Platelet Reactivity and Inflammatory Phenotype Induced by Full-Length Spike SARS-CoV-2 Protein and Its RBD Domain

A state of immunothrombosis has been reported in COVID-19. Platelets actively participate in this process. However, little is known about the ability of SARS-CoV-2 virus proteins to induce platelet activity. Platelet-rich plasma (PRP) was incubated with spike full-length protein and the RBD domain in independent assays. We evaluated platelet activation through the expression of P-selectin and activation of glicoprotein IIbIIIa (GP IIbIIIa), determined by flow cytometry and the ability of the proteins to induce platelet aggregation. We determined concentrations of immunothrombotic biomarkers in PRP supernatant treated with the proteins. We determined that the spike full-length proteins and the RBD domain induced an increase in P-selectin expression and GP IIbIIIa activation (p < 0.0001). We observed that the proteins did not induce platelet aggregation, but favored a pro-aggregating state that, in response to minimal doses of collagen, could re-establish the process (p < 0.0001). On the other hand, the viral proteins stimulated the release of interleukin 6, interleukin 8, P-selectin and the soluble fraction of CD40 ligand (sCD40L), molecules that favor an inflammatory state p < 0.05. These results indicate that the spike full-length protein and its RBD domain can induce platelet activation favoring an inflammatory phenotype that might contribute to the development of an immunothrombotic state.     

Mechanism of [CO2] Enrichment Alleviated Drought Stress in the Roots of Cucumber Seedlings Revealed via Proteomic and Biochemical Analysis

Cucumber is one of the most widely cultivated greenhouse vegetables, and its quality and yield are threatened by drought stress. Studies have shown that carbon dioxide concentration ([CO₂]) enrichment can alleviate drought stress in cucumber seedlings; however the mechanism of this [CO₂] enrichment effect on root drought stress is not clear. In this study, the effects of different drought stresses (simulated with 0, 5% and 10% PEG 6000, i.e., no, moderate, and severe drought stress) and [CO₂] (400 μmol·mol⁻¹ and 800 ± 40 μmol·mol⁻¹) on the cucumber seedling root proteome were analyzed using the tandem mass tag (TMT) quantitative proteomics method. The results showed that after [CO₂] enrichment, 346 differentially accumulating proteins (DAPs) were found only under moderate drought stress, 27 DAPs only under severe drought stress, and 34 DAPs under both moderate and severe drought stress. [CO₂] enrichment promoted energy metabolism, amino acid metabolism, and secondary metabolism, induced the expression of proteins related to root cell wall and cytoskeleton metabolism, effectively maintained the balance of protein processing and degradation, and enhanced the cell wall regulation ability. However, the extent to which [CO₂] enrichment alleviated drought stress in cucumber seedling roots was limited under severe drought stress, which may be due to excessive damage to the seedlings.     

Rice β-Glucosidase 4 (Os1βGlu4) Regulates the Hull Pigmentation via Accumulation of Salicylic Acid

Salicylic acid (SA) is a stress hormone synthesized in phenylalanine ammonia-lyase (PAL) and the branching acid pathway. SA has two interconvertible forms in plants: SAG (SA O-β-glucoside) and SA (free form). The molecular mechanism of conversion of SA to SAG had been reported previously. However, which genes regulate SAG to SA remained unknown. Here, we report a cytoplasmic β-glucosidase (β-Glu) which participates in the SA pathway and is involved in the brown hull pigmentation in rice grain. In the current study, an EMS-generated mutant brown hull 1 (bh1) displayed decreased contents of SA in hulls, a lower photosynthesis rate, and high-temperature sensitivity compared to the wild type (WT). A plaque-like phenotype (brown pigmentation) was present on the hulls of bh1, which causes a significant decrease in the seed setting rate. Genetic analysis revealed a mutation in LOC_Os01g67220, which encodes a cytoplasmic Os1βGlu4. The knock-out lines displayed the phenotype of brown pigmentation on hulls and decreased seed setting rate comparable with bh1. Overexpression and complementation lines of Os1βGlu4 restored the phenotype of hulls and normal seed setting rate comparable with WT. Subcellular localization revealed that the protein of Os1βGlu4 was localized in the cytoplasm. In contrast to WT, bh1 could not hydrolyze SAG into SA in vivo. Together, our results revealed the novel role of Os1βGlu4 in the accumulation of flavonoids in hulls by regulating the level of free SA in the cellular pool.     

