Platelet changes and bleeding symptoms in children, adolescents, and adults with 22q11.2 deletion syndrome.

BACKGROUND: The deletion region of 22q11.2 deletion syndrome (22q11.2DS) contains a gene encoding glycoprotein Ibß (GPIbß), which is required to express the GPIb/IX/V complex on the platelet surface. Therefore, patients with 22q11.2DS may have congenital platelet disorders. However, information is limited on platelets and bleeding symptoms. In this study, we investigated clinical information, including bleeding symptoms, platelet counts, and GPIb expression levels in children and adolescents/adults with 22q11.2DS. PROCEDURE: Thirty-two patients with 22q11.2DS were enrolled in a prospective cohort study between 2022 and 2023 at outpatient clinics within our institute. RESULTS: The median platelet counts in adolescents/adults with 22q11.2DS were significantly lower than those in children (p < .0001). A gradual decrease was found along with increasing age (p = .0006). Values of median GPIb expression on platelet surfaces (66% in children and 70% in adolescents/adults) were significantly lower than those in healthy controls (p < .0001 and p = .0002). Bleeding symptoms included surgery-related bleeding (52%), purpura (31%), and epistaxis (22%); most of them were minor. The median International Society on Thrombosis and Hemostasis bleeding assessment tool score was not significantly different between children and adolescents/adults (p = .2311). CONCLUSION: Although there was an age-related decrease in platelet count and a disease-related decrease in GPIb expression, no difference in bleeding symptoms was found between children and adolescents/adults. 22q11.2DS overall had minor bleeding symptoms in daily life, and the disease had little effect on spontaneous bleeding. However, some patients had major bleeding events; further accumulation of data on hemostasis during surgery and trauma is required.

Diagnostic efficacy of ACA, aß2-GP1, hs-CRP, and Hcy for cerebral infarction and their relationship with the disease severity.

OBJECTIVE: To evaluate the diagnostic efficacy of anti-cardiolipin antibodies (ACA), anti-ß2-glycoprotein I antibodies (aß2-GP1), high-sensitivity C-reactive protein (hs-CRP), and homocysteine (Hcy) in cerebral infarction and to explore their relationship with disease severity. METHODS: Medical records of 67 cerebral infarction patients admitted from May 2020 to January 2023 and 50 healthy individuals undergoing health checkups were retrospectively analyzed. The levels of ACA, aß2-GP1, hs-CRP, and Hcy were compared, their correlation with National Institutes of Health Stroke Scale (NIHSS) scores was assessed, and their diagnostic efficacy across different disease severities were evaluated. A joint predictive score formula, defined as -6.054712173 + aß2-GP1*1.906727231 + Hcy*0.576221974, which combines aß2-GP1 and Hcy levels, was developed to assess the likelihood of cerebral infarction in our study population. RESULTS: The levels of ACA, aß2-GP1, hs-CRP and Hcy, and joint predictive score were significantly higher in the patient group (all P < 0.001). ROC analysis yielded AUCs of 0.887 for ACA, 0.894 for aß2-GP1, 0.899 for hs-CRP, 0.880 for Hcy, and 0.954 for the joint predictive score. Delong's test showed no statistical difference in most indicators compared to the joint predictive score (P > 0.05), except aß2-GP1 (P < 0.05). Pearson's correlation analysis indicated that aß2-GP1, Hcy, and the joint predictive score were positively correlated of with NIHSS score (all P < 0.05), while ACA and hs-CRP were not (P > 0.05). Notable differences in aß2-GP1 and the joint predictive score were observed among varying severity levels (P < 0.01), with the joint predictive score showing superior diagnostic efficacy in distinguishing between mild and moderate/severe cases (P < 0.01). CONCLUSION: ACA, aß2-GP1, hs-CRP, and Hcy are effective biomarkers for diagnosing cerebral infarction, and are positively correlated with disease severity. The joint predictive score demonstrates enhanced accuracy in discerning degree of severity.

Bernard-Soulier syndrome caused by two novel heterozygous GP1BA gene mutations: a case report and literature review.

