Biology subjects

Ali, N.

Publications and source records attributed to Ali, N..

31 records · Page 2Linked to original sources

bioRxiv2023

Altered co-expression patterns of synovial fluid proteins related to the immune system and extracellular matrix organization in late stage OA, compared to non-OA controls

ObjectiveSynovial fluid contains proteins that may have been released from surrounding tissues, our aim was to gain new insights into the proteomic profiles of human synovial fluid in knees with and without osteoarthritis (OA). MethodsWe used synovial fluid from 11 patients with end-stage medial compartment knee OA, aspirated during total knee replacement, and from 13 deceased donors who had no prior history of knee OA (healthy controls). These samples were analyzed using high-multiplex immunoassays Olink(R). The differential expression of proteins between the groups was analyzed using a linear mixed effects model. The linear associations between pairs of protein expressions were estimated with a linear regression model. ResultsWe found that almost half of the detected proteins were differentially expressed between the OA and non-OA controls. The proteins that were most elevated in the OA group compared to controls were tartrate-resistant acid phosphatase type 5 (fold change 10.6, 95% CI [6.6-17.0]), coagulation factor XI (4.3 [2.6-6.8]) and urokinase-type plasminogen activator (4.3 [2.3-6.8]). The proteins with lower levels in OA compared to controls were fatty acid-binding protein, adipocyte (0.03 [0.02-0.05]), myocilin (0.05 [0.03-0.08]) and carbonic anhydrase 3 (0.14 [0.09-0.23]). The protein-protein co-expression analysis suggests an overall lower number of protein pairs that show co-expression in OA. ConclusionThere is a substantial change in protein abundance in synovial fluid in end-stage knee OA, suggesting that global joint homeostasis is severely deranged. Our findings suggest altered co-expression between the immune response and extracellular matrix organization in end-stage knee OA, in comparison to non-OA controls.

molecular biology↗

bioRxiv2022

Immunomodulatory effects of Simvastatin in Toll-like receptor-2 stimulated Rheumatoid arthritis synoviocytes

Fibroblast-like synoviocytes (FLS) are major effector cells in Rheumatoid arthritis (RA). Toll-like receptor-2 (TLR2) activation of FLS plays a key role in RA pathogenesis. Recently, statins have been shown to modulate RA. We investigated the immunomodulatory effects of simvastatin (SMV) in vitro, in FLS cell lines from control and RA subjects, in the presence of TLR2 activation. Normal and RA FLS cultures (1x105 cells/well) were treated with SMV then activated with TLR2 ligand (Pam3CSK4 100g/ml) for 24-hours. SMV caused dose-dependent cytotoxicity in both cell lines as assessed by Alamar-blue assay and increased ROS production measured by flow cytometry. Gene expression of anti-oxidants including GSR, TXNRD1, PRDX1, SOD2 and MT3 was upregulated. BNIP3 expression was increased 2200-fold in RA-FLS after SMV/TLR2 treatment pointing to apoptosis as a cause of observed cytotoxicity. Apoptosis increased in both cell lines with SMV to 25%. Akt decreased dose-dependently in RA FLS assessed by western blots. IP-1, IP-2 and IP3 decreased with SMV/TLR2 treatment indicating cell survival while IL-6 increased dose-dependently. Thus, SMV induces apoptosis in FLS by upregulating BNIP3 and inhibiting Akt. FLS react to survive by increasing IL-6, decreasing IP-1 and IP-2 and upregulating anti-oxidant genes. SMV has immunomodulatory properties and may be used in RA as adjunctive therapy in combination with other more potent disease-modifying medications.

immunology↗

bioRxiv2022

Genome-scale CRISPR screening reveals that C3aR signaling is critical for rapid capture of fungi by macrophages

