Biology subjects

Hagan, R.

Publications and source records attributed to Hagan, R..

6 recordsLinked to original sources

bioRxiv2026

Wavelet Transform improves time-frequency characterization of the Electroretinographic Oscillatory Potentials as compared to Short-Time Fourier Transform across experimental and clinical models

Purpose: To compare continuous wavelet transform (CWT) and short-time Fourier transform (STFT) analyses of the oscillatory potentials (OPs) of the dark-adapted full-field electroretinogram (FFERG). Methods: FFERG recordings from Royal College of Surgeons (RCS) and healthy Brown Norway (BN) rats, wild-type and Atoh7 enhancer-knockout (KO) mice, and patients with diabetic retinopathy and healthy controls were analyzed with STFT and with CWT employing the real Morlet wavelet, after 60 Hz high-pass and sharp 60 Hz notch filtering. Time- and frequency-localized OP features were quantified and the methods compared on detection of transient OP signals. Both were additionally validated using synthetic OP-like bursts of known frequency embedded in each cohort's recorded noise. Results: CWT revealed distinct time-frequency patterns and disease-specific OP alterations across all rat, mouse, and human cohorts, with improved temporal localization and frequency resolution, whereas STFT merged transient features and was more susceptible to noise. CWT-derived dominant-frequency differences reached significance in all three models; the corresponding STFT difference reached significance only in rats (separate tests; methods not directly compared). In synthetic bursts, both methods recovered injected frequencies to within 1-2.5 Hz, but only CWT resolved two bursts separated by 15 ms. Conclusions: CWT employing the real Morlet wavelet provides a robust, physiologically relevant evaluation of OPs, with improved temporal localization and reduced spectral dispersion relative to STFT in clinical and experimental settings. Translational Relevance: CWT analysis may improve the characterization of inner retinal dysfunction by preserving transient OP time-frequency structure, as illustrated in preclinical rodent models and diabetic retinopathy patients.

physiology↗

bioRxiv2025

An unclean slate, discrepancies between food input and recovered protein signal from experimental foodcrusts

Organic residues are a rich source of biomolecular information on ancient diets. In particular, foodcrusts, charred residues on ceramics, are commonly analysed for their lipid content and to a lesser extent protein in order to identify foods, culinary practices and material culture use in past populations. However, the composition of foodcrusts and the factors behind their formation are not well understood. Here we analyse proteomic data (available via ProteomeXchange with identifier PXD059930) from foodcrusts made using a series of mixtures of protein- (salmon flesh), lipid- (beef fat) and carbohydrate-rich (beetroot) foods to investigate the relationship between the biomolecular composition of the input and the recovered signal using conventional methods applied to archaeological material. Additionally, using 3D modelling we quantify the volume of foodcrust generated by different ingredient combinations The results highlight biases in the data obtained in the analyses of organic residues both in terms of identified resources reflecting the cooked foodstuffs, e.g. an overrepresentation of fish proteins, as well as with regards to the abundance of foodcrust, for example mixtures of only salmon and beef fat resulted in relatively small amounts of foodcrust, and suggest caution in interpreting the composition of residues formed from complex mixtures of foodstuffs.

biochemistry↗

bioRxiv2021

MK2 Expression Promotes Non-Small Cell Lung Cancer Cell Death and Predicts Survival

Non-small cell lung cancers demonstrate intrinsic resistance to cell death even in response to chemotherapy. Previous work suggested that defective nuclear translocation of active caspase 3 may play a role in resistance to cell death. Separately, our group has identified that mitogen activated protein kinase activated protein kinase 2 (MK2) is required for nuclear translocation of active caspase 3 in the execution of apoptosis. This study demonstrates a relatively low expression of MK2 in non-small cell lung carcinoma cell lines compared to small cell carcinoma cell lines. Further, overexpression of MK2 in non-small cell lung carcinoma cell lines results in increased caspase 3 activity and caspase 3 mediated cell death. Higher MK2 transcript levels were observed in patients with earlier-stage non-small cell lung cancer. Higher expression of MK2 is associated with better survival in patients with early stage non-small cell lung cancer across two independent clinical datasets. Using data sets spanning multiple cancer types, we observed improved survival with higher MK2 expression was unique to lung adenocarcinoma. Mechanistically, MK2 promotes nuclear translocation of caspase 3 leading to PARP1 cleavage and execution of cell death. While MK2 can directly phosphorylate caspase 3, neither phosphorylation status of caspase 3 nor the kinase activity of MK2 impacts caspase 3 activation, nuclear translocation and execution of cell death. Rather, a non-kinase function of MK2, specifically trafficking via its nuclear localization sequence, is required for caspase 3 mediated cell death. In summary this study highlights the importance of a non-enzymatic function of MK2 in the execution of apoptosis, which may be leveraged in the adjunctive treatment of NSCLC or other conditions where regulation of apoptosis is crucial.

cancer biology↗

bioRxiv2020

Tracing production instability in a clonally-derived CHO cell line using single cell transcriptomics

