Type 1 collagen: Synthesis, structure and key functions in bone mineralization.

Collagen is a highly abundant protein in the extracellular matrix of humans and mammals, and it plays a critical role in maintaining the body's structural integrity. Type I collagen is the most prevalent collagen type and is essential for the structural integrity of various tissues. It is present in nearly all connective tissues and is the main constituent of the interstitial matrix. Mutations that affect collagen fiber formation, structure, and function can result in various bone pathologies, underscoring the significance of collagen in sustaining healthy bone tissue. Studies on type 1 collagen have revealed that mutations in its encoding gene can lead to diverse bone diseases, such as osteogenesis imperfecta, a disorder characterized by fragile bones that are susceptible to fractures. Knowledge of collagen's molecular structure, synthesis, assembly, and breakdown is vital for comprehending embryonic and foetal development and several aspects of human physiology. In this review, we summarize the structure, molecular biology of type 1 collagen, its biomineralization and pathologies affecting bone.

An efficient, catalyst-free and aqueous ethanol-mediated synthesis of 5-((2-aminothiazol-5-yl)(phenyl)methyl)-6-hydroxypyrimidine-2,4(1H,3H)-dione derivatives and their antioxidant activity.

In this study, we effectively developed a catalyst-free multicomponent synthesis of 5-((2-aminothiazol-5-yl)(phenyl)methyl)-6-hydroxypyrimidine-2,4(1H,3H)-dione derivatives employing 2-aminothiazole, N',N'-dimethyl barbituric acid/barbituric acid and different aldehydes at 80 °C in an aqueous ethanol medium (1 : 1) using group-assisted purification (GAP) chemistry. The essential characteristics of this methodology include superior green credential parameters, metal-free multicomponent synthesis, faster reaction times, greater product yields, simple product purification without column chromatography and higher product yields. All of the synthesized compounds were analyzed against the HepG2 cell line. Compounds 4j and 4k shows good anti-proliferative effects on HepG2 cells. Furthermore, the ABTS and DPPH scavenging assays were used to determine the antioxidant activity of all compounds (4a-r). In both ABTS and DPPH radical scavenging assays, compounds 4e, 4i, 4j, 4o and 4r exhibit excellent potency compared to the standard ascorbic acid.

Fabrication of ECM Mimicking Bioactive Scaffold: A Regenerative Approach for MSC Mediated Applications.

Biomaterials are feasible resources that aids to replace damaged structures in our bodies. The most biologically active flora is Aloe vera which has many bioactive compounds that are anti-inflammatory, antimicrobial, and have ECM mimicking protein content which helps in the healing of wounds and also acts as an ECM factor for stem cell homing and differentiation. The Aloe vera containing 10 w/v of gelatin was lyophilized. Scaffolds had sharper morphology, greater hydrophilic properties, and a Young's modulus of 6.28 MPa and 15.9 MPa of higher tensile strength are desirable. In tissue engineering and regenerative medicine, biologically active scaffolds have been producing hopeful outcomes in both restoration and replacement, respectively. The objective of the present investigation is to test the idea that incorporating gelatin to Aloe vera scaffolds might enhance their structure, good biocompatibility, and possibly even bioactivity. The SEM picture of the composite scaffold revealed pore walls. The scaffolds had linked pores with diameters ranging from 93 to 296 µm. Aloe vera and the matrix interact well, according to the FTIR study, which could lead to a reduction in the amount of water-binding sites and a reduction in the material's ability to absorb water. Aloe vera with 10% gelatin (AV/G) scaffold was investigated for different biological reactions of human gingival tissue mesenchymal stem cells (MSCs) in terms of cell proliferation, morphology, and cell migration. The results demonstrated the potential of the AV/G scaffold as a biomaterial that offers new insight in the field of tissue engineering.

Chick Embryo Ex Vivo Aortic Sprouting Assays for Cardiovascular Research.