The fungal pathogen Histoplasma capsulatum (Hc) invades, replicates within, and destroys macrophages. To interrogate the molecular mechanisms underlying this interaction, we conducted a host-directed CRISPR-Cas9 screen and identified 361 genes that modify macrophage susceptibility to Hc infection, greatly expanding our understanding of host gene networks targeted by Hc. We identified pathways that have not been previously implicated in Hc interaction with macrophages, including the ragulator complex (involved in nutrient stress sensing), glycosylation enzymes, protein degradation machinery, mitochondrial respiration genes, solute transporters, and the ER membrane complex (EMC). The highest scoring protective hits included the complement C3a receptor (C3aR), a G-protein coupled receptor (GPCR) that recognizes the complement fragment C3a. Although it is known that the complement system reacts with the fungal surface, leading to opsonization and release of small peptide fragments such as C3a, a role for C3aR in macrophage susceptibility to fungi has not been elucidated. We demonstrated that whereas C3aR is dispensable for macrophage phagocytosis of bacteria and latex beads, it is critical for optimal macrophage capture of pathogenic fungi, including Hc, the ubiquitous fungal pathogen Candida albicans, and the causative agent of Valley Fever Coccidioides posadasii. We showed that C3aR localizes to the early phagosome during H. capsulatum infection where it coordinates the formation of actin-rich membrane protrusions that promote Hc capture. We also showed that the EMC promotes surface expression of C3aR, likely explaining its identification in our screen. Taken together, our results provide new insight into host processes that affect Hc-macrophage interactions and uncover a novel and specific role for C3aR in macrophage recognition of fungi.

microbiology↗

bioRxiv2021

Identification of marker-trait associated SNPs for fruit weight and plant height in a wild and domesticated jujube fruit tree (Ziziphus spp.) collection using genotyping-by-sequencing

Ziziphus are economically and nutritionally important fruiting plants that were domesticated in China around 7000 years ago. We identified genetic diversity in 141 accessions of four, different species collected in Pakistan and in China, including wild species of Ziziphus mauritiana, Z. nummularia and Z. oxyphllya, and both wild and domesticated Z. jujuba Mill. Population structure, phylogenetic analysis and principal coordinates analysis based on 10,889 high-quality SNPs derived from genotyping-by-sequencing indicated that the accessions clustered into two major groups. The wild Pakistani Z. jujuba and Z. nummularia exhibited higher genetic diversity and polymorphic information content (0.37 and 0.23 respectively) than other species. We further conducted a genome-wide association study and discovered six highly significant marker-trait associations for fruit weight and plant height in this population. Our study provides important information for future breeding of Ziziphus species.

plant biology↗

bioRxiv2021

Mineral crystal thickness in calcified cartilage and subchondral bone in healthy and osteoarthritic knees

Osteoarthritis (OA) is the most common joint disease globally. In OA, articular cartilage degradation is often accompanied with sclerosis of the subchondral bone. However, the association between OA and tissue mineralization at the nanostructural level is currently not understood. Especially, it is technically challenging to identify calcified cartilage, where relevant but poorly understood pathological processes like tidemark multiplication and advancement occur. Here, we used state-of-the-art micro-focus small-angle X-ray scattering with high 5{micro}m spatial resolution to determine mineral crystal thickness in human subchondral bone and calcified cartilage. Specimens with a wide spectrum of OA severities were acquired from the medial and lateral compartments of medial compartment knee OA patients (n=15) and cadaver knees (n=10). For the first time, we identified a well-defined layer of calcified cartilage associated with pathological tidemark multiplication, containing 0.32nm thicker crystals compared to the rest of calcified cartilage. In addition, we found 0.2nm thicker mineral crystals in both tissues of the lateral compartment in OA compared with healthy knees, indicating a loading-related disease process since the lateral compartment is typically less loaded in medial compartment knee OA. Furthermore, the crystal thickness of the subchondral bone was lower with increasing histopathological OA severity. In summary, we report novel changes in mineral crystal thickness during OA. Our data suggest that unloading in the knee is associated with the growth of mineral crystals, which is especially evident in the calcified cartilage. In the subchondral bone, mineral crystals become thinner with increasing OA severity, which indicates new bone formation with sclerosis. One Sentence SummaryMineral crystal thickness increases with osteoarthritis in the lateral condyle that is typically unloaded.

