Morphological and biochemical responses of Vicia faba (faba beans) grown on fly ash amended soil in the presence of Rhizobium leguminosarum and arbuscular mycorrhizal fungus.

An experiment was conducted in the greenhouse to investigate the feasibility of Vicia faba grown on different fly ash concentrations (0-30%) and dual inoculation with Rhizobium and arbuscular mycorrhizal fungi (AMF). Sampling was done 45 days after sowing to analyse the plant growth parameters, photosynthetic attributes (total chlorophyll and carotenoids content), protein content, nitrogen (N) and phosphorus (P) content, defensive factors (antioxidant activity and proline content) and damage markers (lipid peroxidation, reactive oxygen species and cell viability). The results revealed that the application of fly ash (FA) alone did not result in any significant improvement in growth, biochemical and physiological parameters. However, dual inoculation showed a synergistic impact on legume growth, photosynthetic pigments, protein, proline, and cell viability. Rhizobium, AMF and 10% FA showed maximum enhancement in all attributes mentioned. 20% and 30% fly doses showed a reduction in growth, photosynthesis and antioxidants and caused oxidative stress via lipid peroxidation. The results showed that the synergistic or combined interactions between all three variables of the symbiotic relationship (Rhizobium-legume-AMF) boosted plant productivity.

Precision Genome Editing Techniques in Gene Therapy: Current State and Future Prospects.

Precision genome editing is a rapidly evolving field in gene therapy, allowing for the precise modification of genetic material. The CRISPR and Cas systems, particularly the CRISPR-- Cas9 system, have revolutionized genetic research and therapeutic development by enabling precise changes like single-nucleotide substitutions, insertions, and deletions. This technology has the potential to correct disease-causing mutations at their source, allowing for the treatment of various genetic diseases. Programmable nucleases like CRISPR-Cas9, transcription activator-like effector nucleases (TALENs), and zinc finger nucleases (ZFNs) can be used to restore normal gene function, paving the way for novel therapeutic interventions. However, challenges, such as off-target effects, unintended modifications, and ethical concerns surrounding germline editing, require careful consideration and mitigation strategies. Researchers are exploring innovative solutions, such as enhanced nucleases, refined delivery methods, and improved bioinformatics tools for predicting and minimizing off-target effects. The prospects of precision genome editing in gene therapy are promising, with continued research and innovation expected to refine existing techniques and uncover new therapeutic applications.

Chloroquine and cytosolic galectins affect endosomal escape of antisense oligonucleotides after Stabilin-mediated endocytosis.

Non-DNA-binding Stabilin-2/HARE receptors expressed on liver sinusoidal endothelial cells specifically bind to and internalize several classes of phosphorothioate antisense oligonucleotides (PS-ASOs). After Stabilin-mediated uptake, PS-ASOs are trafficked within endosomes (>97%-99%), ultimately resulting in destruction in the lysosome. The ASO entrapment in endosomes lowers therapeutic efficacy, thereby increasing the overall dose for patients. Here, we use confocal microscopy to characterize the intracellular route transverse by PS-ASOs after Stabilin receptor-mediated uptake in stable recombinant Stabilin-1 and -2 cell lines. We found that PS-ASOs as well as the Stabilin-2 receptor transverse the classic path: clathrin-coated vesicle-early endosome-late endosome-lysosome. Chloroquine exposure facilitated endosomal escape of PS-ASOs leading to target knockdown by more than 50% as compared to untreated cells, resulting in increased PS-ASO efficacy. We also characterize cytosolic galectins as novel contributor for PS-ASO escape. Galectins knockdown enhances ASO efficacy by more than 60% by modulating EEA1, Rab5C, and Rab7A mRNA expression, leading to a delay in the endosomal vesicle maturation process. Collectively, our results provide additional insight for increasing PS-ASO efficacy by enhancing endosomal escape, which can further be utilized for other nucleic acid-based modalities.

Akkermansia muciniphila and its membrane protein ameliorates intestinal inflammatory stress and promotes epithelial wound healing via CREBH and miR-143/145.