BACKGROUND: Bernard-Soulier syndrome (BSS) is a rare inherited macrothrombocytopenia, usually autosomal recessive, which is characterized by prolonged bleeding, thrombocytopenia, and abnormally large platelets. METHODS: For more than 6 years, we misdiagnosed a patient with BSS without an obvious bleeding tendency as having idiopathic thrombocytopenia purpura (ITP), prior to obtaining a genetic analysis. On admission, routine hematology showed a platelet count of 30 × 109/L and mean platelet volume (MPV) of 14.0 fL. RESULTS: Whole-exome sequencing revealed two likely pathogenic heterozygous mutations (c.95_101del and c.1012del) in GP1BA. Flow cytometry analysis of platelet membrane glycoproteins indicated that the expression of GP1b was 0.28% of the normal level. Platelet aggregation tests indicated that platelet aggregation was inhibited by ristocetin- (1.7%), ADP- (14.5%), and arachidonic acid- (5.6%) induced platelet aggregation. A literature review identified reports on 53 mutations in the GP1BA gene in 253 patients, 29 mutations in the GP1BB gene in 90 patients, and 32 mutations in the GP9 gene in 114 patients. CONCLUSION: This case report describes two novel gene mutation sites that have not been reported previously, enriching understanding of the GP1BA mutation spectrum.

Genetic Polymorphisms of GP1BA, PEAR1, and PAI-1 may be Associated with Serum sIgE and Blood Eosinophil Levels in Chinese Patients with Allergic Diseases.

BACKGROUND: It has been suggested that genetic factors may be substantially linked to allergy disorders. OBJECTIVE: This study aims to investigate the relationship between the serum specific Immunoglobulin E (sIgE), blood eosinophil, and the polymorphisms of glycoprotein Ib alpha gene (GP1BA) rs6065, platelet endothelial aggregation receptor 1 gene (PEAR1) rs12041331, and plasminogen activator inhibitor 1 gene (PAI-1) rs1799762. METHODS: From the Peking Union Medical College Hospital, this study enrolled 60 healthy participants and 283 participants with allergic diseases. TaqMan-minor groove binder (MGB) quantitative polymerase chain reaction (qPCR) was used to examine the gene polymorphisms in each group. RESULTS: The TaqMan-MGB qPCR results were completely consistent with the DNA sequencing results, according to other studies in this medical center (Kappa =1, p <0.001). The GP1BA rs6065, PEAR1 rs12041331, and PAI-1 rs1799762 polymorphisms did not show different distribution between allergy patients and healthy individuals. Concerning allergy patients, the CT (n=33) genotype of GP1BA rs6065 had higher blood eosinophil level than the CC (n=250) genotype (0.59, IQR 0.32-0.72 vs 0.31, IQR 0.15-0.61, *109/L, p =0.005). The serum sIgE of AA (n=46) genotype of PEAR1 rs12041331 was lower (median 3.7, interquartile quartiles (IQR) 0.2-16.8, kU/L) than the GA (n=136) and GG (n=101) genotypes (GA median 16.3, IQR 3.1-46.3, kU/L, p = 0.002; GG median 12.9, IQR 3.0-46.9, kU/L, p =0.003). The GA genotypes of PEAR1 rs12041331were with higher blood eosinophil levels (median 0.42, IQR 0.17-0.74 *109/L) than the AA genotype (median 0.25, IQR 0.15-0.41*109/L, p =0.012). The sIgE of the 5G5G (n=44) genotype of PAI-1 rs1799762 was lower (median 5.0, IQR 0.1-22.8, kU/L) than the 4G5G (n=144) (median 17.3, IQR 3.7-46.0, kU/L, p = 0.012). CONCLUSION: The GP1BA rs6065, PEAR1 rs12041331, and PAI-1 rs1799762 polymorphisms may be associated with the genetic susceptibility of serum sIgE or blood eosinophil in Chinese allergic disease patients.

Platelet Glycoprotein-Ib (GPIb) May Serve as a Bridge between Type 2 Diabetes Mellitus (T2DM) and Atherosclerosis, Making It a Potential Target for Antiplatelet Agents in T2DM Patients.