biophysics↗

bioRxiv2021

Role of distinct fibroblast lineages and immune cells in dermal repair following UV radiation induced tissue damage

Solar ultraviolet radiation (UVR) is a major source of skin damage, resulting in inflammation, premature ageing and cancer. While several UVR-induced changes, including extracellular matrix reorganisation and epidermal DNA damage, have been documented, the role of different fibroblast lineages and their communication with immune cells has not been explored. We show that acute and chronic UVR exposure led to selective loss of fibroblasts from the upper dermis in human and mouse skin. Lineage tracing and in vivo live imaging revealed that repair following acute UVR is predominantly mediated by papillary fibroblast proliferation and migration. In contrast, chronic UVR exposure led to a permanent loss of papillary fibroblasts, with expansion of fibroblast membrane protrusions partially compensating for the reduction in cell number. Although UVR strongly activated Wnt-signalling in skin, stimulation of fibroblast proliferation by epidermal {beta}-catenin stabilisation did not support papillary dermis repair. Acute UVR triggered an infiltrate of neutrophils and T cell subpopulations and increased pro-inflammatory prostaglandin signalling in skin. Depletion of CD4 and CD8 positive cells resulted in increased papillary fibroblast depletion, which correlated with an increase in DNA damage and reduction in fibroblast proliferation. Conversely, topical COX-2 inhibition prevented fibroblast depletion and neutrophil infiltration after UVR. We conclude that loss of papillary fibroblasts is primarily induced by a deregulated inflammatory response, with infiltrating T cells supporting fibroblast survival upon UVR-induced environmental stress. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=91 SRC="FIGDIR/small/447606v1_ufig1.gif" ALT="Figure 1"> View larger version (26K): org.highwire.dtl.DTLVardef@d246bborg.highwire.dtl.DTLVardef@121b5faorg.highwire.dtl.DTLVardef@324114org.highwire.dtl.DTLVardef@14e9eb7_HPS_FORMAT_FIGEXP M_FIG C_FIG

cell biology↗

bioRxiv2021

Altered Sub-Genomic RNA Expression in SARS-CoV-2 B.1.1.7 Infections

SARS-CoV-2 lineage B.1.1.7 viruses are more transmissible, may lead to greater clinical severity, and result in modest reductions in antibody neutralization. subgenomic RNA (sgRNA) is produced by discontinuous transcription of the SARS-CoV-2 genome and is a crucial step in the SARS-CoV-2 life cycle. Applying our tool (periscope) to ARTIC Network Oxford Nanopore genomic sequencing data from 4400 SARS-CoV-2 positive clinical samples, we show that normalised sgRNA expression profiles are significantly increased in B.1.1.7 infections (n=879). This increase is seen over the previous dominant circulating lineage in the UK, B.1.177 (n=943), which is independent of genomic reads, E gene cycle threshold and days since symptom onset at sampling. A noncanonical sgRNA which could represent ORF9b is found in 98.4% of B.1.1.7 SARS-CoV-2 infections compared with only 13.8% of other lineages, with a 16-fold increase in median expression. We hypothesise that this is a direct consequence of a triple nucleotide mutation in nucleocapsid (28280:GAT>CAT, D3L) creating a transcription regulatory-like sequence complementary to a region 3 of the genomic leader. These findings provide a unique insight into the biology of B.1.1.7 and support monitoring of sgRNA profiles in sequence data to evaluate emerging potential variants of concern. One Sentence SummaryThe recently emerged and more transmissible SARS-CoV-2 lineage B.1.1.7 shows greater subgenomic RNA expression in clinical infections and enhanced expression of a noncanonical subgenomic RNA near ORF9b.

