Proposal and confirmation of N-(2-carboxyethyl)proline as a human endogenous metabolite.

RATIONALE: Malondialdehyde, one of the peroxidation products of polyunsaturated fatty acids, has been widely reported as an oxidative stress biomarker in many diseases. However, malondialdehyde is inherently unstable in biological matrices, which renders its measurement unreliable with all the reported analytical methods. To find an alternative oxidative stress biomarker, we envisioned that N-(2-carboxyethyl)proline, a modified conjugate of malondialdehyde and proline, could be a stable candidate for this purpose. METHODS: The proposed compound was chemically synthesized, and liquid chromatography-mass spectrometry methods were developed and used to search for the compound in human biological samples. RESULTS: An endogenous metabolite in human feces and urine samples was found to match the synthetic N-(2-carboxyethyl)proline by chromatographic retention and the fragmentation pattern of its molecular ion. CONCLUSION: The results confirmed that N-(2-carboxyethyl)proline was a new metabolite in human feces and urine samples. In addition, our results demonstrated a case of successful identification of true unknown metabolite by knowledge-based hypothesis of possible metabolites followed by experimental confirmation with a synthetic standard.

A review on spent Mn-containing Li-ion batteries: Recovery technologies, challenges, and future perspectives.

Mn-containing Li-ion batteries have become primary power sources for electronic devices and electric vehicles because of their high-energy density, extended cycle life, low cost, and heightened safety. In recent years, Li-ion batteries (LIBs) have undergone rapid updates, paralleling the swift advancement of the lithium battery industry, resulting in a growing accumulation of LIB scraps annually, necessitating comprehensive recovery strategies. This article reviews the recent progress in recovering spent Mn-containing LIBs (SM-LIBs), specifically focusing on LiMn2O4 and ternary LiCoxMnyNizO2 (NCM). Initially, the study analyzes the current resource profile of SM-LIBs and elucidates their service mechanisms. Subsequently, the study explores the recovery of SM-LIBs, discussing various methods such as the hydrometallurgical approach, combined pyrolytic treatment-wet leaching process, bioleaching pathway, and electrochemical extraction. These discussions include recovery processes, reaction principles, and technological features. In addition, this study evaluates the potential applications of these recovery technologies, considering aspects such as complexity, economic viability, energy consumption, environmental sustainability, and scalability. Finally, it summarizes the challenges associated with the comprehensive recovery and resource utilization of SM-LIBs and offers insights into future directions.

Mimic Pork Rinds from Plant-Based Gel: The Influence of Sweet Potato Starch and Konjac Glucomannan.

This study investigated the effect of sweet potato starch (SPS) and konjac glucomannan (KGM) on the textural, color, sensory, rheological properties, and microstructures of plant-based pork rinds. Plant-based gels were prepared using mixtures of soy protein isolate (SPI), soy oil, and NaHCO3 supplemented with different SPS and KGM concentrations. The texture profile analysis (TPA) results indicated that the hardness, cohesiveness, and chewiness of the samples improved significantly after appropriate SPS and KGM addition. The results obtained via a colorimeter showed no significant differences were found in lightness (L*) between the samples and natural pork rinds after adjusting the SPS and KGM concentrations. Furthermore, the rheological results showed that adding SPS and KGM increased both the storage modulus (G') and loss modulus (G''), indicating a firmer gel structure. The images obtained via scanning electron microscopy (SEM) showed that the SPS and KGM contributed to the formation of a more compact gel structure. A mathematical model allowed for a more objective sensory evaluation, with the 40% SPS samples and the 0.4% KGM samples being considered the most similar to natural pork rinds, which provided a comparable texture, appearance, and mouthfeel. This study proposed a possible schematic model for the gelling mechanism of plant-based pork rinds: the three-dimensional network structures of the samples may result from the interaction between SPS, SPI, and soybean oil, while the addition of KGM and NaHCO3 enabled a more stable gel structure.

Detection of Serum Cross-Reactive Antibodies and Memory Response to SARS-CoV-2 in Prepandemic and Post-COVID-19 Convalescent Samples.

