Trajectories of psychological distress during the COVID-19 pandemic among community-dwelling older adults in Quebec: A longitudinal study.

OBJECTIVE: The COVID-19 pandemic and its associated public health measures may increase the risk for psychological distress among vulnerable older adults. This longitudinal study aimed to identify predictors of psychological distress trajectories among community-dwelling older adults in Quebec, Canada. METHODS: The study spanned four time points across 13 months and three waves of the COVID-19 pandemic. The sample included 645 community-dwelling older adults ages 60 years and older in Quebec. Participants completed telephone-based interviews that included the Kessler 6-item Psychological Distress Scale (K6) to assess psychological distress at each time point as well as information on socioeconomic, medical, psychological and COVID-19 related factors. Group-based trajectory modelling was used to identify distinct trajectories of psychological distress across time. RESULTS: Three group-based trajectories of psychological distress were identified: the resilient (50.5%), reactive (34.9%), and elevated distress groups (14.6%). Individuals with mobility issues, insomnia symptoms, COVID-19 related acute stress, general health anxiety, increased loneliness symptoms, and those unable to use technology to see others were more likely to be in the reactive and elevated groups than the resilient group. Those with past mental health problems had uniquely increased odds of being in the reactive group compared to the resilient group. Individuals living in poverty and those who reported taking psychotropic medication had increased odds of being in the elevated distress group compared to the resilient group. CONCLUSION: These findings characterized distinct trajectories of psychological distress in older adults and identified risk factors for elevated distress levels.

COVID-19 related stress and fears of contamination: the impact of feared self-perceptions.

Specific concerns have been raised for those suffering from obsessive-compulsive disorder (OCD) during the COVID-19 pandemic, particularly those suffering from contamination fear. Investigation in non-clinical and OCD samples have reported an increase in contamination symptoms in association with the severity of the COVID-19 pandemic. In particular, stress related to COVID-19 has been found to be a major predictor of an increase in contamination symptoms. It has also been suggested that these effects may be accounted for by feared-self perceptions, that renders certain individuals more vulnerable to COVID-related stress and its effect on contamination-related symptomatology. We hypothesized that feared self-perceptions would predict COVID-19-related stress and that both feared self-perceptions and COVID-19-related stress would predict contamination symptoms while controlling for age, education and sex. To test this hypothesis, 1137 community participants completed online questionnaires. Path analysis confirmed our hypotheses highlighting the importance of feared self-perceptions during the COVID-19 pandemic in its effect on stress and ensuing symptomatology. Further, women scored higher on questionnaires, but the relationship between feared self-perceptions, COVID-19-related stress and contamination symptoms remained similar. Implications for theory and research are discussed.

Unveiling the bovine embryo transcriptome during the maternal-to-embryonic transition.

Bovine early embryos are transcriptionally inactive and subsist through the initial developmental stages by the consumption of the maternal supplies provided by the oocyte until its own genome activation. In bovine, the activation of transcription occurs during the 8- to 16-cell stages and is associated with a phase called the maternal-to-embryonic transition (MET) where maternal mRNA are replaced by embryonic ones. Although the importance of the MET is well accepted, since its inhibition blocks embryonic development, very little is known about the transcripts expressed at this crucial step in embryogenesis. In this study, we generated and characterized a cDNA library enriched in embryonic transcripts expressed at the MET in bovine. Suppression subtractive hybridization followed by microarray hybridization was used to isolate more than 300 different transcripts overexpressed in untreated late eight-cell embryos compared with those treated with the transcriptional inhibitor, alpha-amanitin. Validation by quantitative RT-PCR of 15 genes from this library revealed that they had remarkable consistency with the microarray data. The transcripts isolated in this cDNA library have an interesting composition in terms of molecular functions; the majority is involved in gene transcription, RNA processing, or protein biosynthesis, and some are potentially involved in the maintenance of pluripotency observed in embryos. This collection of genes associated with the MET is a novel and potent tool that will be helpful in the understanding of particular events such as the reprogramming of somatic cells by nuclear transfer or for the improvement of embryonic culture conditions.