Angiogenesis is the formation of new blood vesicles and is controlled by a dynamic cascade of molecular and cellular activities. The whole procedure can be replicated in vitro under chemically specified conditions by cultivating chick aortic explants in biomatrices. In this technique, angiogenesis is powered by endogenous molecules that the aorta releases to promote its outgrowth. In an ordered series of morphogenetic events, sprouting endothelial cells are strongly associated with macrophages, fibroblasts, and pericytes, recapitulating all the phases of the angiogenic process. The structural, morphologic, and molecular properties of the angiogenic process can be studied and the effectiveness of pro/antiangiogenic drugs can also be evaluated using this aortic culture. We describe in this chapter the basic procedure currently used in our laboratory to measure the angiogenic properties for cardiovascular research.

Electrooxidation of coragen-contaminated wastewater using graphite electrodes and sorbent nano-hydroxyapatite.

The degradation of coragen (C18H14N5O2BrCl2) was tested by the electrooxidation process using graphite electrodes. Further, the advantage of nano-hydroxyapatite (n-Hap), as a cost-effective nano sorbent, in the removal of bromide from coragen was examined. Three different variables such as initial pH, electrolysis time and the current density were used to analyse the effects of the electrolytic process on the degradation of coragen. During electrolysis, under various stages, the parameters such as chemical oxygen demand (COD), chloride and bromide were analysed. The maximum COD, chloride and bromide removal efficiency of 96%, 50% and 99%, respectively, at pH 5, the maximum current density of 7.5 mA cm-2 and 120 min electrolysis time were achieved. Based on the final output of this study, it can be concluded that the electrolysis process can effectively reduce COD, chloride and bromide from coragen in an aqueous medium. Further, the degradation efficiency of the coragen was confirmed through different analyses such as UV spectra, Fourier transform infrared spectroscopy and gas chromotography-mass spectrometry analyses.

Flavonoids: Classification, Function, and Molecular Mechanisms Involved in Bone Remodelling.

Flavonoids are polyphenolic compounds spotted in various fruits, vegetables, barks, tea plants, and stems and many more natural commodities. They have a multitude of applications through their anti-inflammatory, anti-oxidative, anti-carcinogenic properties, along with the ability to assist in the stimulation of bone formation. Bone, a rigid connective body tissue made up of cells embedded in a mineralised matrix is maintained by an assemblage of pathways assisting osteoblastogenesis and osteoclastogenesis. These have a significant impact on a plethora of bone diseases. The homeostasis between osteoblast and osteoclast formation decides the integrity and structure of the bone. The flavonoids discussed here are quercetin, kaempferol, icariin, myricetin, naringin, daidzein, luteolin, genistein, hesperidin, apigenin and several other flavonoids. The effects these flavonoids have on the mitogen activated protein kinase (MAPK), nuclear factor kappa ß (NF-kß), Wnt/ß-catenin and bone morphogenetic protein 2/SMAD (BMP2/SMAD) signalling pathways, and apoptotic pathways lead to impacts on bone remodelling. In addition, these polyphenols regulate angiogenesis, decrease the levels of inflammatory cytokines and play a crucial role in scavenging reactive oxygen species (ROS). Considering these important effects of flavonoids, they may be regarded as a promising agent in treating bone-related ailments in the future.

Evaluation of the angiogenic properties of Brugia malayi asparaginyl-tRNA synthetase and its mutants: A study on the molecular target for antifilarial drug development.

Brugia malayi asparaginyl-tRNA synthetase (BmAsnRS) has been identified as an immunodominant antigen and a physiocrine that mimics Interleukin-8 (IL-8) to induce chemotaxis and angiogenesis in endothelial cells. Computational analyses have shown that the N-terminal region of BmAsnRS has a novel fold, a lysine rich ß-hairpin α-helix, (FLIRTKKDGKQIWE) which is similar to that present in IL-8 chemokine, CXCR1. This novel fold is involved in tRNA binding and is integral for the manifestation of the disease, lymphatic filariasis (LF). Drug discovery programmes carried out so far for LF have not been successful because of the target (BmAsnRS) resistance due to the disease-associated mutation. Mutations in AARS targets have been shown to correlate with several diseases. However, no disease-associated mutational studies have been carried out for LF. BmAsnRS has been an established target for LF. It was proposed, therefore, to study the effect of single point mutations in BmAsnRS so as to elucidate the molecular target. An understanding of the molecular consequences of mutations will provide insight into how resistance develops in addition to the identification of the likely resistance-conferring mutations. Three mutants were, therefore, generated by site-directed mutagenesis using CUPSAT server and their angiogenic properties evaluated. Cytometric analysis of the mutants on endothelial cell cycle was also carried out. CUPSAT prediction of protein stability upon point mutations reveal that two mutants generated are likely resistance-conferring mutations. All the three mutants show significant reduction in their angiogenic properties and reduction in the DNA content in the cells of S and G2/M phases thus showing altered function of the gene encoding the drug target. The resistance- conferring mutants, however, show angiogenic properties nearer to the wild type protein, BmAsnRS. Future work on designing newer drugs may take into consideration these drug resistance-conferring mutations.