Therapeutic Effects of Saponins for the Prevention and Treatment of Cancer by Ameliorating Inflammation and Angiogenesis and Inducing Antioxidant and Apoptotic Effects in Human Cells

Saponins are natural compounds found in plants and have a diverse range of applications. However, the therapeutic potential of saponins in regulating cytotoxicity, angiogenesis, and inflammation in mammalian cells is yet to be explored. Here, we investigated the therapeutic effects of saponins from green tea by exploring the cytotoxic effects of saponins by inducing apoptosis in the human cancer cell lines hepatocellular carcinoma (HEPG2) and colorectal adenocarcinoma (HT29). The anti-angiogenesis effect of saponins was also investigated in human umbilical vein endothelial cells (HUVEC). We explored the ability of saponins to attenuate inflammation in a dose-dependent manner in normal human cells. It was found that saponins exhibit cytotoxic effects in cancer cells and not in normal cells at the same concentration. Cytotoxicity was measured by inducing apoptosis by enhancing caspase-3 (cas-3) activation and B-cell lymphoma-2 (Bcl-2)-associated X protein (BAX) gene expression and suppressing the antiapoptotic protein, Bcl-2. The inhibition of HUVEC proliferation was due to the suppression of the phosphoinositide 3-kinase (PI3K), protein kinase B (AKT), vascular endothelial growth factor receptor-2 (VEGFR-2), and nuclear factor kappa B (NF-κB). We also observed the antioxidant potential of green tea-derived saponins against free radicals in reactive oxygen species (ROS)-induced cells. Here we observed that the saponins exhibited free radical scavenging activities and activated nuclear factorerythroid 2-related factor 2 (NRF-2) leading to the upregulation of antioxidant-related genes in human embryonic kidney 293 (HEK293) cells. Furthermore, we demonstrated that the anti-inflammatory effects were due to the suppression of pro-inflammatory cytokines interleukin (IL)-1β, IL-6, tumor necrosis factor-alpha (TNF-α), and inducible nitric oxide synthase (iNOS) in HEK293 cells. The significance of the work is we are the first to report on the anti-cancer effects of saponins based on the anti-inflammatory, antioxidant, anti-angiogenesis, and apoptosis induction properties. In conclusion, green tea-derived saponins could be effective therapeutics for the treatment of cancer.     

The Emerging Role of Venetoclax-Based Treatments in Acute Lymphoblastic Leukemia

Venetoclax, a B-cell lymphoma (BCL-2) inhibitor, in combination with hypomethylating agents has become the new standard of care in elderly and unfit patients with acute myeloid leukemia, with significantly improved overall survival and quality of life. Studies of venetoclax combined with high-dose chemotherapy are emerging with evidence of higher rates of molecular remission. Recently, a growing number of publications bring forth the use of venetoclax in patients with acute lymphoblastic leukemia (ALL). In the current review, we present the biological rationale of BCL-2 inhibition in ALL, how the interplay of BH3 proteins modulate the response and the current clinical experience with various combinations.     

Targeting PSEN1 by lnc-CYP3A43-2/miR-29b-2-5p to Reduce β Amyloid Plaque Formation and Improve Cognition Function

Presenilin-1 (PSEN1) is a crucial subunit within the γ-secretase complex and regulates β-amyloid (Aβ) production. Accumulated evidence indicates that n-butylidenephthalide (BP) acts effectively to reduce Aβ levels in neuronal cells that are derived from trisomy 21 (Ts21) induced pluripotent stem cells (iPSCs). However, the mechanism underlying this effect remains unclear. This article aims to investigate the possible mechanisms through which BP ameliorates the development of Alzheimer’s disease (AD) and verify the effectiveness of BP through animal experiments. Results from RNA microarray analysis showed that BP treatment in Ts21 iPSC-derived neuronal cells reduced long noncoding RNA (lncRNA) CYP3A43-2 levels and increased microRNA (miR)-29b-2-5p levels. Bioinformatics tool prediction analysis, biotin-labeled miR-29b-2-5p pull-down assay, and dual-luciferase reporter assay confirmed a direct negative regulatory effect for miRNA29b-2-5p on lnc-RNA-CYP3A43-2 and PSEN1. Moreover, BP administration improved short-term memory and significantly reduced Aβ accumulation in the hippocampus and cortex of 3xTg-AD mice but failed in miR-29b-2-5p mutant mice generated by CRISP/Cas9 technology. In addition, analysis of brain samples from patients with AD showed a decrease in microRNA-29b-2-5p expression in the frontal cortex region. Our results provide evidence that the LncCYP3A43-2/miR29-2-5p/PSEN1 network might be involved in the molecular mechanisms underlying BP-induced Aβ reduction.     