Type 2 diabetes mellitus (T2DM) is a persistent metabolic condition that contributes to the development of cardiovascular diseases. Numerous studies have provided evidence that individuals with T2DM are at a greater risk of developing cardiovascular diseases, typically two to four times more likely than those without T2DM, mainly due to an increased risk of atherosclerosis. The rupture of an atherosclerotic plaque leading to pathological thrombosis is commonly recognized as a significant factor in advancing cardiovascular diseases caused by TD2M, with platelets inducing the impact of plaque rupture in established atherosclerosis and predisposing to the primary expansion of atherosclerosis. Studies suggest that individuals with T2DM have platelets that display higher baseline activation and reactivity than those without the condition. The expression enhancement of several platelet receptors is known to regulate platelet activation signaling, including platelet glycoprotein-Ib (GPIb). Furthermore, the high expression of platelet GP1b has been reported to increase the risk of platelet adhesion, platelet-leucocyte interaction, and thrombo-inflammatory pathology. However, the study exploring the role of GP1b in promoting platelet activation-induced cardiovascular diseases in T2DM patients is still limited. Therefore, we summarize the important findings regarding pathophysiological continuity between T2DM, platelet GPIb, and atherosclerosis and highlight the potential therapy targeting GPIb as a novel antiplatelet agent for preventing further cardiovascular incidents in TD2M patients.

Structurally Different Yet Functionally Similar: Aptamers Specific for the Ebola Virus Soluble Glycoprotein and GP1,2 and Their Application in Electrochemical Sensing.

The Ebola virus glycoprotein (GP) gene templates several mRNAs that produce either the virion-associated transmembrane protein or one of two secreted glycoproteins. Soluble glycoprotein (sGP) is the predominant product. GP1 and sGP share an amino terminal sequence of 295 amino acids but differ in quaternary structure, with GP1 being a heterohexamer with GP2 and sGP a homodimer. Two structurally different DNA aptamers were selected against sGP that also bound GP1,2. These DNA aptamers were compared with a 2'FY-RNA aptamer for their interactions with the Ebola GP gene products. The three aptamers have almost identical binding isotherms for sGP and GP1,2 in solution and on the virion. They demonstrated high affinity and selectivity for sGP and GP1,2. Furthermore, one aptamer, used as a sensing element in an electrochemical format, detected GP1,2 on pseudotyped virions and sGP with high sensitivity in the presence of serum, including from an Ebola-virus-infected monkey. Our results suggest that the aptamers interact with sGP across the interface between the monomers, which is different from the sites on the protein bound by most antibodies. The remarkable similarity in functional features of three structurally distinct aptamers suggests that aptamers, like antibodies, have preferred binding sites on proteins.

The impact of SELP gene Thr715Pro polymorphism on sP-selectin level and association with cardiovascular disease in Saudi diabetic patients: A cross-sectional case-control study.

Background: Cardiovascular diseases (CVD) are leading cause of mortality in patients with type 2 diabetes mellitus (T2DM). Increased soluble sP-selectin and 715Thr > Pro polymorphism were studied in CVD and T2DM, but association between them hasn't been explored in Saudi Arabia. We aimed to assess sP-selectin levels in T2DM and T2DM-associated CVD patients in comparison to healthy control cohort. Also, we sought to investigate relationship between Thr715Pro polymorphism and sP-selectin levels and disease state. Methods: This is a cross-sectional case-control study. sP-selectin level (measured by Enzyme-linked immunosorbent assay) and prevalence of Thr715Pro polymorphism (assessed by Sanger sequencing) were investigated in 136 Saudi participants. The study comprised 3 groups: group1 included 41 T2DM patients; group 2 (48 T2DM patients with CVD), and group 3 (47 healthy controls). Results: sP-selectin levels were significantly higher in diabetics and diabetics + CVD groups as compared to the corresponding control. In addition, results showed that the prevalence of 715Thr > Pro polymorphism is 11.75 % in the study population amongst the three study groups (9.55 % Thr/Pro, and 2.2 % Pro/Pro). No statistical difference was found between sP-selectin levels in subject carrying the wildtype genotype of this polymorphism and these who carry the mutant gene. There could be an association between this polymorphism and T2DM, whilst the polymorphism may protect diabetic patients from having CVD. However, odds ratio is not statistically significant in both cases. Conclusion: Our study supports the previous researches' results that Thr715Pro is neither influencing the sP-selectin level nor the risk of CVD in T2DM patients.

