SpotLight Proteomics Identifies Variable Sequences of Blood Antibodies Specific Against Deamidated Human Serum Albumin.

Spontaneous deamidation of asparaginyl residues in proteins, if not repaired or cleared, can set in motion a cascade that leads to deteriorated health. Previously, we have discovered that deamidated human serum albumin (HSA) is elevated in the blood of patients with Alzheimer's disease and other neurodegenerative diseases, while the level of endogenous antibodies against deamidated HSA is significantly diminished, creating an imbalance between the risk factor and the defense against it. Endogenous antibodies against deamidated proteins are still unexplored. In the current study, we employed the SpotLight proteomics approach to identify novel amino acid sequences in antibodies specific to deamidated HSA. The results provide new insights into the clearance mechanism of deamidated proteins, a possible avenue for prevention of neurodegeneration.

Heart rate deceleration and acceleration capacities associated with circadian rhythm of blood pressure in essential hypertension.

BACKGROUND: This study aimed to investigate the potential association between the circadian rhythm of blood pressure and deceleration capacity (DC)/acceleration capacity (AC) in patients with essential hypertension. METHODS: This study included 318 patients with essential hypertension, whether or not they were being treated with anti-hypertensive drugs, who underwent 24-hour ambulatory blood pressure monitoring (ABPM). Patients were categorized into three groups based on the percentage of nocturnal systolic blood pressure (SBP) dipping: the dipper, non-dipper and reverse dipper groups. Baseline demographic characteristics, ambulatory blood pressure monitoring parameters, Holter recordings (including DC and AC), and echocardiographic parameters were collected. RESULTS: In this study, the lowest DC values were observed in the reverse dipper group, followed by the non-dipper and dipper groups (6.46 ± 2.06 vs. 6.65 ± 1.95 vs. 8.07 ± 1.79 ms, P < .001). Additionally, the AC gradually decreased (-6.32 ± 2.02 vs. -6.55 ± 1.95 vs. -7.80 ± 1.73 ms, P < .001). There was a significant association between DC (r = .307, P < .001), AC (r=-.303, P < .001) and nocturnal SBP decline. Furthermore, DC (ß = 0.785, P = .001) was positively associated with nocturnal SBP decline, whereas AC was negatively associated with nocturnal SBP (ß = -0.753, P = .002). By multivariate logistic regression analysis, deceleration capacity [OR (95% CI): 0.705 (0.594-0.836), p < .001], and acceleration capacity [OR (95% CI): 1.357 (1.141-1.614), p = .001] were identified as independent risk factors for blood pressure nondipper status. The analysis of ROC curves revealed that the area under the curve for DC/AC in predicting the circadian rhythm of blood pressure was 0.711/0.697, with a sensitivity of 73.4%/65.1% and specificity of 66.7%/71.2%. CONCLUSIONS: Abnormal DC and AC density were correlated with a blunted decline in nighttime SBP, suggesting a potential association between the circadian rhythm of blood pressure in essential hypertension patients and autonomic nervous dysfunction.

Ultralight Ultrafast Enzymes.

Inorganic materials depleted of heavy stable isotopes are known to deviate strongly in some physicochemical properties from their isotopically natural counterparts. Here we explored for the first time the effect of simultaneous depletion of the heavy carbon, hydrogen, oxygen and nitrogen isotopes on the bacterium E. coli and the enzymes expressed in it. Bacteria showed faster growth, with most proteins exhibiting higher thermal stability, while for recombinant enzymes expressed in depleted media, faster kinetics was discovered. At room temperature, luciferase, thioredoxin and dihydrofolate reductase and Pfu DNA polymerase showed up to a 250 % increase in activity compared to the native counterparts, with an additional ∼50 % increase at 10 °C. Diminished conformational and vibrational entropy is hypothesized to be the cause of the accelerated kinetics. Ultralight enzymes may find an application where extreme reaction rates are required.

Peripheral immunity is associated with the risk of incident dementia.