BACKGROUND: The intestinal epithelial barrier is the interface for interaction between gut microbiota and host metabolic systems. Akkermansia muciniphila (A. muciniphila) is a key player in the colonic microbiota that resides in the mucus layer, whose abundance is selectively decreased in the faecal microbiota of inflammatory bowel disease (IBD) patients. This study aims to investigate the regulatory mechanism among A. muciniphila, a transcription factor cAMP-responsive element-binding protein H (CREBH), and microRNA-143/145 (miR-143/145) in intestinal inflammatory stress, gut barrier integrity and epithelial regeneration. METHODS: A novel mouse model with increased colonization of A muciniphila in the intestine of CREBH knockout mice, an epithelial wound healing assay and several molecular biological techniques were applied in this study. Results were analysed using a homoscedastic 2-tailed t-test. RESULTS: Increased colonization of A. muciniphila in mouse gut enhanced expression of intestinal CREBH, which was associated with the mitigation of intestinal endoplasmic reticulum (ER) stress, gut barrier leakage and blood endotoxemia induced by dextran sulfate sodium (DSS). Genetic depletion of CREBH (CREBH-KO) significantly inhibited the expression of tight junction proteins that are associated with gut barrier integrity, including Claudin5 and Claudin8, but upregulated Claudin2, a tight junction protein that enhances gut permeability, resulting in intestinal hyperpermeability and inflammation. Upregulation of CREBH by A. muciniphila further coupled with miR-143/145 promoted intestinal epithelial cell (IEC) regeneration and wound repair via insulin-like growth factor (IGF) and IGFBP5 signalling. Moreover, the gene expressing an outer membrane protein of A. muciniphila, Amuc_1100, was cloned into a mammalian cell-expression vector and successfully expressed in porcine and human IECs. Expression of Amuc_1100 in IECs could recapitulate the health beneficial effect of A. muciniphila on the gut by activating CREBH, inhibiting ER stress and enhancing the expression of genes involved in gut barrier integrity and IEC's regeneration. CONCLUSIONS: This study uncovers a novel mechanism that links A. muciniphila and its membrane protein with host CREBH, IGF signalling and miRNAs in mitigating intestinal inflammatory stress-gut barrier permeability and promoting intestinal wound healing. This novel finding may lend support to the development of therapeutic approaches for IBD by manipulating the interaction between host genes, gut bacteria and its bioactive components.

Insight into the Tubulin-Targeted Anticancer Potential of Noscapine and its Structural Analogs.

Cancer is known as a notorious disease responsible for threatening millions of lives every year. Natural products which act by disrupting the microtubule assembly and dynamics have proven to be highly successful as anticancer agents but their high toxicity owing to lower selectivity has limited their usage. Recently, Noscapine (NOS), a known anti-tussive, has come out to be an effective anti-tubulin candidate with far lesser toxicity. Since its first report as an anti-mitotic agent in 1998, NOS has been extensively studied and modified by various groups of researchers to optimize its anti-tubulin activity. In this review, the recent advancements about the potential of these therapeutic candidates against various cancers have been compiled and analyzed for their inhibitory mechanism in distinct health conditions. It has been observed that the non-polar substitutions (e.g., halides, aryl groups) at specific sites (9-position and N-sites of isoquinoline ring; and modification of a methoxy group) have an enhanced effect on efficacy. The mechanistic studies of NOS and its modified analogs have shown their inhibitory action primarily through interaction with microtubules dynamics thus disrupting the cell-cycle and leading to apoptosis. This review highlights the latest research in the field by providing a rich resource for the researchers to have a hands-on analysis of NOS analogs and the inhibitory action in comparison to other microtubule disrupting anti-cancer agents. The article also documents the newer investigations in studying the potential of noscapine analogs as possible anti-microbial and antiviral agents.

Cancer Categorization Using Genetic Algorithm to Identify Biomarker Genes.

In the microarray gene expression data, there are a large number of genes that are expressed at varying levels of expression. Given that there are only a few critically significant genes, it is challenging to analyze and categorize datasets that span the whole gene space. In order to aid in the diagnosis of cancer disease and, as a consequence, the suggestion of individualized treatment, the discovery of biomarker genes is essential. Starting with a large pool of candidates, the parallelized minimal redundancy and maximum relevance ensemble (mRMRe) is used to choose the top m informative genes from a huge pool of candidates. A Genetic Algorithm (GA) is used to heuristically compute the ideal set of genes by applying the Mahalanobis Distance (MD) as a distance metric. Once the genes have been identified, they are input into the GA. It is used as a classifier to four microarray datasets using the approved approach (mRMRe-GA), with the Support Vector Machine (SVM) serving as the classification basis. Leave-One-Out-Cross-Validation (LOOCV) is a cross-validation technique for assessing the performance of a classifier. It is now being investigated if the proposed mRMRe-GA strategy can be compared to other approaches. It has been shown that the proposed mRMRe-GA approach enhances classification accuracy while employing less genetic material than previous methods. Microarray, Gene Expression Data, GA, Feature Selection, SVM, and Cancer Classification are some of the terms used in this paper.