Epitope of antiphospholipid antibodies retrieved from peptide microarray based on R39-R43 of ß2-glycoprotein I.

Background: Antiphospholipid antibody (aPL) syndrome (APS) is an autoimmune disease characterized by the presence of antiphospholipid antibodies and thromboembolic or pregnancy complications. Although cryptic epitope R39-R43 belonging to beta-2-glycoprotein 1 (ß2GP1) has been identified as the main antigenic determinant for aPLs, we have recently demonstrated that the epitope is a motif determined by the polarity, rather than by the sequence or charge of amino acids. Objective: In the present study, we wanted to identify the association of residues needed to obtain the highest aPL affinity. Methods: Based on the epitope R39-R43 and our identified motif, we generated a printed peptide microarray of 676 different peptides. These peptides have been then screened for their ability to interact with the plasmas from 11 well-characterized APS patients and confirmed by surface plasma resonance assay. Results and Conclusions: We identified a peptide that selectively bound immunoglobulin G (IgG) derived from APS patients with 100 times more affinity than ß2GP1, Domain I, or epitope R39-R43. This peptide is able to inhibit the activity of IgG derived from APS patients in vitro. We have also generated a monoclonal IgG antibody against this peptide. Using both peptide and monoclonal antibody, we have been able to develop a fully standardized indirect colorimetric immunoassay with highly sensitivity. The identification of the optimized peptide offers a new standardized and accurate tool for diagnostics of APS. Furthermore, having increased affinity for aPL, this peptide could represent a useful tool as prevention strategy for APS and an alternative to the use of anticoagulants.

Biological, clinical features and modelling of heterozygous variants of glycoprotein Ib platelet subunit alpha (GP1BA) and glycoprotein Ib platelet subunit beta (GP1BB) genes responsible for constitutional thrombocytopenia.

Constitutional thrombocytopenias are rare disorders, often difficult to discriminate from acquired thrombocytopenias. More than 80 genes have been described as being at the origin of these diseases. Among them, several variants of the glycoprotein Ib platelet subunit alpha (GP1BA) and glycoprotein Ib platelet subunit beta (GP1BB) genes, coding for the GpIb-IX-V glycoprotein complex, have been reported in the literature. The study reported here aimed at describing newly identified monoallelic anomalies affecting the GP1BA and GP1BB genes on a clinical, biological and molecular level. In a cohort of nine patients with macrothrombocytopenia, eight heterozygous variants of the GP1BA or GP1BB genes were identified. Five of them had never been described in the heterozygous state. Computer modelling disclosed structure/function relationships of these five variants.

Augmented Therapeutic Potential of EC-Synthetic Retinoids in Caco-2 Cancer Cells Using an In Vitro Approach.

Colorectal cancer therapies have produced promising clinical responses, but tumor cells rapidly develop resistance to these drugs. It has been previously shown that EC19 and EC23, two EC-synthetic retinoids, have single-agent preclinical anticancer activity in colorectal carcinoma. Here, isobologram analysis revealed that they have synergistic cytotoxicity with retinoic acid receptor (RAR) isoform-selective agonistic retinoids such as AC261066 (RARß2-selective agonist) and CD437 (RARγ-selective agonist) in Caco-2 cells. This synergism was confirmed by calculating the combination index (lower than 1) and the dose reduction index (higher than 1). Flow cytometry of combinatorial IC50 (the concentration causing 50% cell death) confirmed the cell cycle arrest at the SubG0-G1 phase with potentiated apoptotic and necrotic effects. The reported synergistic anticancer activity can be attributed to their ability to reduce the expression of ATP-binding cassette (ABC) transporters including P-glycoprotein (P-gp1), breast cancer resistance protein (BCRP) and multi-drug resistance-associated protein-1 (MRP1) and Heat Shock Protein 70 (Hsp70). This adds up to the apoptosis-promoting activity of EC19 and EC23, as shown by the increased Caspase-3/7 activities and DNA fragmentation leading to DNA double-strand breaks. This study sheds the light on the possible use of EC-synthetic retinoids in the rescue of multi-drug resistance in colorectal cancer using Caco-2 as a model and suggests new promising combinations between different synthetic retinoids. The current in vitro results pave the way for future studies on these compounds as possible cures for colorectal carcinoma.