BACKGROUND: A notable feature of coronavirus disease 2019 (COVID-19) is that children are less susceptible to severe disease. Children are known to experience more infections with endemic human coronaviruses (HCoVs) compared to adults. Little is known whether HCoV infections lead to cross-reactive anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antibodies. METHODS: We investigated the presence of cross-reactive anti-SARS-CoV-2 IgG antibodies to spike 1 (S1), S1-receptor-binding domain (S1-RBD), and nucleocapsid protein (NP) by enzyme-linked immunosorbent assays, and neutralizing activity by a SARS-CoV-2 pseudotyped virus neutralization assay, in prepandemic sera collected from children (n = 50) and adults (n = 45), and compared with serum samples from convalescent COVID-19 patients (n = 16). RESULTS: A significant proportion of children (up to 40%) had detectable cross-reactive antibodies to SARS-CoV-2 S1, S1-RBD, and NP antigens, and the anti-S1 and anti-S1-RBD antibody levels correlated with anti-HCoV-HKU1 and anti-HCoV-OC43 S1 antibody titers in prepandemic samples (P < .001). There were marked increases of anti-HCoV-HKU1 and - OC43 S1 (but not anti-NL63 and -229E S1-RBD) antibody titers in serum samples from convalescent COVID-19 patients (P < .001), indicating an activation of cross-reactive immunological memory to ß-coronavirus spike. CONCLUSIONS: We demonstrated cross-reactive anti-SARS-CoV-2 antibodies in prepandemic serum samples from children and young adults. Promoting this cross-reactive immunity and memory response derived from common HCoV may be an effective strategy against SARS-COV-2 and future novel coronaviruses.

Modified Vaccinia Ankara-Vectored Vaccine Expressing Nucleoprotein and Matrix Protein 1 (M1) Activates Mucosal M1-Specific T-Cell Immunity and Tissue-Resident Memory T Cells in Human Nasopharynx-Associated Lymphoid Tissue.

BACKGROUND: Increasing evidence supports a critical role of CD8+ T-cell immunity against influenza. Activation of mucosal CD8+ T cells, particularly tissue-resident memory T (TRM) cells recognizing conserved epitopes would mediate rapid and broad protection. Matrix protein 1 (M1) is a well-conserved internal protein. METHODS: We studied the capacity of modified vaccinia Ankara (MVA)-vectored vaccine expressing nucleoprotein (NP) and M1 (MVA-NP+M1) to activate M1-specific CD8+ T-cell response, including TRM cells, in nasopharynx-associated lymphoid tissue from children and adults. RESULTS: After MVA-NP+M1 stimulation, M1 was abundantly expressed in adenotonsillar epithelial cells and B cells. MVA-NP+M1 activated a marked interferon γ-secreting T-cell response to M1 peptides. Using tetramer staining, we showed the vaccine activated a marked increase in M158-66 peptide-specific CD8+ T cells in tonsillar mononuclear cells of HLA-matched individuals. We also demonstrated MVA-NP+M1 activated a substantial increase in TRM cells exhibiting effector memory T-cell phenotype. On recall antigen recognition, M1-specific T cells rapidly undergo cytotoxic degranulation, release granzyme B and proinflammatory cytokines, leading to target cell killing. CONCLUSIONS: MVA-NP+M1 elicits a substantial M1-specific T-cell response, including TRM cells, in nasopharynx-associated lymphoid tissue, demonstrating its strong capacity to expand memory T-cell pool exhibiting effector memory T-cell phenotype, therefore offering great potential for rapid and broad protection against influenza reinfection.

Local and Systemic Immunity against Respiratory Syncytial Virus Induced by a Novel Intranasal Vaccine. A Randomized, Double-Blind, Placebo-controlled Clinical Trial.

Rationale: Needle-free intranasal vaccines offer major potential advantages, especially against pathogens entering via mucosal surfaces. As yet, there is no effective vaccine against respiratory syncytial virus (RSV), a ubiquitous pathogen of global importance that preferentially infects respiratory epithelial cells; new strategies are urgently required.Objectives: Here, we report the safety and immunogenicity of a novel mucosal RSV F protein vaccine linked to an immunostimulatory bacterium-like particle (BLP).Methods: In this phase I, randomized, double-blind, placebo-controlled trial, 48 healthy volunteers, aged 18-49 years, were randomly assigned to receive placebo or SynGEM (low or high dose) intranasally by prime-boost administration. The primary outcome was safety and tolerability, with secondary objectives assessing virus-specific immunogenicity.Measurements and Main Results: There were no significant differences in adverse events between placebo and vaccinated groups. SynGEM induced systemic plasmablast responses and significant, durable increases in RSV-specific serum antibody in healthy, seropositive adults. Volunteers given low-dose SynGEM (140 µg F, 2 mg BLP) required a boost at Day 28 to achieve plateau responses with a maximum fold change of 2.4, whereas high-dose recipients (350 µg F, 5 mg BLP) achieved plateau responses with a fold change of 1.5 after first vaccination that remained elevated up to 180 days after vaccination, irrespective of further boosting. Palivizumab-like antibodies were consistently induced, but F protein site ∅-specific antibodies were not detected, and virus-specific nasal IgA responses were heterogeneous, with the strongest responses in individuals with lower pre-existing antibody levels.Conclusions: SynGEM is thus the first nonreplicating intranasal RSV subunit vaccine to induce persistent antibody responses in human volunteers.Clinical trials registered with www.clinicaltrials.gov (NCT02958540).

