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Science (New York, N.Y.) Mar 2024The extent to which prophage proteins interact with eukaryotic macromolecules is largely unknown. In this work, we show that cytoplasmic incompatibility factor A (CifA)...
The extent to which prophage proteins interact with eukaryotic macromolecules is largely unknown. In this work, we show that cytoplasmic incompatibility factor A (CifA) and B (CifB) proteins, encoded by prophage WO of the endosymbiont alter long noncoding RNA (lncRNA) and DNA during sperm development to establish a paternal-effect embryonic lethality known as cytoplasmic incompatibility (CI). CifA is a ribonuclease (RNase) that depletes a spermatocyte lncRNA important for the histone-to-protamine transition of spermiogenesis. Both CifA and CifB are deoxyribonucleases (DNases) that elevate DNA damage in late spermiogenesis. lncRNA knockdown enhances CI, and mutagenesis links lncRNA depletion and subsequent sperm chromatin integrity changes to embryonic DNA damage and CI. Hence, prophage proteins interact with eukaryotic macromolecules during gametogenesis to create a symbiosis that is fundamental to insect evolution and vector control.
Topics: Animals; Male; Cytoplasm; DNA; Prophages; RNA, Long Noncoding; Spermatozoa; Wolbachia; Paternal Inheritance; Viral Proteins; Drosophila melanogaster; Bacterial Proteins; Deoxyribonucleases
PubMed: 38452081
DOI: 10.1126/science.adk9469 -
British Journal of Haematology May 2024Prolonged coagulation times, such as activated partial thromboplastin time (APTT) and thrombin time (TT), are common in patients infected with severe fever with... (Observational Study)
Observational Study
Prolonged coagulation times, such as activated partial thromboplastin time (APTT) and thrombin time (TT), are common in patients infected with severe fever with thrombocytopenia syndrome virus (SFTSV) and have been confirmed to be related to patient's poor outcome by previous studies. To find out the reason for prolonged coagulation time in patients with SFTSV infection, and whether it predicts haemorrhagic risk or not. Seventy-eight consecutive patients with confirmed SFTSV infection were enrolled in this prospective, single-centre, observational study. Several global and specific coagulation parameters of these patients on admission were detected, and the haemorrhagic events during hospitalization and their outcomes were recorded. Most of the enrolled patients had prolonged APTT (82.1%) and TT (80.8%), normal prothrombin time (83.3%) and intrinsic coagulation factors above haemostatic levels (97.4%). The heparin-like effect was confirmed by a protamine neutralization test and anti-Xa activity detection in most patients. Interestingly, the APTT and TT results were significantly positively correlated with the levels of endothelial markers and viral load, respectively. The APTT was independently associated with the haemorrhage of patients. The prolonged APTT and TT of SFTS patients may mainly be attributed to endogenous heparinoids and are associated with increased haemorrhagic risk.
Topics: Humans; Male; Female; Middle Aged; Aged; Partial Thromboplastin Time; Hemorrhage; Prospective Studies; Severe Fever with Thrombocytopenia Syndrome; Heparin; Adult; Thrombin Time; Phlebovirus; Blood Coagulation; Risk Factors; Aged, 80 and over
PubMed: 38438264
DOI: 10.1111/bjh.19364 -
Journal of Controlled Release :... Apr 2024Therapeutic proteins often require needle-based injections, which compromise medication adherence especially for those with chronic diseases. Sublingual administration...
Therapeutic proteins often require needle-based injections, which compromise medication adherence especially for those with chronic diseases. Sublingual administration provides a simple and non-invasive alternative. Herein, two novel peptides (lipid-conjugated protamine and a protamine dimer) were synthesized to enable sublingual delivery of proteins through simple physical mixing with the payloads. It was found that the novel peptides promoted intracellular delivery of proteins via increased pore formation on the cell surface. Results from in vitro models of cell spheroids and human sublingual tissue substitute indicated that the novel peptides enhanced protein penetration through multiple cell layers compared to protamine. The novel peptides were mixed with insulin or semaglutide and sublingually delivered to mice for blood glucose (BG) control. The effects of these sublingual formulations were comparable to the subcutaneous preparations and superior to protamine. In addition to peptide drugs, the novel peptides were shown to enable sublingual absorption of larger proteins with molecular weights from 22 to 150 kDa in mice, including human recombinant growth hormone (rhGH), bovine serum albumin (BSA) and Immunoglobulin G (IgG). The novel peptides given sublingually did not induce any measurable toxicities in mice.