Human Milk Oligosaccharide 2′-Fucosyllactose Modulates Local Viral Immune Defense by Supporting the Regulatory Functions of Intestinal Epithelial and Immune Cells

Human milk contains bioactive components that provide protection against viral infections in early life. In particular, intestinal epithelial cells (IEC) have key regulatory roles in the prevention of enteric viral infections. Here we established an in vitro model to study the modulation of host responses against enteric viruses mimicked by poly I:C (pIC). The effects of 2′-fucosyllactose (2′FL), abundantly present in human milk, were studied on IEC and/or innate immune cells, and the subsequent functional response of the adaptive immune cells. IEC were pre-incubated with 2′FL and stimulated with naked or Lyovec™-complexed pIC (LV-pIC). Additionally, monocyte-derived dendritic cells (moDC) alone or in co-culture with IEC were stimulated with LV-pIC. Then, conditioned-moDC were co-cultured with naïve CD4⁺ T helper (Th)-cells. IEC stimulation with naked or LV-pIC promoted pro-inflammatory IL-8, CCL20, GROα and CXCL10 cytokine secretion. However, only exposure to LV-pIC additionally induced IFNβ, IFNλ1 and CCL5 secretion. Pre-incubation with 2′FL further increased pIC induced CCL20 secretion and LV-pIC induced CXCL10 secretion. LV-pIC-exposed IEC/moDC and moDC cultures showed increased secretion of IL-8, GROα, IFNλ1 and CXCL10, and in the presence of 2′FL galectin-4 and -9 were increased. The LV-pIC-exposed moDC showed a more pronounced secretion of CCL20, CXCL10 and CCL5. The moDC from IEC/moDC cultures did not drive T-cell development in moDC/T-cell cultures, while moDC directly exposed to LV-pIC secreted Th1 driving IL-12p70 and promoted IFNγ secretion by Th-cells. Hereby, a novel intestinal model was established to study mucosal host-defense upon a viral trigger. IEC may support intestinal homeostasis, regulating local viral defense which may be modulated by 2′FL. These results provide insights regarding the protective capacity of human milk components in early life.     

Peroxisome Proliferator-Activated Receptor-α (PPARα) Expression in a Clinical Population of Pakistani Patients with Type 2 Diabetes and Dyslipidemia

Poor glycemic control and dyslipidemia are hallmarks of type 2 diabetes mellitus (T2DM), which predispose to cardiovascular diseases. Peroxisome proliferator-activated receptor-α (PPARα) has been associated with atherosclerosis, but its role in T2DM is less clear. Previously, we studied PPARα expression levels in diabetics with and without dyslipidemia (DD). In this study we described the association with fasting blood glucose, HbA1c levels and lipid levels of the study population. Patient demography and biochemical data were collected from hospitals in Islamabad, Pakistan, and RT-PCR data of PPARα expression were retrieved from our previous study from the same cohort. We performed t-tests and regression analysis to evaluate the relationships between PPARα expression and demographic and clinical variables. As expected, body mass index and HbA1c were elevated in T2DM and DD patients compared to controls. Blood lipids (total cholesterol, triglycerides, LDL and HDL) were significantly higher in the DD group compared to the other two groups. In the T2DM and DD groups, the PPARα expression was not associated with any of the physical and biochemical parameters measured in this study. Expression of the PPARα gene was independent of blood lipids and glycemic control in this study. Further research is necessary to better understand the biological parameters of PPARα expression.