Cancer multidrug-resistance reversal by ABCB1 inhibition: A recent update.

Chemotherapy is one of the most common treatments for cancer that uses one or more anti-cancer drugs as a part of the standardized chemotherapy regimen. Cytotoxic chemicals delay and prevent cancer cells from multiplying, invading, and metastasizing. However, the significant drawbacks of cancer chemotherapy are the lack of selectivity of the cytotoxic drugs to tumour cells and normal cells and the development of resistance by cells for the particular drug or the combination of drugs. Multidrug resistance (MDR) is the low sensitivity of specific cells against drugs associated with cancer chemotherapy. The most common mechanisms of anticancer drug resistance are: (a) drug-dependent MDR (b) target-dependent MDR, and (c) drug target-independent MDR. In all the factors, the overexpression of multidrug efflux systems contributes significantly to the increased resistance in the cancer cells. Multidrug resistance due to efflux of anticancer drugs by membrane ABC transporters includes ABCB1, ABCC1, and ABCG2. ABCB1 inhibition can restore the sensitivity of the cancerous cells toward chemotherapeutic drugs. In this review, we discussed ABCB1 inhibitors under clinical studies with their mode of action, potency and selectivity. Also, we have highlighted the contribution of repurposing drugs, biologics and nano formulation strategies to combat multidrug resistance by modulating the ABCB1 activity.

Pseudogene CLEC4GP1 modulates trophoblast cell apoptosis and invasion via IL-15 inhibition.

Preeclampsia (PE) is a pregnancy-associated complication accompanied by gestational hypertension and proteinuria, affecting 2-8% of pregnancies globally. The placental trophoblast cell invasion of decidua and myometrium during early gestation is crucial for healthy placentation. Thus, trophoblast dysfunction might contribute to PE onset. Therefore, further investigations are needed to elucidate the underlying mechanism of trophoblast cell functions. In the present study, we identified a novel pseudogene named C-Type Lectin Domain Family 4 Member G Pseudogene 1 (CLEC4GP1), which was aberrantly expressed in PE placental tissues. In vitro analyses showed that CLEC4GP1 overexpression significantly increased the cell viability and invasiveness and decreased the apoptosis rate of HTR-8/SVneo and JEG-3 cells, while CLEC4GP1 knockdown exerted opposite effects, suggesting the beneficial role of CLEC4GP1 in trophoblast cells. Next, co-expression analysis found that CLEC4GP1 was negatively correlated with Interleukin 15 (IL-15). The expression of IL-15 dramatically increased in PE placental tissues. In HTR-8/SVneo and JEG-3 cells, IL-15 exhibited detrimental effects, opposite to CLEC4GP1, and they were negatively correlated. In addition, CLEC4GP1 attenuates the mRNA stability of IL-16 by inhibiting the binding between human antigen R (HuR) protein and IL-15 RNA. Finally, the obverse effects of CLEC4GP1 and IL-15 were investigated, and results showed that IL-15 reverted CLEC4GP1 induced cellular functions. In brief, these data suggest that CLEC4GP1/IL-15 axis might modulate the occurrence and progression of PE via influencing the trophoblast cell viability, apoptosis, and invasive capability. This study provided cognizance of targeting the CLEC4GP1/IL-15 axis as a novel therapeutic approach to mitigate PE progression.

Antigens and Antibodies of the Antiphospholipid Syndrome as New Allies in the Pathogenesis of COVID-19 Coagulopathy.