Machine Learning Techniques for Human Age and Gender Identification Based on Teeth X-Ray Images.

The use of digital medical images is increasing with advanced computational power that has immensely contributed to developing more sophisticated machine learning techniques. Determination of age and gender of individuals was manually performed by forensic experts by their professional skills, which may take a few days to generate results. A fully automated system was developed that identifies the gender of humans and age based on digital images of teeth. Since teeth are a strong and unique part of the human body that exhibits least subject to risk in natural structure and remains unchanged for a longer duration, the process of identification of gender- and age-related information from human beings is systematically carried out by analyzing OPG (orthopantomogram) images. A total of 1142 digital X-ray images of teeth were obtained from dental colleges from the population of the middle-east part of Karnataka state in India. 80% of the digital images were considered for training purposes, and the remaining 20% of teeth images were for the testing cases. The proposed gender and age determination system finds its application widely in the forensic field to predict results quickly and accurately. The prediction system was carried out using Multiclass SVM (MSVM) classifier algorithm for age estimation and LIBSVM classifier for gender prediction, and 96% of accuracy was achieved from the system.

Stabilin receptors clear LPS and control systemic inflammation.

Lipopolysaccharides (LPSs) cause lethal endotoxemia if not rapidly cleared from blood circulation. Liver sinusoidal endothelial cells (LSEC) systemically clear LPS by unknown mechanisms. We discovered that LPS clearance through LSEC involves endocytosis and lysosomal inactivation via Stabilin-1 and 2 (Stab1 and Stab2) but does not involve TLR4. Cytokine production was inversely related to clearance/endocytosis of LPS by LSEC. When exposed to LPS, Stabilin double knockout mice (Stab DK) and Stab1 KO, but not Stab2 KO, showed significantly enhanced systemic inflammatory cytokine production and early death compared with WT mice. Stab1 KO is not significantly different from Stab DK in circulatory LPS clearance, LPS uptake and endocytosis by LSEC, and cytokine production. These data indicate that (1) Stab1 receptor primarily facilitates the proactive clearance of LPS and limits TLR4-mediated inflammation and (2) TLR4 and Stab1 are functionally opposing LPS receptors. These findings suggest that endotoxemia can be controlled by optimizing LPS clearance by Stab1.

Prominent Receptors of Liver Sinusoidal Endothelial Cells in Liver Homeostasis and Disease.

Liver sinusoidal endothelial cells (LSECs) are the most abundant non-parenchymal cells lining the sinusoidal capillaries of the hepatic system. LSECs are characterized with numerous fenestrae and lack basement membrane as well as a diaphragm. These unique morphological characteristics of LSECs makes them the most permeable endothelial cells of the mammalian vasculature and aid in regulating flow of macromolecules and small lipid-based structures between sinusoidal blood and parenchymal cells. LSECs have a very high endocytic capacity aided by scavenger receptors (SR), such as SR-A, SR-B (SR-B1 and CD-36), SR-E (Lox-1 and mannose receptors), and SR-H (Stabilins). Other high-affinity receptors for mediating endocytosis include the FcγRIIb, which assist in the antibody-mediated removal of immune complexes. Complemented with intense lysosomal activity, LSECs play a vital role in the uptake and degradation of many blood borne waste macromolecules and small (<280 nm) colloids. Currently, seven Toll-like receptors have been investigated in LSECs, which are involved in the recognition and clearance of pathogen-associated molecular pattern (PAMPs) as well as damage associated molecular pattern (DAMP). Along with other SRs, LSECs play an essential role in maintaining lipid homeostasis with the low-density lipoprotein receptor-related protein-1 (LRP-1), in juxtaposition with hepatocytes. LSECs co-express two surface lectins called L-Specific Intercellular adhesion molecule-3 Grabbing Non-integrin Receptor (L-SIGN) and liver sinusoidal endothelial cell lectin (LSECtin). LSECs also express several adhesion molecules which are involved in the recruitment of leukocytes at the site of inflammation. Here, we review these cell surface receptors as well as other components expressed by LSECs and their functions in the maintenance of liver homeostasis. We further discuss receptor expression and activity and dysregulation associated with the initiation and progression of many liver diseases, such as hepatocellular carcinoma, liver fibrosis, and cirrhosis, alcoholic and non-alcoholic fatty liver diseases and pseudocapillarization with aging.