Central immunity components especially microglia in dementia have been well studied and corresponding immunotherapy gradually caught the attention. However, few studies focused on peripheral immunity and dementia. To address the issue, we examined the longitudinal association between incident dementia and peripheral immunity markers encompassing immune cell counts, and their derived ratios including neutrophil-to-lymphocyte ratio (NLR), platelet-to-lymphocyte ratio (PLR), systemic immune-inflammation index (SII), and lymphocyte-to-monocyte ratio (LMR), utilizing data of 361,653 participants from the UK Biobank (UKB). During a median follow-up of 8.99 years, 4239 participants developed dementia. The results revealed that increased innate immunity markers were associated with higher dementia risk (per SD increment hazard ratio [HR]; 95% confidence interval [CI] 1.14; 1.09-1.19 for neutrophils, 1.16; 1.11-1.20 for NLR and 1.11; 1.07-1.16 for SII), while increased adaptive immunity markers were associated with lower dementia risk (0.93; 0.90-0.97 for lymphocytes and 0.94; 0.90-0.98 for LMR). Our study pinpoints the differential role of innate and adaptive immunity in dementia incidence, which may provide some new perspectives in etiology and therapy of dementia.

Testing the link between isoaspartate and Alzheimer's disease etiology.

Isoaspartate (isoAsp) is a damaging amino acid residue formed in proteins as a result of spontaneous deamidation. IsoAsp disrupts protein structures, making them prone to aggregation. Here we strengthened the link between isoAsp and Alzheimer's disease (AD) by novel approaches to isoAsp analysis in human serum albumin (HSA), the most abundant blood protein and a major carrier of amyloid beta (Aß) and phosphorylated tau (p-tau) in blood. We discovered a reduced amount of anti-isoAsp antibodies (P < 0.0001), an elevated isoAsp level in HSA (P < 0.001), more HSA aggregates (P < 0.0001), and increased levels of free Aß (P < 0.01) in AD blood compared to controls. We also found that deamidation significantly reduces HSA capacity to bind with Aß and p-tau (P < 0.05). These suggest the presence in AD of a bottleneck in clearance of Aß and p-tau, leading to their increased concentrations in the brain and facilitating their aggregations there.

MiR-375 silencing attenuates pro-inflammatory macrophage response and foam cell formation by targeting KLF4.

Macrophage mediated inflammation and foam cell formation play crucial roles in the development of atherosclerosis. MiR-375 is a small noncoding RNA that significantly implicated in multiple tumor regulation and has been emerged as a novel biomarker for type 2 diabetes. However, the exact role of miR-375 on macrophage activation remains unknown. In the present study, we observed that miR-375 expression showed an up-regulated expression in atherosclerotic aortas, as well as in bone marrow derived macrophages (BMDMs) and mouse peritoneal macrophages (MPMs) isolated from ApoE deficiency mice and was gradually increased followed the Ox-LDL treated time. Functionally, miR-375 inhibition significantly decreased foam cell formation accompanied by up-regulated genes expression involved in cholesterol efflux but reduced genes expression implicated in cholesterol influx. Moreover, miR-375 silencing increased resolving M2 macrophage but reduced pro-inflammatory M1 macrophage markers expression. Such above effects can be reversed by miR-375 overexpression. Mechanistically, we noticed that miR-375 knockdown promoted KLF4 expression which was required for the ameliorated effect of miR-375 silencing on macrophage activation. Importantly, the consistent results in mRNA expression of M1 and M2 markers were observed in vivo, and miR-375-/-ApoE-/- mice significant decreased atherosclerotic lesions in the whole aorta and aortic sinus. Taken together, these evidences suggested that miR-375 knockdown attenuated macrophage activation partially through activation of KLF4-dependent mechanism.

First Experimental Evidence for Reversibility of Ammonia Loss from Asparagine.