Topics: Animals; Mice; Humans; Administration, Sublingual; Peptides; Immunoglobulin G; Protamines
PubMed: 38423473
DOI: 10.1016/j.jconrel.2024.02.042 -
JTCVS Open Feb 2024Professional standards recommend stopping cardiotomy suction at the termination of cardiopulmonary bypass before protamine administration based on perceived safety...
OBJECTIVE
Professional standards recommend stopping cardiotomy suction at the termination of cardiopulmonary bypass before protamine administration based on perceived safety concerns. This study evaluated a multidisciplinary collaborative quality-improvement intervention promoting this agreed-upon cardiotomy suction practice during coronary artery bypass grafting (CABG).
METHODS
A statewide intervention (eg, unblinded surgeon and perfusionist feedback, evidence-based lectures, evaluating barriers to change) involved 32 centers participating in the PERForm (ie, Perfusion Measures and Outcomes) Registry to standardize cardiotomy suction practices at cardiopulmonary bypass termination during CABG. Four non-Michigan registry participating centers were not exposed to collaborative learning. Cardiotomy suction practice was defined as the absence of or stopping cardiotomy suction before protamine administration. The practice changes attributed to the intervention, including Michigan and non-Michigan comparisons, were evaluated with the change of time effect modeled using splines. Multivariable regression was used to evaluate the intervention's associated impact (eg, mortality, reoperation, transfusion).
RESULTS
Among 10,394 patients undergoing CABG at Michigan centers, 80.7% achieved agreed-upon cardiotomy suction practices. The Michigan centers had nonsignificant changes in agreed-upon cardiotomy suction practices during the preintervention period ( = .24), with significant increased monthly change in practice thereafter, absent adjusted morbidity and mortality increases. The Michigan centers achieved a significantly greater adjusted monthly improvement in agreed-upon practices relative to non-Michigan centers within 7 months after the intervention (adjusted odds ratio for change of trends: 2.53, < .001).
CONCLUSIONS
This initiative demonstrates the effectiveness of multidisciplinary collaborative quality improvement in advancing agreed-upon cardiotomy suction practices without negatively impacting clinical outcomes.
PubMed: 38420528
DOI: 10.1016/j.xjon.2023.11.005 -
Biochemistry and Cell Biology =... Jun 2024Insects are the largest group of animals when it comes to the number and diversity of species. Yet, with the exception of , no information is currently available on the...
Insects are the largest group of animals when it comes to the number and diversity of species. Yet, with the exception of , no information is currently available on the primary structure of their sperm nuclear basic proteins (SNBPs). This paper represents the first attempt in this regard and provides information about six species of Neoptera: , and . The SNBPs of these species were characterized by acetic acid urea gel electrophoresis (AU-PAGE) and high-performance liquid chromatography fractionated. Protein sequencing was obtained using a combination of mass spectrometry sequencing, Edman N-terminal degradation sequencing and genome mining. While the SNBPs of several of these species exhibit a canonical arginine-rich protamine nature, a few of them exhibit a protamine-like composition. They appear to be the products of extensive cleavage processing from a precursor protein which are sometimes further processed by other post-translational modifications that are likely involved in the chromatin transitions observed during spermiogenesis in these organisms.
Topics: Animals; Male; Amino Acid Sequence; Protamines; Nuclear Proteins; Insect Proteins; Insecta; Molecular Sequence Data; Spermatozoa
PubMed: 38408323
DOI: 10.1139/bcb-2023-0363 -
Journal of Medicinal Chemistry Mar 2024Unfractionated heparin (UFH) and low-molecular-weight heparins (LMWHs) are widely applied for surgical procedures and extracorporeal therapies, which, however, suffer...