genomics↗

bioRxiv2021

ASSOCIATION STUDY OF SIX CANDIDATE GENES WITH MAJOR DEPRESSIVE DISORDER IN THE NORTH-WESTERN POPULATION OF PAKISTAN

People around the world are currently affected by Major Depressive Disorder (MDD). Despite its many aspects, symptoms, manifestations and impacts, efforts have been made to identify the root causes of the disorder. In particular, genetic studies have concentrated on identifying candidate genes for MDD and exploring associations between these genes and some specific group of individuals. The aim of this research was to find out the association between single nucleotide polymorphisms in 6 candidate genes linked to the neurobiology of major depressive disorder in the North-Western population of Pakistan. We performed a case-control analysis, with 400 MDD and 232 controls. A trained psychiatrist or clinical psychologists evaluated the patients. Six polymorphisms were genotyped and tested for allele and genotype association with MDD. There were no statistical variations between MDD patients and healthy controls for genotypic and allelic distribution of all the polymorphisms observed. Thus, our analysis does not support the major role of these polymorphisms in contributing to MDD susceptibility, although it does not preclude minor impact. The statistically significant correlation between six polymorphisms and major depressive disorder in the studied population was not observed. There are inconsistencies in investigations around the world. Future research, including GWAS and association analysis on larger scale should be addressed for further validation and replication of the present findings.

genetics↗

bioRxiv2020

ProteoMill: Efficient network-based functional analysis portal for proteomics data

Functional analysis has become a common approach to incorporate biological knowledge into the analysis of omics data, and to explore molecular events that govern a disease state. It is though only one step in a wider analytical pipeline that typically requires use of multiple individual analysis software. There is currently a need for a well-integrated omics analysis tool that performs all the steps. The ProteoMill portal is developed as an R Shiny application and integrates all necessary steps from data-upload, converting identifiers, to quality control, differential expression and network-based functional analysis into a single fast, interactive easy to use workflow. Further, it maintains annotation data sources up to date, overcoming a common problem with use of outdated information, and seamlessly integrates multiple R-packages for an improved user-experience. The functionality provided in this software can benefit researchers by facilitating the exploratory analysis of proteomics data. ProteoMill is available for free at https://proteomill.com.

bioinformatics↗

bioRxiv2020

Proteomics profiling of human synovial fluid suggests global increased protein interplay in early-osteoarthritis (OA) and lost in late-stage OA

The underlying molecular mechanisms in osteoarthritis (OA) development are largely unknown. This study explores the proteome and the pairwise interplay of proteins on a global level in synovial fluid from patients with late-stage knee OA (arthroplasty), early knee OA (arthroscopy due to degenerative meniscal tear) and from deceased controls without knee OA. Synovial fluid samples were analyzed using state-of-the-art mass spectrometry with data-independent acquisition. The differential expression of the proteins detected was clustered and evaluated with data mining strategies and a multilevel model. Group-specific slopes of associations were estimated between expressions of each pair of identified proteins to assess the co-expression (i.e. interplay) between the proteins in each group. More proteins were increased in early-OA vs controls than late-stage OA vs controls. For most of these proteins, the fold changes between late-stage OA vs controls and early stage OA vs controls were remarkably similar suggesting potential involvement in the OA process. Further, for the first time this study illustrated distinct patterns in protein co-expression suggesting that the global interplay between the protein machinery is increased in early-OA and lost in late-stage OA. Further efforts should probably focus on earlier stages of the disease than previously considered.

molecular biology↗

bioRxiv2020

Two pectate lyases from Caldicellulosiruptor bescii with the same CALG domain had distinct properties on plant biomass degradation