Revealing the bovine embryo transcript profiles during early in vivo embryonic development.

Gene expression profiling is proving to be a powerful approach for the identification of molecular mechanisms underlying complex cellular functions such as the dynamic early embryonic development. The objective of this study was to perform a transcript abundance profiling analysis of bovine early embryonic development in vivo using a bovine developmental array. The molecular description of the first week of life at the mRNA level is particularly challenging when considering the important fluctuations in RNA content that occur between developmental stages. Accounting for the different intrinsic RNA content between developmental stages was achieved by restricting the reaction time during the global amplification steps and by using spiked controls and reference samples. Analysis based on intensity values revealed that most of the transcripts on the array were present at some point during in vivo bovine early embryonic development, while the varying number of genes detected in each developmental stage confirmed the dynamic profile of gene expression occurring during embryonic development. Pair-wise comparison of gene expression showed a marked difference between oocytes and blastocysts profiles, and principal component analysis revealed that the majority of the transcripts could be regrouped into three main clusters representing distinct RNA abundance profiles. Overall, these data provide a detailed temporal profile of the abundance of mRNAs revealing the richness of signaling processes in early mammalian development. Results presented here provide better knowledge of bovine in vivo embryonic development and contribute to the progression of our current knowledge regarding the first week of life in mammals.

The dynamics of gene products fluctuation during bovine pre-hatching development.

Early embryonic development, spanning fertilization to blastocyst hatching, is a very dynamic developmental window that is characterized, especially in large mammals, by a period of transcriptional incompetence that ends during the maternal to embryonic transition (MET). Prior to the MET, the first cell cycles are supported by stored RNA and proteins pools accumulated during oogenesis. Therefore, RNA and protein content are different between developmental stages. It is also known that the stability of the stored mRNA and the mechanisms for translation recruitment are partly controlled by the length of the poly(A) tail. To date, little is known about RNA and protein content fluctuations during the pre-hatching period. In this report we present measurements of total RNA, mRNA, poly(A) bearing mRNA and protein contents, as well as estimations of the proportions of both mRNA fractions to total RNA contents within these developmental stages. We found that while the ontogenic profiles of the different transcript contents were expected, their amounts were considerably lower than the reported values. Additionally, low 28S rRNA abundance and a tendency for diminishing protein content prior to the MET, suggest a limited potential for ribosomal turnover and translation. We consider the overall fluctuations in RNA and protein contents to be reference points that are essential for downstream interpretation of gene expression data across stages whether it be through candidates or high throughput approaches.

The DNA damage-inducible C. elegans tankyrase is a nuclear protein closely linked to chromosomes.

Tankyrases are protein members of the poly(ADP-ribose) polymerase family bearing several ankyrin domain and a WGR domain. They have functional role in telomere maintenance, are found at centrosome, and are associated with vesicular secretion. This diversity in localization and function makes it difficult to identify a unified role for tankyrases. We have shown that the C. elegans orthologue PME-5 is among the most transcriptionally up-regulated genes following ionizing radiations, linking a tankyrase with DNA damage response. Our analysis showed that the up-regulation of PME-5 is translated at the protein level, suggesting an effective role in DNA damage response or DNA repair. In order to gain more information on the potential role of PME-5 in DNA damage response, we analyzed its sub-cellular localization. Using immunostaining as well as gfp reporter assay, we have shown a nuclear localization for PME-5. Moreover, we showed that PME-5 is a ubiquitous nuclear protein expressed throughout the development of the worm and is closely linked to chromatin and condensed chromosomes. Taken together, our data suggest that C. elegans can be used to study the nuclear roles of tankyrase.

Identification of differentially expressed markers in human follicular cells associated with competent oocytes.