LncRNA MALAT1 Promotes Tumor Angiogenesis by Regulating MicroRNA-150-5p/VEGFA Signaling in Osteosarcoma: In-Vitro and In-Vivo Analyses.

The present study aims to analyze the expression of long noncoding RNA (lncRNA) metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) in human osteosarcoma (OS) cells and to investigate its role in OS-induced angiogenesis. MALAT1 expression in OS cells was significantly higher than in normal osteoblasts. The functional analysis indicated that MALAT1 appears to enhance OS-induced angiogenesis, in vitro and in vivo analyses, endothelial cell proliferation and migration, chick embryo angiogenesis assay, and zebrafish xenograft model. Mechanistically, silencing MALAT1 downregulated vascular endothelial growth factor A (VEGFA) expression and upregulated miR-150-5p expression in OS cells, and MALAT1-mediated angiogenic induction by VEGFA in OS microenvironment. Moreover, MALAT1 directly targeted miR-150-5p and miR-150-5p directly target VEGFA in OS. Overexpression of miR-150-5p downregulates VEGFA expression in OS. More notably, we showed that MALAT1 induced angiogenesis in OS microenvironment by upregulating the expression of VEGFA via targeting miR-150-5p. Overall, our findings suggest that MALAT1 promotes angiogenesis by regulating the miR-150-5p/VEGFA signaling in OS microenvironment. The findings of the molecular mechanisms of MALAT1 in tumor angiogenesis offer a new viewpoint on OS treatment.

Recent Insight on the Non-coding RNAs in Mesenchymal Stem Cell-Derived Exosomes: Regulatory and Therapeutic Role in Regenerative Medicine and Tissue Engineering.

Advances in the field of regenerative medicine and tissue engineering over the past few decades have paved the path for cell-free therapy. Numerous stem cell types, including mesenchymal stem cells (MSCs), have been reported to impart therapeutic effects via paracrine secretion of exosomes. The underlying factors and the associated mechanisms contributing to these MSC-derived exosomes' protective effects are, however, poorly understood, limiting their application in the clinic. The exosomes exhibit a diversified repertoire of functional non-coding RNAs (ncRNAs) and have the potential to transfer these biologically active transcripts to the recipient cells, where they are found to modulate a diverse array of functions. Altered expression of the ncRNAs in the exosomes has been linked with the regenerative potential and development of various diseases, including cardiac, neurological, skeletal, and cancer. Also, modulating the expression of ncRNAs in these exosomes has been found to improve their therapeutic impact. Moreover, many of these ncRNAs are expressed explicitly in the MSC-derived exosomes, making them ideal candidates for regenerative medicine, including tissue engineering research. In this review, we detail the recent advances in regenerative medicine and summarize the evidence supporting the altered expression of the ncRNA repertoire specific to MSCs under different degenerative diseases. We also discuss the therapeutic role of these ncRNA for the prevention of these various degenerative diseases and their future in translational medicine.

Bio-hybrid hydrogel comprising collagen-capped silver nanoparticles and melatonin for accelerated tissue regeneration in skin defects.