A variety of mechanisms including transcriptional silencing, gene copy loss and increased susceptibility to cellular stress have been associated with a sudden or gradual loss of monoclonal antibody (mAb) production in Chinese hamster ovary (CHO) cell lines. In this study, we utilised single cell RNA-seq (scRNA-seq) to study a clonally-derived CHO cell line that underwent production instability leading to a dramatic reduction of the levels of mAb produced. From the scRNA-seq data we identified sub clusters associated with variations in the mAb transgenes and observed that heavy chain gene expression was significantly lower than that of the light chain across the population. Using trajectory inference, the evolution of the cell line was reconstructed and was found to correlate with a reduction in heavy and light chain gene expression. Genes encoding for proteins involved in the response to oxidative stress and apoptosis were found to increase in expression as cells progressed along the trajectory. Future studies of CHO cell lines using this technology have the potential to dramatically enhance our understanding of the characteristics underpinning efficient manufacturing performance as well as product quality. HighlightsO_LIA clonally-derived CHO cell line in our laboratory had undergone production instability - in that the amount of intact monoclonal antibody had reduced dramatically to levels at which reliable quantitation was no longer possible. We were, however, able to detect mAb heavy and light chain protein, as well as dimerised light chain species in the cell culture media. C_LIO_LISingle cell RNA-seq was utilised to capture > 3,800 gene expression profiles from the cell line at 72hrs post seeding. C_LIO_LIAnalyses of the scRNA-seq data uncovered transcriptional heterogeneity and revealed the presence of multiple intra cell line clusters. The heavy chain transcript was detected at a significantly lower level in comparison to light chain transcripts. Light chain gene expression was not only more abundant, but also expressed more uniformly across the cell population. C_LIO_LIUsing unsupervised trajectory analysis, the emergence of heterogeneity in the cell population was traced from those cells most similar to the original isolated clone to those where transcription of the mAb heavy and light chain was undetectable. C_LIO_LISubsequent analysis of CHO cell gene expression patterns revealed a correlation between the progression of cells along the trajectory and the upregulation of genes involved in the cellular response to oxidative stress. C_LI

cell biology↗

bioRxiv2019

Sub physiological temperature induces pervasive alternative splicing in Chinese hamster ovary cells

RNA sequencing (RNASeq) has been widely used to associate alterations in Chinese hamster ovary (CHO) cell gene expression with bioprocess phenotypes, however alternative mRNA splicing, has thus far, received little attention. In this study, we utilised RNASeq for transcriptomic analysis of a monoclonal antibody producing CHOK1 cell line subjected to a temperature shift. More than 2,365 instances of differential splicing were observed 24hrs after the reduction of cell culture temperature. 1,163 of these alternative splicing events were identified in genes where no changes in abundance were detected by standard differential expression analysis. Ten examples of alternative splicing were selected for independent validation using qPCR in the monoclonal antibody producing CHOK1 line used for RNASeq and a further 2 CHOK1 cell lines. This analysis provided evidence that exon skipping and mutually exclusive splicing events occur in genes linked to the cellular response to changes in temperature and mitochondrial function. While further work is required to determine the impact of changes in mRNA sequence on cellular phenotype, this study demonstrates that alternative splicing analysis can be utilised to gain a deeper understanding of post-transcriptional regulation in CHO cells during biopharmaceutical production.

cell biology↗

bioRxiv2017

Dehydration Bouts Prompt Increased Activity And Blood Feeding By Mosquitoes

Mosquitoes are prone to dehydration and respond to this stress through multiple mechanisms, but previous studies have examined very specific responses and fail to provide an encompassing view of the role that dehydration has on mosquito biology. This study examined underlying changes in biology of the northern house mosquito, Culex pipiens, associated with short bouts of dehydration. We show that dehydration increased blood feeding propensity of mosquitoes, which was the result of both enhanced activity and a higher tendency to land on a host. Mosquitoes exposed to dehydrating conditions with access to water or rehydrated individuals experience no water loss and failed to display behavioral changes. RNA-seq and metabolome analyses following dehydration indicated that factors associated with energy metabolism are altered, specifically the breakdown of trehalose to yield glucose, which likely underlies changes in mosquito activity. Suppression of trehalose breakdown by RNA interference reduced phenotypes associated with dehydration. Comparable results were noted for two other mosquito species, suggesting this is a general response among mosquitoes. Lastly, field-based mesocosm studies using C. pipiens revealed that dehydrated mosquitoes were more likely to host feed, and disease modeling indicates dehydration bouts may increase transmission of West Nile virus. These results suggest that periods of dehydration prompt mosquitoes to utilize blood feeding as a mechanism to obtain water. This dehydration-induced increase in blood feeding is likely to intensify disease transmission during periods of low water availability.\n\nSignificanceDehydration stress has substantial impacts on the biology of terrestrial invertebrates. To date, no studies have elucidated the difference between dehydration exposure and realized water loss in relation to mosquito behavior and physiology. Our experiments show that direct dehydration stress increases mosquito activity and subsequent blood feeding, likely as a mechanism to locate and utilize a bloodmeal for rehydration. These dehydration-induced phenotypes were linked to altered carbohydrate metabolism that acts as a source of energy. This study provides important insight into the impact of mosquito-dehydration dynamics on disease transmission that is likely general among mosquitoes.

physiology↗