Galvanic Replacement-Mediated Synthesis of Ni-Supported Pd Nanoparticles with Strong Metal-Support Interaction for Methanol Electro-oxidation.

Herein, well-defined Pd nanoparticles (NPs) developed on Ni substrate (Pd NPs/Ni) are synthesized via a facile galvanic replacement reaction (GRR) route performed in ethaline-based deep eutectic solvent (DES). For comparison, a Pd NPs/Ni composite is also prepared by the GRR method conducted in an aqueous solution. The Pd NPs/Ni obtained from the ethaline-DES is catalytically more active and durable for the methanol electro-oxidation reaction (MOR) than those of the counterpart derived from conventional aqueous solution and commercial Pd/C under alkaline media. Detailed kinetic analysis indicates that the unique solvent environment offered by ethaline plays vital roles in adjusting the reactivity of the active species and their mass transport properties to control over the genesis of the Pd NPs/Ni nanocomposite. The resulting Pd NPs/Ni catalyst possesses a homogeneous dispersion of Pd NPs with a strong Pd (metal)-Ni (support) interaction. This structure enhances the charge transfer between the support and the active phases, and optimizes the adsorption energy of OH- and CO on the surface, leading to superior electrocatalytic performance. This work provides a novel GRR strategy performed in ethaline-DES to the rational design and construction of advanced metal/support catalysts with strong interaction for improving the activity and durability for MOR.

Activation and Induction of Antigen-Specific T Follicular Helper Cells Play a Critical Role in Live-Attenuated Influenza Vaccine-Induced Human Mucosal Anti-influenza Antibody Response.

There is increasing interest recently in developing intranasal vaccines against respiratory tract infections. The antibody response is critical for vaccine-induced protection, and T follicular helper cells (TFH) are considered important for mediating the antibody response. Most data supporting the role for TFH in the antibody response are from animal studies, and direct evidence from humans is limited, apart from the presence of TFH-like cells in blood. We studied the activation and induction of TFH and their role in the anti-influenza antibody response induced by a live-attenuated influenza vaccine (LAIV) in human nasopharynx-associated lymphoid tissue (NALT). TFH activation in adenotonsillar tissues was analyzed by flow cytometry, and anti-hemagglutinin (anti-HA) antibodies were examined following LAIV stimulation of tonsillar mononuclear cells (MNC). Induction of antigen-specific TFH by LAIV was studied by flow cytometry analysis of induced TFH and CD154 expression. LAIV induced TFH proliferation, which correlated with anti-HA antibody production, and TFH were shown to be critical for the antibody response. Induction of TFH from naive T cells by LAIV was shown in newly induced TFH expressing BCL6 and CD21, followed by the detection of anti-HA antibodies. Antigen specificity of LAIV-induced TFH was demonstrated by expression of the antigen-specific T cell activation marker CD154 upon challenge by H1N1 virus antigen or HA. LAIV-induced TFH differentiation was inhibited by BCL6, interleukin-21 (IL-21), ICOS, and CD40 signaling blocking, and that diminished anti-HA antibody production. In conclusion, we demonstrated the induction by LAIV of antigen-specific TFH in human NALT that provide critical support for the anti-influenza antibody response. Promoting antigen-specific TFH in NALT by use of intranasal vaccines may provide an effective vaccination strategy against respiratory infections in humans.IMPORTANCE Airway infections, such as influenza, are common in humans. Intranasal vaccination has been considered a biologically relevant and effective way of immunization against airway infection. The vaccine-induced antibody response is crucial for protection against infection. Recent data from animal studies suggest that one type of T cells, TFH, are important for the antibody response. However, data on whether TFH-mediated help for antibody production operates in humans are limited due to the lack of access to human immune tissue containing TFH In this study, we demonstrate the induction of TFH in human immune tissue, providing critical support for the anti-influenza antibody response, by use of an intranasal influenza vaccine. Our findings provide direct evidence that TFH play a critical role in vaccine-induced immunity in humans and suggest a novel strategy for promoting such cells by use of intranasal vaccines against respiratory infections.