Hydrogel-based drug delivery systems have emerged as a promising platform for chronic tissue defects owing to their inherent ability to inhibit pathogenic infection and accelerate rapid tissue regeneration. Here, we fabricated a stable bio-hybrid hydrogel system comprising collagen, aminated xanthan gum, bio-capped silver nanoparticles and melatonin with antimicrobial, antioxidant and anti-inflammatory properties. Highly colloidal bio-capped silver nanoparticles were synthesized using collagen as a reducing cum stabilizing agent for the first time while aminated xanthan gum was synthesized using ethylenediamine treatment on xanthan gum. The synthesized bio-hybrid hydrogel exhibits better gelation, surface morphology, rheology and degelation properties. In vitro assessment of bio-hybrid hydrogel demonstrates excellent bactericidal efficiency against both common wound and multidrug-resistant pathogens and biocompatibility properties. In vivo animal studies demonstrate rapid tissue regeneration, collagen deposition and angiogenesis at the wound site predominantly due to the synergistic effect of silver nanoparticles and melatonin in the hydrogel. This study paves the way for developing biologically functional bio-nano hydrogel systems for promoting effective care for various ailments, including infected chronic wounds.

Application of Artificial Neural Network as a nonhazardous alternative on kinetic analysis and modeling for green synthesis of cobalt nanocatalyst from Ocimum tenuiflorum.

The present paper is dedicated to analyze non-hazardous kinetic behaviour and modelling of green synthesized cobalt nanocatalyst (CoNCs), using an Artificial Neural Network (ANN). In order to supplement the trace metal in other applications, CoNCs were rapidly synthesized with a Cobalt sulphate solution at room temperature between 30 and 35 ºC at pH 7.2 under less reaction time. The Levenberg - Marquardt algorithm (LM) is used to investigate the experimental values by applying ANN. The results of variance using logistic ANN model depicts that the maximum nanoparticles were synthesized at its optimized stipulation of 0.5 h stirring time, 25 mL volume of extract and 20 mL volume of cobalt sulphate. The developed ANN model proved to be an efficient size determining tool in the biosynthesis of cobalt nanocatalyst. Experimental behavior using potentiometric analysis confirms that the linearity in CoNCs formation and size coincides (5-38 nm)with the predicted values of the ANN model. Techno economic analysis proved that, green synthesis reduced 30-40% in raw material cost and 60% in energy consumption.

Bioactive Zinc(II) complex incorporated PCL/gelatin electrospun nanofiber enhanced bone tissue regeneration.

Bone tissue regeneration is augmented by biocompatible nanofiber scaffolds, that supports reliable and enhanced bone formation. Zinc is an essential mineral that is vital for routine skeletal growth and it emerges to be able to improve bone regeneration. Phytochemicals, particularly flavonoids have achieved prominent interest for their therapeutic ability, they have demonstrated promising effects on bone by encouraging osteoblastogenesis, which finally leads to bone formation. In this study, we have synthesized bioactive zinc(II) quercetin complex material and used for nanofibers scaffold fabrication to enhance bone tissue regeneration property. Two derivatives of zinc(II) quercetin complexes [(Zn(quercetin) (H2O)2) (Zn+Q), and Zn(quercetin)(phenanthroline) (Zn+Q(PHt)) have been synthesized and characterized using UV-Visible spectrophotometer and Fourier Transform-IR spectroscopy. The UV-Visible absorption and IR spectra prove the B-ring chelation of the flavonoid quercetin to zinc(II) rather C-ring chelation. The potential ability of the above synthesized metal complexes on osteogenesis and angiogenesis have been studied. Besides the bioactivity of the metal complexes, the control quercetin has also been examined. The chick embryo chorioallantoic membrane (CAM) assay demonstrated that the angiogenic parameters were increased by the (Zn+Q(PHt)) complex. Amongst, (Zn+Q(PHt)) complex showed significant activity and thereby this complex has been further examined for the bone tissue activity by incorporating the complex into a nanofiber through electrospinning method. At the molecular level, Runx2, mRNA and protein, ALP and type 1 collagen mRNAs, and osteoblast-specific microRNA, pre-mir-15b were examined using real time RT-PCR and Western blot assay. Histology studies showed that the (PCL/gelatin/Zn+Q(PHt)) was biocompatibility in-ovo. Overall, the present study showed that quercetin-zinc complex (Zn+Q(PHt)) incorporated into PCL/gelatin nanofiber can act as a pharmacological agent for treating bone associated defects and promote bone regeneration.

Identification and analysis of circulating long non-coding RNAs with high significance in diabetic cardiomyopathy.