BACKGROUND: The development of an accurate method for selection of high-quality embryos is essential to achieve high pregnancy rates with single embryo transfer in human IVF. The developmental competence of the oocyte is acquired during follicle maturation and strong communication also exists between the follicular cells (FCs) and the oocytes; thus oocyte developmental competence may be determined by markers expressed in the surrounding FCs. METHODS: From consenting patients (n = 40), FCs were recovered on a per follicle basis by individual follicle puncture. Hybridization analyses using a custom-made complementary DNA microarray containing granulosa/cumulus expressed sequence tags (ESTs) from subtracted libraries and an Affymetrix GeneChip were performed to identify specific genes expressed in follicles leading to a pregnancy. The selected candidate genes were validated by quantitative-PCR (Q-PCR). RESULTS: Subtractive libraries prepared from pooled samples representing pregnant versus non-pregnant patients produced 1694 ESTs. Hybridization data analysis discriminated 115 genes associated with competent follicles. Selected candidates were confirmed by Q-PCR: 3-beta-hydroxysteroid dehydrogenase 1 (P = 0.0078), Ferredoxin 1 (P = 0.0203), Serine (or cysteine) proteinase inhibitor clade E member 2 (P = 0.0499), Cytochrome P450 aromatase (P = 0.0359) and Cell division cycle 42 (P = 0.0396). CONCLUSIONS: Microarray technologies are useful to mine the transcriptome of FCs expressed in follicles associated with competent oocytes and could be used to improve embryo selection with the objective of successful single embryo transfer.

The C. elegans gene pme-5: molecular cloning and role in the DNA-damage response of a tankyrase orthologue.

Tankyrases are recently identified proteins characterized by ankyrin repeats and a poly(ADP-ribose) polymerase (PARP) signature motif. In vertebrates, tankyrases mediate protein-protein interactions via the ankyrin domain. Many partners have been identified that could function in telomere maintenance, signal transduction in vesicular transport, and cell death. To further our knowledge of tankyrases and to study their function in development, we sought and found a tankyrase-related gene in Caenorhabditis elegans that we named pme-5 (poly(ADP-ribose) metabolism enzyme-5). The protein encoded includes a large ankyrin domain and a catalytic PARP domain containing the well-conserved PARP signature sequence and the regulatory region. Unlike other tankyrases, PME-5 lacks a sterile-alpha module (SAM), but has a coiled coil domain which may mediate oligomerization. We also found that pme-5 mRNA is alternatively spliced at the fifth exon, producing a long (PME-5L) and a short (PME-5S) transcript. Both isoforms are constitutively expressed during the life cycle of C. elegans. We also show DNA damage increases expression of pme-5, a response that requires the DNA damage checkpoint gene hus-1. Moreover, DNA damage-induced germ cell apoptosis was slightly increased in pme-5(RNAi) hermaphrodites. Altogether, these data indicate that pme-5 is part of a DNA damage response pathway which leads to apoptosis in C. elegans.

Potential and limitations of bovine-specific arrays for the analysis of mRNA levels in early development: preliminary analysis using a bovine embryonic array.

New insights into the early development of large mammals are becoming available through the measurement of differential mRNA levels in oocytes and preimplantation embryos. These advances in knowledge are rapidly picking up in pace, mainly owing to the advantages brought by new molecular biology approaches being developed. The possibility of amplifying the starting material and therefore making measurements in single embryo units is now feasible. With these tools, the evaluation of variations in gene expression patterns during the preimplantation period or the impact of culture on mRNA levels is now possible. However, it is important to keep in mind that these methods still have limitations associated with sample preparation or the use of the appropriate controls. Even proper methods of analysis are very important to achieve the full benefit of the application of these tools. The present paper describes some of the potential, as well as limitations, of mRNA level analysis in early embryos, especially for microarray analysis. We have generated a bovine cDNA array (>2000 clones) that contains expressed sequence tags (ESTs) collected from various preimplantation development stages. Using this chip, we have initiated the characterisation of global mRNA level patterns of several key developmental stages from the immature oocyte to the blastocyst stage. As expected, the hybridisation results indicate very different expression profiles involving hundreds of genes when comparing oocyte and blastocyst samples to a reference mRNA sample made from a pool of ESTs from pooled somatic tissues. Although this array is still in its preliminary stage and the EST bank has not been processed to contain only unigenes, it is already a very useful tool for discovering candidate genes that may play important roles during early embryonic life.