High prevalence of both criteria and extra-criteria antiphospholipid antibodies (aPL) has been reported in COVID-19 patients. However, the differences in aPL prevalence decreased when an age-matched control group was included. The association of aPL with thrombotic events in COVID-19 is very heterogeneous. This could be influenced by the fact that most of the studies carried out were conducted on small populations enriched with elderly patients in which aPL was measured only at a single point and they were performed with non-standardized assays. The few studies that confirmed aPL in a second measurement showed that aPL levels hardly changed, with the exception of the lupus anticoagulant that commonly reduced. COVID-19 coagulopathy is an aPL-independent phenomenon closely associated with the onset of the disease. Thrombosis occurs later in patients with aPL presence, which is likely an additional prothrombotic factor. B2-glycoprotein deficiency (mainly aPL antigen caused both by low production and consumption) is very common during the SARS-CoV2 infection and has been associated with a greater predisposition to COVID-19 complications. This could be a new prothrombotic mechanism that may be caused by the blockage of its physiological functions, the anticoagulant state being the most important.

NXN Gene Epigenetic Changes in an Adult Neurogenesis Model of Alzheimer's Disease.

In view of the proven link between adult hippocampal neurogenesis (AHN) and learning and memory impairment, we generated a straightforward adult neurogenesis in vitro model to recapitulate DNA methylation marks in the context of Alzheimer's disease (AD). Neural progenitor cells (NPCs) were differentiated for 29 days and Aß peptide 1-42 was added. mRNA expression of Neuronal Differentiation 1 (NEUROD1), Neural Cell Adhesion Molecule 1 (NCAM1), Tubulin Beta 3 Class III (TUBB3), RNA Binding Fox-1 Homolog 3 (RBFOX3), Calbindin 1 (CALB1), and Glial Fibrillary Acidic Protein (GFAP) was determined by RT-qPCR to characterize the culture and framed within the multistep process of AHN. Hippocampal DNA methylation marks previously identified in Contactin-Associated Protein 1 (CNTNAP1), SEPT5-GP1BB Readthrough (SEPT5-GP1BB), T-Box Transcription Factor 5 (TBX5), and Nucleoredoxin (NXN) genes were profiled by bisulfite pyrosequencing or bisulfite cloning sequencing; mRNA expression was also measured. NXN outlined a peak of DNA methylation overlapping type 3 neuroblasts. Aß-treated NPCs showed transient decreases of mRNA expression for SEPT5-GP1BB and NXN on day 9 or 19 and an increase in DNA methylation on day 29 for NXN. NXN and SEPT5-GP1BB may reflect alterations detected in the brain of AD human patients, broadening our understanding of this disease.

Deep-Time Structural Evolution of Retroviral and Filoviral Surface Envelope Proteins.

The retroviral surface envelope protein subunit (SU) mediates receptor binding and triggers membrane fusion by the transmembrane (TM) subunit. SU evolves rapidly under strong selective conditions, resulting in seemingly unrelated SU structures in highly divergent retroviruses. Structural modeling of the SUs of several retroviruses and related endogenous retroviral elements with AlphaFold 2 identifies a TM-proximal SU ß-sandwich structure that has been conserved in the orthoretroviruses for at least 110 million years. The SU of orthoretroviruses diversified by the differential expansion of the ß-sandwich core to form domains involved in virus-host interactions. The ß-sandwich domain is also conserved in the SU equivalent GP1 of Ebola virus although with a significantly different orientation in the trimeric envelope protein structure relative to the ß-sandwich of human immunodeficiency virus type 1 gp120, with significant evidence for divergent rather than convergent evolution. The unified structural view of orthoretroviral SU and filoviral GP1 identifies an ancient, structurally conserved, and evolvable domain underlying the structural diversity of orthoretroviral SU and filoviral GP1. IMPORTANCE The structural relationships of SUs of retroviral groups are obscured by the high rate of sequence change of SU and the deep-time divergence of retroviral lineages. Previous data showed no structural or functional relationships between the SUs of type C gammaretroviruses and lentiviruses. A deeper understanding of structural relationships between the SUs of different retroviral lineages would allow the generalization of critical processes mediated by these proteins in host cell infection. Modeling of SUs with AlphaFold 2 reveals a conserved core domain underlying the structural diversity of orthoretroviral SUs. Definition of the conserved SU structural core allowed the identification of a homologue structure in the SU equivalent GP1 of filoviruses that most likely shares an origin, unifying the SU of orthoretroviruses and GP1 of filoviruses into a single protein family. These findings will allow an understanding of the structural basis for receptor-mediated membrane fusion mechanisms in a broad range of biomedically important retroviruses.