Unfractionated heparin (UFH) and low-molecular-weight heparins (LMWHs) are widely applied for surgical procedures and extracorporeal therapies, which, however, suffer bleeding risk. Protamine, the only clinically approved antidote, can completely neutralize UFH, but only partially neutralizes LMWHs, and also has a number of safety drawbacks. Here, we show that caltrop-like multicationic small molecules can completely neutralize both UFH and LMWHs. In vitro and ex vivo assays with plasma and whole blood and in vivo assays with mice and rats support that the lead compound is not only superior to protamine by displaying higher neutralization activity and broader therapeutic windows but also biocompatible. The effective neutralization dose and the maximum tolerated dose of the lead compound are determined to be 0.4 and 25 mg/kg in mice, respectively, suggesting good promise for further preclinical studies.
Topics: Rats; Mice; Animals; Heparin; Heparin, Low-Molecular-Weight; Antidotes; Protamines; Biological Assay; Anticoagulants
PubMed: 38407934
DOI: 10.1021/acs.jmedchem.3c02224 -
International Journal of Molecular... Feb 2024Chromatin status is critical for sperm fertility and reflects spermatogenic success. We tested a multivariate approach for studying pig sperm chromatin structure to...
Chromatin status is critical for sperm fertility and reflects spermatogenic success. We tested a multivariate approach for studying pig sperm chromatin structure to capture its complexity with a set of quick and simple techniques, going beyond the usual assessment of DNA damage. Sperm doses from 36 boars (3 ejaculates/boar) were stored at 17 °C and analyzed on days 0 and 11. Analyses were: CASA (motility) and flow cytometry to assess sperm functionality and chromatin structure by SCSA (%DFI, DNA fragmentation; %HDS, chromatin maturity), monobromobimane (mBBr, tiol status/disulfide bridges between protamines), chromomycin A3 (CMA3, protamination), and 8-hydroxy-2'-deoxyguanosine (8-oxo-dG, DNA oxidative damage). Data were analyzed using linear models for the effects of boar and storage, correlations, and multivariate analysis as hierarchical clustering and principal component analysis (PCA). Storage reduced sperm quality parameters, mainly motility, with no critical oxidative stress increases, while chromatin status worsened slightly (%DFI and 8-oxo-dG increased while mBBr MFI-median fluorescence intensity-and disulfide bridge levels decreased). Boar significantly affected most chromatin variables except CMA3; storage also affected most variables except %HDS. At day 0, sperm chromatin variables clustered closely, except for CMA3, and %HDS and 8-oxo-dG correlated with many variables (notably, mBBr). After storage, the relation between %HDS and 8-oxo-dG remained, but correlations among other variables disappeared, and mBBr variables clustered separately. The PCA suggested a considerable influence of mBBr on sample variance, especially regarding storage, with SCSA and 8-oxo-dG affecting between-sample variability. Overall, CMA3 was the least informative, in contrast with results in other species. The combination of DNA fragmentation, DNA oxidation, chromatin compaction, and tiol status seems a good candidate for obtaining a complete picture of pig sperm nucleus status. It raises many questions for future molecular studies and deserves further research to establish its usefulness as a fertility predictor in multivariate models. The usefulness of CMA3 should be clarified.
Topics: Swine; Male; Animals; Chromatin; Flow Cytometry; 8-Hydroxy-2'-Deoxyguanosine; Biofilms; Semen; Bioreactors; Spermatozoa; DNA; DNA Fragmentation; Disulfides; Bridged Bicyclo Compounds
PubMed: 38396632
DOI: 10.3390/ijms25041953 -
International Journal of Reproductive... Dec 2023[This corrects the article DOI: 10.18502/ijrm.v13i8.7507.].
Corrigendum to "Protective effects of extract on protamine gene expression, testicular function, and testicular histology in doxorubicin-treated adult rats: An experimental study" [Int J Reprod BioMed 2020; 18: 667-682].
[This corrects the article DOI: 10.18502/ijrm.v13i8.7507.].
PubMed: 38370490
DOI: 10.18502/ijrm.v21i12.15044 -
F&S Science May 2024To determine if early spermatocytes can be enriched from a human testis biopsy using fluorescence-activated cell sorting (FACS).
OBJECTIVE
To determine if early spermatocytes can be enriched from a human testis biopsy using fluorescence-activated cell sorting (FACS).