Validation of a Metabolite Panel for a More Accurate Estimation of Glomerular Filtration Rate Using Quantitative LC-MS/MS.

BACKGROUND: Clinical practice guidelines recommend estimation of glomerular filtration rate (eGFR) using validated equations based on serum creatinine (eGFRcr), cystatin C (eGFRcys), or both (eGFRcr-cys). However, when compared with the measured GFR (mGFR), only eGFRcr-cys meets recommended performance standards. Our goal was to develop a more accurate eGFR method using a panel of metabolites without creatinine, cystatin C, or demographic variables. METHODS: An ultra-performance liquid chromatography-tandem mass spectrometry assay for acetylthreonine, phenylacetylglutamine, pseudouridine, and tryptophan was developed, and a 20-day, multiinstrument analytical validation was conducted. The assay was tested in 2424 participants with mGFR data from 4 independent research studies. A new GFR equation (eGFRmet) was developed in a random subset (n = 1615) and evaluated in the remaining participants (n = 809). Performance was assessed as the frequency of large errors [estimates that differed from mGFR by at least 30% (1 - P30); goal <10%]. RESULTS: The assay had a mean imprecision (≤10% intraassay, ≤6.9% interassay), linearity over the quantitative range (r 2 > 0.98), and analyte recovery (98.5%-113%). There was no carryover, no interferences observed, and analyte stability was established. In addition, 1 - P30 in the validation set for eGFRmet (10.0%) was more accurate than eGFRcr (13.1%) and eGFRcys (12.0%) but not eGFRcr-cys (8.7%). Combining metabolites, creatinine, cystatin C, and demographics led to the most accurate equation (7.0%). Neither equation had substantial variation among population subgroups. CONCLUSIONS: The new eGFRmet equation could serve as a confirmatory test for GFR estimation.

LC-MS/MS method with chemical derivatization for quantitation of L-threonate in human plasma.

An LC-MS/MS-based bioanalytical method has been developed to measure the concentration of L-threonate at its endogenous level in human plasma. Following isotope dilution and protein precipitation, the samples were acetylated and chromatographed under reversed-phase conditions for baseline separation of the derivatized L-threonate and its stereoisomer D-erythronate. The method was assessed by a fit-for-purpose validation with a calibration range from 100 to 10,000 ng/mL. The intra-run coefficients of variation (CVs) were <3.6% and the inter-run CV was 3.2% for the QC samples at endogenous level. At the lower limit of quantitation, the intra-run CV was 6.1% and the average inaccuracy was -1.4%. This method provides an efficient and reliable quantitation of L-threonate and could be useful to certain biomarker investigators.

Plant-expressed Fc-fusion protein tetravalent dengue vaccine with inherent adjuvant properties.

Dengue is a major global disease requiring improved treatment and prevention strategies. The recently licensed Sanofi Pasteur Dengvaxia vaccine does not protect children under the age of nine, and additional vaccine strategies are thus needed to halt this expanding global epidemic. Here, we employed a molecular engineering approach and plant expression to produce a humanized and highly immunogenic poly-immunoglobulin G scaffold (PIGS) fused to the consensus dengue envelope protein III domain (cEDIII). The immunogenicity of this IgG Fc receptor-targeted vaccine candidate was demonstrated in transgenic mice expressing human FcγRI/CD64, by induction of neutralizing antibodies and evidence of cell-mediated immunity. Furthermore, these molecules were able to prime immune cells from human adenoid/tonsillar tissue ex vivo as evidenced by antigen-specific CD4+ and CD8+ T-cell proliferation, IFN-γ and antibody production. The purified polymeric fraction of dengue PIGS (D-PIGS) induced stronger immune activation than the monomeric form, suggesting a more efficient interaction with the low-affinity Fcγ receptors on antigen-presenting cells. These results show that the plant-expressed D-PIGS have the potential for translation towards a safe and easily scalable single antigen-based tetravalent dengue vaccine.

MERS-CoV infection in humans is associated with a pro-inflammatory Th1 and Th17 cytokine profile.