Diabetic cardiomyopathy (DCM) lacks diagnostic biomarkers. Circulating long non-coding RNAs (lncRNAs) can serve as valuable diagnostic biomarkers in cardiovascular disease. To seek potential lncRNAs as a diagnostic biomarker for DCM, we investigated the genome-wide expression profiling of circulating lncRNAs and mRNAs in type 2 diabetic db/db mice with and without DCM and performed bioinformatic analyses of the deregulated lncRNA-mRNA co-expression network. Db/db mice had obesity and hyperglycemia with normal cardiac function at 6 weeks of age (diabetes without DCM) but with an impaired cardiac function at 20 weeks of age (DCM) on an isolated Langendorff apparatus. Compared with the age-matched controls, 152 circulating lncRNAs, 127 mRNAs and 3355 lncRNAs, 2580 mRNAs were deregulated in db/db mice without and with DCM, respectively. The lncRNA-mRNA co-expression network analysis showed that five deregulated lncRNAs, XLOC015617, AK035192, Gm10435, TCR-α chain, and MouselincRNA0135, have the maximum connections with differentially expressed mRNAs. Bioinformatic analysis revealed that these five lncRNAs were highly associated with the development and motion of myofilaments, regulation of inflammatory and immune responses, and apoptosis. This finding was validated by the ultrastructural examination of myocardial samples from the db/db mice with DCM using electron microscopy and changes in the expression of myocardial tumor necrosis factor-α and phosphorylated p38 mitogen-activated protein kinase in db/db mice with DCM. These results indicate that XLOC015617, AK035192, Gm10435, TCR-α chain, and MouselincRNA0135 are crucial circulating lncRNAs in the pathogenesis of DCM. These five circulating lncRNAs may have high potential as a diagnostic biomarker for DCM.

Assessment of human ovarian follicular fluid derived mesenchymal stem cells in chitosan/PCL/Zn scaffold for bone tissue regeneration.

Bone tissue engineering compasses the use of mesenchymal stem cells (MSCs) along with engineered biomaterial construct to augment bone regeneration. Till now, MSCs were isolated from various sources and used in cellular constructs. For the first time, in this study, MSCs were isolated from human Ovarian Follicular Fluid (OFF) and characterized by CD 44+ and CD 105+ markers via confocal microscopy and flow cytometry. Additionally, MSCs stemness, proliferation and colony-forming unit ability, multi-lineage differentiation potential were also studied. To test its suitability for bone tissue engineering applications, we grew the MSCs with the conditioned medium obtained from biocomposite scaffold by fusing a natural polymer, Chitosan (CS) and a synthetic polymer, Polycaprolactone (PCL) and the scaffold were coated with Zinc divalent ions to impart osteogenic properties. The physico-chemical characterization of scaffold, such as FTIR, XRD, and SEM studies was carried out. The biological characterization showed that the scaffolds were compatible with MSCs and promoted osteoblast differentiation which was confirmed at both cellular and molecular levels. The cellular construct increased calcium deposition, analyzed by alizarin red staining and ALP activity at cellular level. At the molecular level, the osteoblast markers expression such as Runx2 and type 1 collagen mRNAs, and osteonectin (ON) and osteocalcin (OC) secretory proteins were increased in the presence of scaffold. Overall, the current study recommends that MSCs can be easily obtained from human waste OFF, and grown in standard in vitro conditions. Successful growth of such MSCs with CS/PCL/Zn scaffold opens new avenues in utilizing the cell source for bone tissue engineering.

Cloning, expression and purification of recombinant dermatopontin in Escherichia coli.

