other miscellaneous functions. It was further deciphered that altered energy metabolism during hypertrophy in comparison to MI may be predominantly attributed to induced glucose oxidation level, via reduced phosphorylation of pyruvate dehydrogenase E1 component subunit ß (PDHE1-B) protein during hypertrophy. This study reports for the first time the global changes in rat cardiac proteome during two etiologically different cardiac diseases and identifies key signaling regulators modulating ontogeny of these two diseases culminating in heart failure. This study also pointed toward differential activation of PDHE1-B that accounts for upregulation of glucose oxidation during hypertrophy. Downstream analysis of altered proteome and the associated modulators would enhance our present knowledge regarding altered pathophysiology of these two etiologically different cardiac disease forms.

Hybrid data acquisition and processing strategies with increased throughput and selectivity: pSMART analysis for global qualitative and quantitative analysis.

Data-dependent acquisition (DDA) and data-independent acquisition strategies (DIA) have both resulted in improved understanding of proteomics samples. Both strategies have advantages and disadvantages that are well-published, where DDA is typically applied for deep discovery and DIA may be used to create sample records. In this paper, we present a hybrid data acquisition and processing strategy (pSMART) that combines the strengths of both techniques and provides significant benefits for qualitative and quantitative peptide analysis. The performance of pSMART is compared to published DIA strategies in an experiment that allows the objective assessment of DIA performance with respect to interrogation of previously acquired MS data. The results of this experiment demonstrate that pSMART creates fewer decoy hits than a standard DIA strategy. Moreover, we show that pSMART is more selective, sensitive, and reproducible than either standard DIA or DDA strategies alone.

Altered expression of platelet proteins and calpain activity mediate hypoxia-induced prothrombotic phenotype.

Oxygen-compromised environments, such as high altitude, air travel, and sports, and pathological conditions, such as solid tumors, have been suggested to be prothrombotic. Despite the indispensable role of platelets in thrombus formation, the studies linking hypoxia, platelet reactivity, and thrombus formation are limited. In the present study, platelet proteome/reactivity was analyzed to elucidate the acute hypoxia-induced prothrombotic phenotype. Rats exposed to acute simulated hypoxia (282 torr/8% oxygen) demonstrated a decreased bleeding propensity and increased platelet reactivity. Proteomic analysis of hypoxic platelets revealed 27 differentially expressed proteins, including those involved in coagulation. Among these proteins, calpain small subunit 1, a 28-kDa regulatory component for calpain function, was significantly upregulated under hypoxic conditions. Moreover, intraplatelet Ca(2+) level and platelet calpain activity were also found to be in accordance with calpain small subunit 1 expression. The inhibition of calpain activity demonstrated reversal of hypoxia-induced platelet hyperreactivity. The prothrombotic role for calpain was further confirmed by an in vivo model of hypoxia-induced thrombosis. Interestingly, patients who developed thrombosis while at extreme altitude had elevated plasma calpain activities and increased soluble P-selectin level. In summary, this study suggests that augmented calpain activity is associated with increased incidence of thrombosis under hypoxic environments.

PPAR signaling pathway is a key modulator of liver proteome in pups born to vitamin B(12) deficient rats.

Maternal nutritional deficiency in-utero is known to predict risk of complex disorders like cardiovascular disease, diabetes and many neurological disorders in the offspring and vitamin B12 is one such critical micronutrient. Here we performed 2D-DIGE followed by MALDI TOF/TOF analysis to identify proteins that are differentially expressed in liver of pups born to mothers fed vitamin B12 deficient diet vis-à-vis control diet. To further establish causality, we analyzed the effect of B12 rehabilitation at parturition on the protein levels and the phenotype in pups. We identified 38 differentially expressed proteins that were enriched in pathways involved in the regulation of amino acid, lipid and carbohydrate metabolism. Further, three enzymes in the ß-oxidation pathway (hydroxyacyl-coenzyme A dehydrogenase, medium-chain specific acyl-CoA dehydrogenase, 3-ketoacyl-CoA thiolase) were down-regulated in pups born to mothers fed vitamin B12 deficient diet. We observed age-dependent differential expression of peroxisome proliferator activated-receptor (PPAR) α and γ in the deficient pups. Interestingly, expression of 27 proteins that were differentially expressed was restored to the control levels after rehabilitation of female rats with vitamin B12 from parturition. Our study thus provides the first evidence that maternal vitamin B12 deficiency influences lipid and other micronutrient metabolism in pups through regulation of PPAR signaling pathway. BIOLOGICAL SIGNIFICANCE: Maternal vitamin B12 deficiency has been shown to predict the onset of complex disorders like atherosclerosis, type II diabetes etc. in the next generation during their adulthood. We have shown earlier that pups born to female rats fed with vitamin B12 deficient diet were obese and developed high levels of other intermediate traits such as triglycerides, cholesterol etc. that are related to the risk of diabetes and cardiovascular disorders. In this piece of work using differential proteomic approach we have identified the altered metabolic processes in the liver of vitamin B12 deficient pups. We have also documented that the proteins involved in ß-oxidation pathway are down-regulated. Further, differential expression of PPARα and PPARγ was evidently documented as the master regulator for the alteration of lipid, amino acid and carbohydrate metabolism during maternal vitamin B12 deficiency.

A systematic analysis of eluted fraction of plasma post immunoaffinity depletion: implications in biomarker discovery.

Plasma is the most easily accessible source for biomarker discovery in clinical proteomics. However, identifying potential biomarkers from plasma is a challenge given the large dynamic range of proteins. The potential biomarkers in plasma are generally present at very low abundance levels and hence identification of these low abundance proteins necessitates the depletion of highly abundant proteins. Sample pre-fractionation using immuno-depletion of high abundance proteins using multi-affinity removal system (MARS) has been a popular method to deplete multiple high abundance proteins. However, depletion of these abundant proteins can result in concomitant removal of low abundant proteins. Although there are some reports suggesting the removal of non-targeted proteins, the predominant view is that number of such proteins is small. In this study, we identified proteins that are removed along with the targeted high abundant proteins. Three plasma samples were depleted using each of the three MARS (Hu-6, Hu-14 and Proteoprep 20) cartridges. The affinity bound fractions were subjected to gelC-MS using an LTQ-Orbitrap instrument. Using four database search algorithms including MassWiz (developed in house), we selected the peptides identified at <1% FDR. Peptides identified by at least two algorithms were selected for protein identification. After this rigorous bioinformatics analysis, we identified 101 proteins with high confidence. Thus, we believe that for biomarker discovery and proper quantitation of proteins, it might be better to study both bound and depleted fractions from any MARS depleted plasma sample.