Immunization with GP1 but Not Core-like Particles Displaying Isolated Receptor-Binding Epitopes Elicits Virus-Neutralizing Antibodies against Junín Virus.

New World arenaviruses are rodent-transmitted viruses and include a number of pathogens that are responsible for causing severe human disease. This includes Junín virus (JUNV), which is the causative agent of Argentine hemorrhagic fever. The wild nature and mobility of the rodent reservoir host makes it difficult to control the disease, and currently passive immunization with high-titer neutralizing antibody-containing plasma from convalescent patients is the only specific therapy. However, dwindling supplies of naturally available convalescent plasma, and challenges in developing similar resources for other closely related viruses, have made the development of alternative antibody-based therapeutic approaches of critical importance. In this study, we sought to induce a neutralizing antibody response in rabbits against the receptor-binding subunit of the viral glycoprotein, GP1, and the specific peptide sequences in GP1 involved in cellular receptor contacts. While these specific receptor-interacting peptides did not efficiently induce the production of neutralizing antibodies when delivered as a particulate antigen (as part of hepatitis B virus core-like particles), we showed that recombinant JUNV GP1 purified from transfected mammalian cells induced virus-neutralizing antibodies at high titers in rabbits. Further, neutralization was observed across a range of unrelated JUNV strains, a feature that is critical for effectiveness in the field. These results underscore the potential of GP1 alone to induce a potent neutralizing antibody response and highlight the importance of epitope presentation. In addition, effective virus neutralization by rabbit antibodies supports the potential applicability of this species for the future development of immunotherapeutics (e.g., based on humanized monoclonal antibodies). Such information can be applied in the design of vaccines and immunogens for both prevention and specific therapies against this and likely also other closely related pathogenic New World arenaviruses.

A GP1BA Variant in a Czech Family with Monoallelic Bernard-Soulier Syndrome.

Bernard-Soulier syndrome (BSS) is a rare inherited disorder characterized by unusually large platelets, low platelet count, and prolonged bleeding time. BSS is usually inherited in an autosomal recessive (AR) mode of inheritance due to a deficiency of the GPIb-IX-V complex also known as the von Willebrand factor (VWF) receptor. We investigated a family with macrothrombocytopenia, a mild bleeding tendency, slightly lowered platelet aggregation tests, and suspected autosomal dominant (AD) inheritance. We have detected a heterozygous GP1BA likely pathogenic variant, causing monoallelic BSS. A germline GP1BA gene variant (NM_000173:c.98G > A:p.C33Y), segregating with the macrothrombocytopenia, was detected by whole-exome sequencing. In silico analysis of the protein structure of the novel GPIbα variant revealed a potential structural defect, which could impact proper protein folding and subsequent binding to VWF. Flow cytometry, immunoblot, and electron microscopy demonstrated further differences between p.C33Y GP1BA carriers and healthy controls. Here, we provide a detailed insight into its clinical presentation and phenotype. Moreover, the here described case first presents an mBSS patient with two previous ischemic strokes.

18F-GP1 Positron Emission Tomography and Bioprosthetic Aortic Valve Thrombus.