Dermatopontin (DPT) is an extracellular matrix (ECM) protein with diversified pharmaceutical applications. It plays important role in cell adhesion/migration, angiogenesis and ECM maintenance. The recombinant production of this protein will enable further exploration of its multifaceted functions. In this study, DPT protein has been expressed in Escherichia coli (E.coli) aiming at cost effective recombinant production. The E.coli GJ1158 expression system was transformed with constructed recombinant vector (pRSETA-DPT) and protein was expressed as inclusion bodies on induction with NaCl. The inclusion bodies were solubilised in urea and renaturation of protein was done by on-column refolding procedure in Nickel activated Sepharose column. The refolded Histidine-tagged DPT protein was purified and eluted from column using imidazole and its purity was confirmed by analytical techniques. The biological activity of the protein was confirmed by collagen fibril assay, wound healing assay and Chorioallantoic Membrane (CAM) angiogenesis assay on comparison with standard DPT. The purified DPT was found to enhance the collagen fibrillogenesis process and improved the migration of human endothelial cells. About 73% enhanced wound closure was observed in purified DPT treated endothelial cells as compared to control. The purified DPT also could induce neovascularisation in the CAM model. At this stage, scaling up the production process for DPT with appropriate purity and reproducibility will have a promising commercial edge.

Feasibility of biodiesel production from waste cooking oil: lab-scale to pilot-scale analysis.

In the last few decades, consciousness of fossil fuel resources and increased environmental concerns have given the need for emergence of alternative fuel. Biodiesel is one of the potential renewable energies produced from edible and non-edible biomass which could be a potential alternative for petrol-derived diesel. In this work, initially the process of biodiesel production from waste cooking oil using potassium hydroxide as catalyst and the process parameters were studied in laboratory. The maximum biodiesel yield of 97% was attained at 75 °C with 1 wt% catalyst concentration and oil-methanol molar ratio of 1:06 at 350 rpm and 90 min. Also, these process conditions were used for biodiesel production in the pilot plant and obtained 97% yield. Overall, mass balance for the pilot plant was studied to analyze the product yield loss. The fatty acid methyl ester formation in the plant was confirmed by characterization with FTIR and 1H NMR. Further, the quality of biodiesel produced was compared for its physiochemical properties with the ASTM standards.

Prophylactic supplementation of 20-HETE ameliorates hypoxia/reoxygenation injury in pulmonary vascular endothelial cells by inhibiting apoptosis.

Hypoxia reoxygenation (HR) injury perturbs structural and functional syncytium in lung tissues. It is commonly implicated in conditions such as stroke, lung transplant or severe pneumonia. In the present study, we investigated the cytoprotective action of 20-hydroxyeicosatetraenoic acid (20-HETE) on pulmonary vascular endothelial cells (PMVECs) under normoxic and hypoxic niche followed by HR. 20-HETE pretreatment showed a protective effect at a concentration of 1µM as there was a marked increase (20%) in the cell viability compared to control and HR groups. Pretreatment of 20-HETE in HR induced injury decreased ROS production dictated its antioxidant property. Similarly, SOD and ATP levels were also downregulated by 20-HETE pretreatment. Cell apoptosis was detected by TUNEL assay, Acridine orange, and procaspase-3 cleavage, caspase-3 activity assay, respectively. JC-1 mitochondrial membrane potential assay and protein expression pattern of BCL-2, and BAD phosphorylation status were examined. The results showed that HR induced significant increase of apoptotic PMVECs, while 20-HETE pretreatment attenuated the effects. Further, 20-HETE pretreatment activated PI3K/Akt and HIF-1α signaling pathway to exhibit its protective effects against HR-induced oxidative stress and apoptosis. Overall, the results concluded the potent antioxidant role of 20-HETE in aiding cytoprotection upon HR injury.

Genome-wide differential expression profiling of lncRNAs and mRNAs associated with early diabetic cardiomyopathy.

Diabetic cardiomyopathy is one of the main causes of heart failure and death in patients with diabetes. There are no effective approaches to preventing its development in the clinic. Long noncoding RNAs (lncRNA) are increasingly recognized as important molecular players in cardiovascular disease. Herein we investigated the profiling of cardiac lncRNA and mRNA expression in type 2 diabetic db/db mice with and without early diabetic cardiomyopathy. We found that db/db mice developed cardiac hypertrophy with normal cardiac function at 6 weeks of age but with a decreased diastolic function at 20 weeks of age. LncRNA and mRNA transcripts were remarkably different in 20-week-old db/db mouse hearts compared with both nondiabetic and diabetic controls. Overall 1479 lncRNA transcripts and 1109 mRNA transcripts were aberrantly expressed in 6- and 20-week-old db/db hearts compared with nondiabetic controls. The lncRNA-mRNA co-expression network analysis revealed that 5 deregulated lncRNAs having maximum connections with differentially expressed mRNAs were BC038927, G730013B05Rik, 2700054A10Rik, AK089884, and Daw1. Bioinformatics analysis revealed that these 5 lncRNAs are closely associated with membrane depolarization, action potential conduction, contraction of cardiac myocytes, and actin filament-based movement of cardiac cells. This study profiles differently expressed lncRNAs in type 2 mice with and without early diabetic cardiomyopathy and identifies BC038927, G730013B05Rik, 2700054A10Rik, AK089884, and Daw1 as the core lncRNA with high significance in diabetic cardiomyopathy.