Impact of DNA sequence and oligonucleotide length on a polythiophene-based fluorescent DNA biosensor.

DNA hybridization is a universal and specific mechanism for the recognition of biological targets. Some cationic polythiophene transducers sensitive to DNA structure have been previously utilized to detect such biomolecules. Further characterization of these systems indicates that both DNA sequence composition and length modulate the biosensor performance. It appears that different repeated sequence patterns cause different conformational changes of the polythiophene, from a more relaxed form to an extremely rigid one. A length difference between the DNA oligonucleotide probe and target has a detrimental effect on the fluorescent signal, but it can be attenuated by changing the sequence composition of the protruding target sequence. This demonstrates that the nature of DNA can be critical for hybridization-based detection systems.

Combining suppressive subtractive hybridization and cDNA microarrays to identify dietary phosphorus-responsive genes of the rainbow trout (Oncorhynchus mykiss) kidney.

Phosphorus (P)-responsive genes and how they regulate renal adaptation to phosphorous-deficient diets in animals, including fish, are not well understood. RNA abundance profiling using cDNA microarrays is an efficient approach to study nutrient-gene interactions and identify these dietary P-responsive genes. To test the hypothesis that dietary P-responsive genes are differentially expressed in fish fed varying P levels, rainbow trout were fed a practical high-P diet (R20: 0.96% P) or a low-P diet (R0: 0.38% P) for 7 weeks. The differentially-expressed genes between dietary groups were identified and compared from the kidney by combining suppressive subtractive hybridization (SSH) with cDNA microarray analysis. A number of genes were confirmed by real-time PCR, and correlated with plasma and bone P concentrations. Approximately 54 genes were identified as potential dietary P-responsive after 7 weeks on a diet deficient in P according to cDNA microarray analysis. Of 18 selected genes, 13 genes were confirmed to be P-responsive at 7 weeks by real-time PCR analysis, including: iNOS, cytochrome b, cytochrome c oxidase subunit II , alpha-globin I, beta-globin, ATP synthase, hyperosmotic protein 21, COL1A3, Nkef, NDPK, glucose phosphate isomerase 1, Na+/H+ exchange protein and GDP dissociation inhibitor 2. Many of these dietary P-responsive genes responded in a moderate way (R0/R20 ratio: <2-3 or >0.5) and in a transient manner to dietary P limitation. In summary, renal adaptation to dietary P deficiency in trout involves changes in the expression of several genes, suggesting a profile of metabolic stress, since many of these differentially-expressed candidates are associated with the cellular adaptative responses.

The influence of follicle size, FSH-enriched maturation medium, and early cleavage on bovine oocyte maternal mRNA levels.

Transcription is arrested in the bovine oocyte within the first few hours of in vitro maturation, thus the stored maternal mRNAs accumulated in the oocyte are essential to sustain development until the Maternal-Zygotic Transition. In vivo matured oocytes have superior blastocyst formation rates than in vitro matured oocytes, suggesting that the mRNA content of these oocytes is of higher quality. To determine which transcripts may be associated with developmental competence, a Suppressive Subtractive Hybridization was performed between oocytes collected by ovariectomy at 6 hr post-LH surge and oocytes from slaughterhouse collected after 6 hr of maturation, resulting in a library enriched in these functionally important mRNAs. The clones were spotted onto a cDNA microarray and transcripts potentially associated with developmental competence were hybridized onto these slides. Hybridizations were performed with transcripts up-regulated in oocytes cultured for 6 hr in the presence or absence of rFSH in vitro, and secondly with transcripts up regulated in early-cleaving embryos versus those at the one-cell stage at 36 hr postfertilization. From these hybridizations, 13 candidates were selected. Their functional association with embryonic competence was validated by measuring their relative transcript levels by quantitative real-time PCR in eight different conditions: oocytes cultured with or without rFSH, early--versus late-cleaving embryos, and oocytes from different follicle sizes (1-3, 3-5, 5-8, and >8 mm of diameter). The gene candidates CCNB2, PTTG1, H2A, CKS1, PSMB2, SKIIP, CDC5L, RGS16, and PRDX1 showed a significant quantitative association with competence compared to BMP15, GDF9, CCNB1, and STK6.