The Middle East respiratory syndrome coronavirus (MERS-CoV) has been recognized as a highly pathogenic virus to humans that infects the respiratory tract and is associated with high morbidity and mortality. Studies in animal models suggest that MERS-CoV infection induces a strong inflammatory response, which may be related to the severity of disease. Data showing the cytokine profiles in humans during the acute phase of MERS-CoV infection are limited. In this study, we have analyzed the profile of cytokine responses in plasma samples from patients with confirmed MERS-CoV infections (n = 7) compared to healthy controls (n = 13). The cytokine profiles, including T helper (Th) 1, Th2 and Th17 responses, were analyzed using cytometric bead array (CBA). A prominent pro-inflammatory Th1 and Th17 response was clearly seen in patients with MERS-CoV infection, with markedly increased concentrations of IFN-γ, TNF-α, IL-15 and IL-17 compared to controls. IL-12 expression levels showed no difference between patients with MERS-CoV infection and the healthy controls despite the significantly increased levels of IFN-α2 and IFN-γ (P < .01). No changes were observed in the levels of IL-2, IL-4, IL-5, IL-13, and TGF-α (P > .05). Our results demonstrate a marked pro-inflammatory cytokine response during the acute phase of MERS-CoV infection in humans.

Infection with 2009 H1N1 influenza virus primes for immunological memory in human nose-associated lymphoid tissue, offering cross-reactive immunity to H1N1 and avian H5N1 viruses.

Influenza is a highly contagious mucosal infection in the respiratory tract. The 2009 pandemic H1N1 (pH1N1) influenza virus infection resulted in substantial morbidity and mortality in humans. Little is known on whether immunological memory develops following pH1N1 infection and whether it provides protection against other virus subtypes. An enzyme-linked immunosorbent spot assay was used to analyze hemagglutinin (HA)-specific memory B cell responses after virus antigen stimulation in nose-associated lymphoid tissues (NALT) from children and adults. Individuals with serological evidence of previous exposure to pH1N1 showed significant cross-reactive HA-specific memory B cell responses to pH1N1, seasonal H1N1 (sH1N1), and avian H5N1 (aH5N1) viruses upon pH1N1 virus stimulation. pH1N1 virus antigen elicited stronger cross-reactive memory B cell responses than sH1N1 virus. Intriguingly, aH5N1 virus also activated cross-reactive memory responses to sH1N1 and pH1N1 HAs in those who had previous pH1N1 exposure, and that correlated well with the memory response stimulated by pH1N1 virus antigen. These memory B cell responses resulted in cross-reactive neutralizing antibodies against sH1N1, 1918 H1N1, and aH5N1 viruses. The 2009 pH1N1 infection appeared to have primed human host with B cell memory in NALT that offers cross-protective mucosal immunity to not only H1N1 but also aH5N1 viruses. These findings may have important implications for future vaccination strategies against influenza. It will be important to induce and/or enhance such cross-protective mucosal memory B cells.

Characterisation of regulatory T cells in nasal associated lymphoid tissue in children: relationships with pneumococcal colonization.

Regulatory T cells (Treg) diminish immune responses to microbial infection, which may contribute to preventing inflammation-related local tissue damage and autoimmunity but may also contribute to chronicity of infection. Nasopharyngeal carriage of pneumococcus is common in young children and can persist for long periods but it is unknown whether the presence of Treg in the nasopharynx contributes to this persistence. We have investigated the numbers and activities of Foxp3+Treg in adenoidal tissues and their association with pneumococcal carriage in children. Expression of Treg cell-related markers including Foxp3, CD25, CD39, CD127 and CLTA4 were analysed by flow-cytometry in adenoidal mononuclear cells (MNC) and PBMC from children. Unfractionated MNC or Treg-depleted MNC were stimulated with a pneumococcal whole cell antigen (WCA) and T cell proliferation measured. Cytokine production by MNC was measured using a cytometric bead array. Higher numbers of CD25(high)Foxp3(high) Treg expressing higher CD39 and CTLA4 were found in adenoidal MNC than in PBMC. Children with pneumococcus positive nasopharyngeal cultures had higher proportions of Treg and expressed higher levels of CD39 and CTLA-4 than those who were culture negative (-). WCA induced adenoidal Treg proliferation which produce IL10 but not IL17, and CD4 T cell proliferation in Treg-depleted MNC was greater in pneumococcal culture positive than negative children. Significant numbers of Treg with an effector/memory phenotype which possess a potent inhibitory effect, exist in adenoidal tissue. The association of pneumococcal carriage with an increased frequency of adenoidal Treg suggests that Treg in nasal-associated lymphoid tissue (NALT) may contribute to the persistence of pneumococcus in children. Further studies to determine what component and mechanisms are involved in the promotion of Treg in NALT may lead to novel therapeutic or vaccination strategy against upper respiratory infection.