Microarray analysis of long non-coding RNA and mRNA expression profiles in diabetic cardiomyopathy using human induced pluripotent stem cell-derived cardiomyocytes.

AIM: This study aims to investigate the altered expression signature of long non-coding RNAs, mRNAs and deregulated pathways related to diabetic cardiomyopathy disease pathogenesis. METHOD: We utilize the previously established in vitro diabetic cardiomyopathy model of human induced pluripotent stem cell-derived human cardiomyocytes to perform long non-coding RNA and mRNA expression analysis on glucose (11 mM), endothelin-1 (10 nM) and cortisol (1 µM) stimulated human induced pluripotent stem cell-derived human cardiomyocytes to interrogate diabetic cardiomyopathy associated RNA expression profile. RESULT: Out of 20,730 mRNAs and 40,173 long non-coding RNAs being screened, 2046 long non-coding RNAs and 1582 mRNAs were differentially regulated (fold change > 2, p < 0.05) between diabetic cardiomyopathy and control group, of which more than half were intergenic and antisense long non-coding RNAs. Most of the coding transcripts were associated with processes like inflammation, structural reorganization, metabolism, smooth muscle contraction, focal adhesion and repair contributing towards the development of diabetic cardiomyopathy. The subgroup analysis further revealed 411 long non-coding RNAs being co-expressed with neighbouring genes. However, our coding-non-coding co-expression analysis showed an overall 48,155 co-expression network connections. In addition to that, the long non-coding RNAs with highest network connections were profoundly enriched for focal adhesion, cell-matrix adhesion and muscle contraction. CONCLUSION: These results provide comprehensive data about the pathways and regulatory mechanisms associated with diabetic cardiomyopathy and indicate that long non-coding RNAs may play a crucial role in diabetic cardiomyopathy.

Immune Response to Brugia malayi Asparaginyl-tRNA Synthetase in Balb/c Mice and Human Clinical Samples of Lymphatic Filariasis.

Background: Lymphatic filariasis (LF) is a global health problem, with a peculiar nature of parasite-specific immunosuppression that promotes long-term pathology and disability. Immune modulation in the host by parasitic antigens is an integral part of this disease. The current study attempts to dissect the immune responses of aminoacyl-tRNA synthetases (AARS) with emphasis on Brugia malayi asparaginyl-tRNA synthetase (BmAsnRS), since it is one among the highly expressed excretory/secretory proteins expressed in all stages of the parasite life cycle, whereas its role in filarial pathology has not been elaborately studied. Methods and Results: In this study, recombinant BmAsnRS (rBmAsnRS) immunological effects were studied in semipermissive filarial animal model Balb/c mice and on clinically defined human samples for LF. In mice study, humoral responses showed considerable titer levels with IgG2a isotype followed by IgG2b and IgG1. Immunoreactivity studies with clinical samples showed significant humoral responses especially in endemic normal with marked levels of IgG1 and IgG2 followed by IgG3. The cell-mediated immune response, evaluated by splenocytes and peripheral blood mononuclear cells proliferation, did not yield significant difference when compared with control groups. Cytokine profiling and qRT-PCR analysis of mice samples immunized with rBmAsnRS showed elevated levels of IFN-γ, IL-10, inhibitory factor-cytotoxic T lymphocyte-associated protein-A (CTLA-4) and Treg cell marker-Forkhead Box P3 (FoxP3). Conclusions: These observations suggest that rBmAsnRS has immunomodulatory effects with modified Th2 response along with suppressed cellular proliferation indicating the essence of this molecule for immune evasion by the parasite.