Pectin deconstruction is the initial step in breaking the recalcitrance of plant biomass by using selected microorganisms that carry pectinolytic enzymes. Pectate lyases that cleave -1,4-galacturonosidic linkage of pectin are widely used in industries, such as paper making and fruit softening. However, reports on pectate lyases with high thermostability are few. Two pectate lyases (CbPL3 and CbPL9) from a thermophilic bacterium Caldicellulosiruptor bescii were investigated. Although these two enzymes belonged to different families of polysaccharide lyase, both were Ca2+-dependent. Similar biochemical properties were shown under optimized conditions 80 {degrees}C-85 {degrees}C and pH 8-9. However, the degradation products on pectin and polygalacturonic acids (pGA) were different, revealing the distinct mode of action. A concanavalin A-like lectin/glucanase (CALG) domain, located in the N-terminus of two CbPLs, shares 100% amino acid identity. CALG-truncated mutant of CbPL9 showed lower activities than the wild-type, whereas the CbPL3 with CALG knock-out portion was reported with enhanced activities, thereby revealing the different roles of CALG in two CbPLs. I-TASSER predicted that the CALG in two CbPLs is structurally close to the family 66 carbohydrate binding module (CBM66). Furthermore, substrate-binding assay indicated that the catalytic domains in two CbPLs had strong affinities on pectate-related substrates, but CALG showed weak interaction with a number of lignocellulosic carbohydrates, except sodium carboxymethyl cellulose and sodium alginate. Finally, scanning electron microscope analysis and total reducing sugar assay showed that the two enzymes could improve the saccharification of switchgrass. The two CbPLs are impressive sources for degradation of plant biomass. ImportanceThermophilic proteins could be implemented in diverse industrial applications. We sought to characterize two pectate lyases, CbPL3 and CbPL9, from a thermophilic bacterium Caldicellulosiruptor bescii. The two enzymes had high optimum temperature, low optimum pH, and good thermostability at evaluated temperature. A family-66 carbohydrate binding module (CBM66) was identified in two CbPLs with sharing 100% amino acid identity. Deletion of CBM66 obviously decreased the activity of CbPL9, but increase the activity and thermostability of CbPL3, suggesting the different roles of CBM66 in two enzymes. Moreover, the degradation products by two CbPLs were different. These results revealed these enzymes could represent a potential pectate lyase for applications in paper and textile industries.

biochemistry↗

bioRxiv2019

Trisk95 as Novel Skin Mirror for Normal and Diabetic Systemic Glucose Level

Coping with diabetes requires frequent and even today mostly invasive blood glucose-based monitoring. Partly due to this invasive nature and the associated reduced skin wound healing and increased risk of infection, non-invasive glucose monitoring technologies would represent considerable progress. Edited keratinocytes may enable such a function.\n\nTo address this hypothesis, we conducted a proteomic screen in the skin by making use of the experimental in vivo mouse model of type I diabetes alongside controls. We identified Trisk 95 as the only protein whose expression is induced in response to high blood glucose. A luciferase reporter assay demonstrated that induction of Trisk 95 expression occurs not only at the protein level but also transcriptionally. This induction was associated with a marked elevation in the Fluo-4 signal, suggesting a role for intracellular calcium changes in the signalling cascade. Strikingly, these changes lead concurrently to fragmentation of the mitochondria. As judged from the knockout findings, both the calcium flux and the mitochondrial phenotype were dependent on Trisk 95 function, since the phenotypes in question were abolished.\n\nThe data demonstrate that the skin represents an organ that reacts robustly and thus mirrors changes in systemic blood glucose levels. The findings are also consistent with a channelling model of Trisk 95 that serves as an insulin-independent but glucose-responsive biomarker taking part in releasing calcium from the cellular stores in the skin. The skin cells may thus provide a novel mean for glucose monitoring when analysing changes in labelled Trisk 95 and calcium. By that, this study is the first proof of the concept of our registered patent (No. PCT FI2016/050917), which proposes the use of cells as biosensors for developing personalized health-monitoring devices.

cell biology↗

bioRxiv2018

Bright split red fluorescent proteins with enhanced complementation efficiency for the tagging of endogenous proteins and visualization of synapses

Self-associating split fluorescent proteins (FPs) have been widely used for labeling proteins, scaffolding protein assembly and detecting cell-cell contacts. Newly developed self-associating split FPs, however, have suffered from suboptimal fluorescence signal. Here, by investigating the complementation process, we have demonstrated two approaches to improve split FPs: assistance through SpyTag/SpyCatcher interaction and directed evolution. The latter has yielded two split sfCherry3 variants with substantially enhanced overall brightness, facilitating the tagging of endogenous proteins by gene editing. Based on sfCherry3, we have further developed a new red-colored trans-synaptic marker called Neuroligin-1 sfCherry3 Linker Across Synaptic Partners (NLG-1 CLASP) for multiplexed visualization of neuronal synapses in living animals, demonstrating its broad applications.

cell biology↗