Ammonia loss from L-asparaginyls is a nonenzymatic reaction spontaneously occurring in all proteins and eventually resulting in damaging isoaspartate residues that hamper protein function and induce proteinopathy related to aging. Here, we discuss theoretical considerations supporting the possibility of a full repair reaction and present the first experimental evidence of its existence. If confirmed, the true repair of L-asparaginyl deamidation could open new avenues for preventing aging and neurodegenerative diseases.

First Immunoassay for Measuring Isoaspartate in Human Serum Albumin.

Isoaspartate (isoAsp) is a damaging amino acid residue formed in proteins mostly as a result of spontaneous deamidation of asparaginyl residues. An association has been found between isoAsp in human serum albumin (HSA) and Alzheimer's disease (AD). Here we report on a novel monoclonal antibody (mAb) 1A3 with excellent specificity to isoAsp in the functionally important domain of HSA. Based on 1A3 mAb, an indirect enzyme-linked immunosorbent assay (ELISA) was developed, and the isoAsp occupancy in 100 healthy plasma samples was quantified for the first time, providing the average value of (0.74 ± 0.13)%. These results suggest potential of isoAsp measurements for supplementary AD diagnostics as well as for assessing the freshness of stored donor blood and its suitability for transfusion.

Sodium sulfite-driven Helicobacter pylori eradication: Unraveling oxygen dynamics through multi-omics investigation.

Due to the emergence and spread of multidrug resistance in Helicobacter pylori (H. pylori), its eradication has become difficult. Sodium sulfite (SS), a widely used food additive for ensuring food safety and storage, has been recognized as an effective nonbactericidal agent for H. pylori eradication. However, the mechanism by which H. pylori adapts and eventually succumbs under low- or no-oxygen conditions remains unknown. In this study, we aimed to evaluate the anti-H. pylori effect of SS and investigated the multiomics mechanism by which SS kills H. pylori. The results demonstrated that SS effectively eradicated H. pylori both in vitro and in vivo. H. pylori responds to the oxygen changes regulated by SS, downregulates the HcpE gene, which is responsible for redox homeostasis in bacteria, decreases the activities of enzymes related to oxidative stress, and disrupts the outer membrane structure, increasing susceptibility to oxidative stress. Furthermore, SS downregulates the content of cytochrome C in the microaerobic respiratory chain, leading to a sharp decrease in ATP synthesis. Consequently, the accumulation of triglycerides (TGs) in bacteria due to oxidative stress supports anaerobic respiration, meeting their energy requirements. The multifaceted death of H. pylori caused by SS does not result in drug resistance. Thus, screening of the redox homeostasis of HcpE as a new target for H. pylori infection treatment could lead to the development of a novel approach for H. pylori eradication therapy.

Study on the Strength and Failure Characteristics of Silty Mudstone Using Different Unloading Paths.

To investigate the strength and failure characteristics of silty mudstone using different stress paths, silt-like mudstone specimens were subjected to triaxial unloading tests. The results indicate the following. (1) When subjected to equivalent initial deviator stress levels and differing confining pressures, the peak stress, residual stress, and elastic modulus, exhibited during unloading, increased concordantly with greater initial confining pressure. Both the peak strain and residual strain increased with rising initial confining pressure. The increase in peak strain and residual strain initially decelerated, then noticeably increased, before ultimately decreasing again. Additionally, the unloading failure time and strain rate demonstrated a negative correlation as the confining pressure increased. (2) Under different initial deviatoric stress conditions, the peak stress, residual stress, and residual strain, under unloading confining pressure conditions, decreased as the initial deviatoric stress levels elevated. Conversely, the peak strain and elastic modulus initially increased, then decreased under increasing initial deviatoric stress conditions. The unloading failure time and strain rate were both observed to decrease as the initial deviatoric stress levels increased. (3) Utilizing the Mohr stress circle enabled the characterization of the shear strength variation in the specimens during the unloading process. The cohesion and internal friction angle remained relatively consistent across the different unloading stress paths appraised, with cohesion being greater in path I versus path II, whereas the internal friction angle exhibited an inverse relationship. (4) The specimen failed during unloading due to lateral expansion caused by unloading confining pressure and collapse failure. The failure fracture surfaces predominantly manifested shear failure morphologies.