BACKGROUND: Bioprosthetic valve thrombosis may have implications for valve function and durability. OBJECTIVES: Using a novel glycoprotein IIb/IIIa receptor radiotracer 18F-GP1, we investigated whether positron emission tomography (PET)-computed tomography (CT) could detect thrombus formation on bioprosthetic aortic valves. METHODS: Ex vivo experiments were performed on human platelets and explanted bioprosthetic aortic valves. In a prospective cross-sectional study, patients with either bioprosthetic or normal native aortic valves underwent echocardiography, CT angiography, and 18F-GP1 PET-CT. RESULTS: Flow cytometric analysis, histology, immunohistochemistry, and autoradiography demonstrated selective binding of 18F-GP1 to activated platelet glycoprotein IIb/IIIa receptors and thrombus adherent to prosthetic valves. In total, 75 participants were recruited: 53 with bioprosthetic valves (median time from implantation 37 months [IQR: 12-80 months]) and 22 with normal native aortic valves. Three participants had obstructive valve thrombosis, and a further 3 participants had asymptomatic hypoattenuated leaflet thickening on CT angiography. All bioprosthetic valves, but none of the native aortic valves, demonstrated focal 18F-GP1 uptake on the valve leaflets: median maximum target-to-background ratio 2.81 (IQR: 2.29-3.48) vs 1.43 (IQR: 1.28-1.53) (P < 0.001). Higher 18F-GP1 uptake was independently associated with duration of valve implantation and hypoattenuated leaflet thickening. All 3 participants with obstructive valve thrombosis were anticoagulated for 3 months, leading to resolution of their symptoms, improvement in mean valve gradients, and a reduction in 18F-GP1 uptake. CONCLUSIONS: Adherence of activated platelets is a common and sustained finding on bioprosthetic aortic valves. 18F-GP1 uptake is higher in the presence of thrombus, regresses with anticoagulation, and has potential use as an adjunctive clinical tool. (18F-GP1 PET-CT to Detect Bioprosthetic Aortic Valve Thrombosis; NCT04073875).

Characteristics of purified anti-ß2GPI IgG N-glycosylation associate with thrombotic, obstetric and catastrophic antiphospholipid syndrome.

OBJECTIVE: Anti-ß-2 glycoprotein I (anti-ß2GPI) antibodies, defined as primary pathogenic antibody in antiphospholipid syndrome (APS). It has been reported that IgG Fc N-glycosylation affects IgG effector, we aim to investigate the association of Fc glycosylation profiles of purified anti-ß2GP1 IgG with clinical features of APS. METHODS: We purify anti-ß2GPI IgG and total IgG from 82 APS patients including nine catastrophic antiphospholipid syndrome (CAPS) patients, as well as total IgG from 103 healthy controls to quantitatively analyse all detectable Fc N-glycanforms of all IgG subclasses with Multiple Reaction Monitoring (MRM) method based on UPLC-ESI-QqQ mass spectrometry. RESULTS: Both purified anti-ß2GPI IgG and APS total IgG showed altered N-glycan profiles when compared with healthy control (HC) IgG. Anti-ß2GPI IgG presented with lower galactosylation, increased bisection and core fucosylation compared with APS total IgG and HC IgG. We found higher galactosylation of aß2GPI IgG2 in thrombotic APS compared with the obstetric APS, and lower galactosylation of aß2GPI IgG2 associated with late pregnancy morbidity. Moreover, low galactosylation of all anti-ß2GPI IgG subclasses, increased bisection and core fucosylation of anti-ß2GPI IgG1/2 were strongly associated with CAPS and triple positivity of antiphospholipid antibodies (aPLs). CONCLUSION: We comprehensively characterize the N-Glycans landscape of both anti-ß2GP1 and total IgG in APS. Altered N-glycan profiles of anti-ß2GPI IgG enables enabled the antibodies with proinflammatory properties. Furthermore, we associated levels of IgG Fc-glycosylation with clinical features antiphospholipid syndrome. These findings could increase our understanding of anti-ß2GPI antibody mediated mechanisms in APS and be used to develop diagnostics and new target treatments.

A novel frameshift GP1BB mutation causes autosomal dominant macrothrombocytopenia with decreased vWF receptor expression but normal platelet aggregation.

GP1bß is a component of the von Willebrand factor (vWF) receptor complex that is necessary for platelet formation and activation. A novel frameshift variant in GP1BB has been identified in a family with macrothrombocytopenia. The variant leads to a protein that is 101 amino acids longer than wild type with loss of the transmembrane domain. As there is no defect in platelet aggregation, the family are classified as heterozygous carriers of a Bernard-Soulier syndrome-related mutation. The levels of the vWF receptor on platelets are reduced to 50% of the controls, with the presence of large platelets but normal platelet aggregation demonstrating that decreased vWF receptor expression impacts proplatelet formation but not platelet function.