DESIGN
Potential surface markers for early spermatocytes were identified using bioinformatics analysis of single-cell RNA-sequenced human testis tissue. Testicular sperm extraction samples from three participants with normal spermatogenesis were digested into single-cell suspensions and cryopreserved. Two to four million cells were obtained from each and sorted by FACS as separate biologic replicates using antibodies for the identified surface markers. A portion from each biopsy remained unsorted to serve as controls. The sorted cells were then characterized for enrichment of early spermatocytes.
SETTING
A laboratory study.
PATIENTS
Three men with a diagnosis of obstructive azoospermia (age range, 30-40 years).
INTERVENTION
None.
MAIN OUTCOME MEASURES
Sorted cells were characterized for RNA expression of markers encompassing the stages of spermatogenesis. Sorting markers were validated by their reactivity on human testis formalin-fixed paraffin-embedded tissue.
RESULTS
Serine protease 50 (TSP50) and SWI5-dependent homologous recombination repair protein 1 were identified as potential surface proteins specific for early spermatocytes. After FACS sorting, the TSP50-sorted populations accounted for 1.6%-8.9% of total populations and exhibited the greatest average-fold increases in RNA expression for the premeiotic marker stimulated by retinoic acid (STRA8), by 23-fold. Immunohistochemistry showed the staining pattern for TSP50 to be strong in premeiotic undifferentiated embryonic cell transcription factor 1/doublesex and Mab-3 related transcription factor 1/STRA8 spermatogonia as well as SYCP3/protamine 2 spermatocytes.
CONCLUSION
This work shows that TSP50 can be used to enrich early STRA8-expressing spermatocytes from human testicular biopsies, providing a means for targeted single-cell RNA sequencing analysis and in vitro functional interrogation of germ cells during the onset of meiosis. This could enable investigation into details of the regulatory pathways underlying this critical stage of spermatogenesis, previously difficult to enrich from whole tissue samples.
Topics: Humans; Male; Spermatocytes; Adult; Flow Cytometry; Biopsy; Spermatogenesis; Testis; Azoospermia; Cell Separation; Single-Cell Analysis
PubMed: 38369016
DOI: 10.1016/j.xfss.2024.02.002 -
Microvascular Research May 2024Coronary artery bypass grafting (CABG) is considered the choice treatment for patients suffering from coronary artery disease (CAD). In the inflammatory milieu of...
BACKGROUND
Coronary artery bypass grafting (CABG) is considered the choice treatment for patients suffering from coronary artery disease (CAD). In the inflammatory milieu of cardiopulmonary bypass (CPB), systemic inflammatory response syndrome (SIRS) can induce a platelet pro-inflammatory state which could exacerbate post-CABG inflammatory status while affecting hemostatic function in patients. Therefore, focusing on platelets, the study presented here attempted to evaluate the pro-inflammatory and immunomodulatory profile of platelets as well as pro-aggregatory status during CABG.
METHODS
Platelets from patients undergoing CABG were subjected to flowcytometry analysis to evaluate P-selectin and CD40L expressions and PAC-1 binding in five intervals of 24 h before surgery, immediately, 2 h, 24 h, and one week after surgery. Moreover, intra-platelet TGF-β1 was also examined with western blotting.
RESULTS
Data showed increases of P-selectin and CD40L expressions in patients, with the meaningful loss of platelet contents of TGF-β1 after CABG (p < 0.001), where the changes tended to recover by day 7 of surgery while remaining above baseline (p < 0.001). Meanwhile, no significant change in PAC-1 binding capacity was shown.
CONCLUSION
The study presented here suggests that although the release of pro-inflammatory substances from platelets during CABG supports the post-operative inflammatory state, platelets are not pro-aggregatory enough to enhance thrombotic events after surgery. Whilst these observations could be due to successful medical interventions to optimize hemostasis during and after surgery, post-CABG reversal of anticoagulant by protamine is considered as another factor that may also have contributed to preventing pro-aggregatory but not pro-inflammatory and immunomodulatory functions of platelets.
Topics: Humans; Transforming Growth Factor beta1; P-Selectin; CD40 Ligand; Coronary Artery Bypass; Phenotype; Blood Platelets
PubMed: 38360131
DOI: 10.1016/j.mvr.2024.104669