Deamidation-related blood biomarkers show promise for early diagnostics of neurodegeneration.

BACKGROUND: The strongest risk factor of neurodegenerative diseases (NDDs) is aging. Spontaneous asparaginyl deamidation leading to formation of isoaspartate (isoAsp) has been correlated with protein aggregation in NDDs. METHODS: Two cohorts consisting of 140 subjects were studied. Cohort 1 contained patients with AD and healthy controls, while Cohort 2 recruited subjects with mild cognitive impairment (MCI), vascular dementia (VaD), frontotemporal dementia (FTD), Parkinson's disease (PD) and healthy controls. The levels of isoAsp in plasma human albumin (HSA), the most abundant protein in plasma, as well as the levels of immunoglobulin G (IgG) specific against deamidated HSA were measured. Apart from the memory tests, plasma biomarkers for NDDs reported in literature were also quantified, including amyloid beta (Aß) peptides Aß40 and Aß42, neurofilament light protein (NfL), glial fibrillary acidic protein (GFAP) and phosphorylated tau 181 (p-tau181) protein. RESULTS: Deamidation products of blood albumin were significantly elevated in vascular dementia and frontotemporal dementia (P < 0.05), but less so in PD. Intriguingly, the deamidation levels were significantly (P < 0.01) associated with the memory test scores for all tested subjects. Deamidation biomarkers performed superiorly (accuracy up to 92%) compared with blood biomarkers Aß42/Aß40, NfL, GFAP and p-tau181 in separating mild cognitive impairment from healthy controls. CONCLUSION: We demonstrated the diagnostic capacity of deamidation-related biomarkers in predicting NDDs at the early stage of disease, and the biomarker levels significantly correlated with cognitive decline, strongly supporting the role of deamidation in triggering neurodegeneration and early stages of disease development. Prospective longitudinal studies with a longer observation period and larger cohorts should provide a more detailed picture of the deamidation role in NDD progression.

5TNF-α and IL-1ß neutralization ameliorates angiotensin II-induced cardiac damage in male mice.

Inflammation is a key event in hypertensive organ damage, and TNF-α and IL-1ß are elevated in hypertension. In this study, we evaluated the effects of TNF-α and IL-1ß elevation on hypertensive cardiac damage by treatment with a bifunctional inflammatory inhibitor, TNF receptor 2-fragment crystalization-IL-1 receptor antagonist (TFI), which can neutralize these 2 cytokines simultaneously. A mouse hypertension model of angiotensin II (Ang II) infusion (1500 ng/kg·min for 7 d) was induced in wild-type mice. TNF-α and IL-1ß were inhibited by TFI administration (5 mg/kg, every other day), the effects of inhibition on cardiac damage were examined, and its mechanism on inflammatory infiltration was further studied in vivo and in vitro. Ang II infusion induced cardiac injury, including increased macrophage infiltration, expression of inflammatory cytokines (IL-12, IL-6, etc), and cardiac fibrosis, such as elevated α-smooth muscle actin, collagen I, and TGF-ß expression. Importantly, the Ang II-induced cardiac injury was suppressed by TFI treatment. Moreover, TFI reduced the expression of adhesion molecules (intercellular adhesion molecule-1 and vascular cell adhesion molecule-1) and monocyte chemotactic protein-1 expression in Ang II-treated hearts. Additionally, blockade of TNF-α and IL-1ß by TFI reduced monocyte adherence to endothelia cell and macrophage migration. This study demonstrates that blocking TNF-α and IL-1ß by TFI prevents cardiac damage in response to Ang II, and targeting these 2 cytokines simultaneously might be a novel tool to treat hypertensive heart injury.