ChemMatrix, a poly(ethylene glycol)-based support for the solid-phase synthesis of complex peptides.

CM (ChemMatrix) resin is a new, totally poly(ethylene glycol) (PEG)-based resin, made exclusively from primary ether bonds and, therefore, highly chemically stable. It exhibits good loading and is user-friendly because of its free-flowing form upon drying. It performs excellently for the preparation of hydrophobic, highly structured, and poly-Arg peptides, as compared to polystyrene (PS) resins. In the most striking example, stepwise solid-phase assembly of the highly complex beta-amyloid (1-42) peptide resulted in a crude material of 91% purity. In contrast, literature procedures using PS or PEG-PS-based resins for this peptide required convergent approaches, additional time-consuming steps, or both. In addition to the difficulties of its synthesis, characterization of the beta-amyloid (1-42) peptide as a monomer is also a challenge, and methods for characterization by HPLC and MALDI-TOF have also been developed.

Identification of novel and known oocyte-specific genes using complementary DNA subtraction and microarray analysis in three different species.

The main objective of the present study was to identify novel oocyte-specific genes in three different species: bovine, mouse, and Xenopus laevis. To achieve this goal, two powerful technologies were combined: a polymerase chain reaction (PCR)-based cDNA subtraction, and cDNA microarrays. Three subtractive libraries consisting of 3456 clones were established and enriched for oocyte-specific transcripts. Sequencing analysis of the positive insert-containing clones resulted in the following classification: 53% of the clones corresponded to known cDNAs, 26% were classified as uncharacterized cDNAs, and a final 9% were classified as novel sequences. All these clones were used for cDNA microarray preparation. Results from these microarray analyses revealed that in addition to already known oocyte-specific genes, such as GDF9, BMP15, and ZP, known genes with unknown function in the oocyte were identified, such as a MLF1-interacting protein (MLF1IP), B-cell translocation gene 4 (BTG4), and phosphotyrosine-binding protein (xPTB). Furthermore, 15 novel oocyte-specific genes were validated by reverse transcription-PCR to confirm their preferential expression in the oocyte compared to somatic tissues. The results obtained in the present study confirmed that microarray analysis is a robust technique to identify true positives from the suppressive subtractive hybridization experiment. Furthermore, obtaining oocyte-specific genes from three species simultaneously allowed us to look at important genes that are conserved across species. Further characterization of these novel oocyte-specific genes will lead to a better understanding of the molecular mechanisms related to the unique functions found in the oocyte.

A versatile approach to affinitychromic polythiophenes.

The preparation of both postfunctionalizable and chromic poly[3-(N-succinimido-p-phenylcarboxylate(tetraethoxy)oxy)-4-methylthiophene] is reported. The N-hydroxysuccinimide ester side group can easily react with different amine-bearing molecules in the solid state to yield a library of new polythiophene derivatives. The resulting polymers can be dissolved in various solvents, and interactions between the side chains (ligands) and different analytes (targets) can be detected from modifications of both the side-chain and the backbone conformations resulting in important color changes (i.e., affinitychromism). This colorimetric polymeric transducer could therefore lead to highly valuable, versatile, and inexpensive tools for highthroughput screening and drug discovery.