Thermoluminescence Studies of ß and γ-Irradiated Geological Materials for Environment Monitoring.

In the present report, thermally stimulated luminescence (TSL) of quartz and limestone samples irradiated with ß and γ-rays has been investigated. Herein the formation of trap depths and calculation of kinetic parameters of ß and γ - irradiated quartz and limestone samples were studied through thermoluminescence (TL) glow curve analyses. The quartz and limestone samples were collected from various sites of Chhattisgarh (Patharia and Dalli-Rajhara mines). The collected raw samples were annealed at 400 °C. The phase formation of collected samples is confirmed by X-ray diffraction studies. The grain sizes of the samples are determined by using Debye-Scherrer formula. TL glow curves of the collected samples were recorded for various doses of ß and γ-rays. Kinetic parameters such as order of kinetics frequency factor and trap depth were calculated by employing CGCD methods. A comparative study on the TL properties of the geological materials under ß and γ-irradiation was done. The trap model analysis was executed to determine the nature of traps responsible for dominant TL peaks of ß and γ-irradiated limestone and quartz samples.

Affordability versus innovation: Is compulsory licensing the solution?

In an era which marks an exceptional phase of growth in science and technology, the acute disparities in access to healthcare still persist. So where on one hand scientific advancement in medicine aims at increasing life expectancy, on the other hand there are millions who are denied access to existing medicines. Patents on medicines also pose a significant barrier to access new drugs, especially in low and middle income countries which already suffer from poor health financing mechanisms. The patent laws were built on the assumption of incentivizing the innovators by rewarding them with the exclusive right to produce, sell or market the innovation. The basic premise for granting patents was based on the thought that it would increase investment in research and development promoting dynamic gains through newer innovations. However, evidence found to support this justification is meager. So in a situation where the drug gap still persists and we aim to achieve sustainable development goals by 2030, this paper attempts to focus on understanding how compulsory licensing has been used in selected cases to alleviate the major legal and political barriers to access medicines. The methodology comprises of cross-country comparison of patent framework and compulsory licensing cases. The sample selected for study includes both developed as well as developing countries. The aim is to evaluate the policy approaches used by selected countries to grant compulsory licenses and to identify the best practices for evidence-based policy making on international issues related to pharmaceutical patents. In each case, a driving factor has been the international extension of patent laws through trade agreements; first bilaterally (US-Canada) and subsequently internationally (1995 Uruguay round, under which low- and middle-income countries were granted a grace period until 2005 to comply).

Patent implications in the pharmaceutical industry: Are static high prices leading to dynamic innovations?

Agreement on Trade Related aspects of Intellectual Property Rights (TRIPS) was laid on the premises of rewarding monopolistic patent rights to the innovator. Stronger patent protection was advocated to promote technology transfer from developed nations to the rest of the world. To boost domestic innovative potential and to maintain trade ties, most developing countries signed the TRIPS agreement. Impact of patent laws in the pharmaceutical industry was crucial as it posed threat to access health. This paper aims to analyze the impact of pharmaceutical product patent laws incorporated under the TRIPS agreement using 65 countries panel dataset from 1995 to 2016. The data is empirically analyzed using negative binomial regression and Poisson regression. Results clearly indicate that the number of pharmaceutical patents filed in US Patent and Trademark Office (USPTO) has decreased after TRIPS compliance in both low and middle income countries. However, the decline is larger in upper middle and lower middle income countries than in low income countries. The phenomenon of low patent activity has an increasingly declining trend across low and middle income countries. Hence the claimed hypothesis that stronger patent rights would increase innovative potential does not seem to stand true, which raises serious affordability concerns of bringing patents in the pharmaceutical sector.