Live attenuated influenza vaccine induces broadly cross-reactive mucosal antibody responses to different influenza strains in tonsils.

Intranasal live attenuated influenza vaccine (LAIV) was used to stimulate tonsillar monocular cells (MNCs) following isolation. Haemagglutinin (HA) proteins of several influenza strains were used for the detection of HA-specific IgG, IgM and IgA antibodies using ELISA. Significant anti-sH1N1 HA IgG IgA and IgM antibody titres were detected in cell culture supernatants after stimulation (mean ± SE: 0.43 ± 0.09, mean ± SE: 0.23 ± 0.04 and mean ± SE: 0.47 ± 0.05 respectively, p < 0.01). LAIV stimulation of tonsillar MNCs induced significant IgG, IgA and IgM antibodies to the pH1N1 HA (mean ± SE:1.35 ± 0.12), (mean ± SE: 0.35 ± 0.06) and (mean ± SE: 0.58 ± 0.10) respectively, p < 0.01. Surprisingly, LAIV was shown to induce cross-reactive anti-aH5N1 HA antibodies (mean ± SE: 0.84 ± 0.20, p < 0.01) to avian influenza virus (aH5N1). Anti-H2N2 HA IgG antibody was also detected in the cell culture supernatants in a significant level after LAIV stimulation (mean ± SE: 0.93 ± 0.23, p < 0.01). High levels of anti-sH3N2 HA IgG antibody was discovered after LAIV stimulation of tonsillar MNCs, (mean ± SE: 1.2 ± 0.23p < 0.01). The current model of human nasal-associated lymphoid tissue (NALT) to evaluate B cells responses to LAIV was evident that it is a successful model to study future intranasal vaccines.

An integrated process for co-producing fermentable sugars and xylonate from sugarcane bagasse based on xylonic acid assisted pretreatment.

In this study, a renewable organic acid (xylonic acid), which can be prepared by the biooxidation of xylose, is used for pretreating sugarcane bagasse. The effects of reaction temperature and time on the release of fermentable xylose and glucose were investigated. On the basis of guaranteeing the good enzymatic hydrolysis efficiency and minimizing the effects of inhibitors, the pretreatment with 1 % xylnoic acid at 190 °C for 30 min was selected after optimization. In this case, 70 % xylose was released, while enzymatic hydrolysis yield was also up to 86.5 %. Meanwhile, the pretreated hydrolysate liquor was proved that it could be used for producing xylonate by biooxidation of Gluconobacter oxydans. Finally, the sequential process of the xylonic acid pretreatment and saccharification also clear the path for recycling the lignin as value-added bioproducts. Overall, this study presents a green-like strategy for fully exploiting sugarcane bagasse.

Robust memory humoral immune response to SARS-CoV-2 in the tonsils of adults and children.

Background: Adaptive humoral immunity against SARS-CoV-2 has mainly been evaluated in peripheral blood. Human secondary lymphoid tissues (such as tonsils) contain large numbers of plasma cells that secrete immunoglobulins at mucosal sites. Yet, the role of mucosal memory immunity induced by vaccines or natural infection against SARS-CoV-2 and its variants is not fully understood. Methods: Tonsillar mononuclear cells (TMNCs) from adults (n=10) and children (n=11) were isolated and stimulated using positive SARS-CoV-2 nasal swabs. We used endpoint enzyme-linked immunosorbent assays (ELISAs) for the measurement of anti-S1, -RBD, and -N IgG antibody levels and a pseudovirus microneutralization assay to assess neutralizing antibodies (nAbs) in paired serum and supernatants from stimulated TMNCs. Results: Strong systemic humoral response in previously SARS-CoV-2 infected and vaccinated adults and children was observed in accordance with the reported history of the participants. Interestingly, we found a significant increase in anti-RBD IgG (305 and 834 folds) and anti-S1 IgG (475 and 443 folds) in the stimulated TMNCs from adults and children, respectively, compared to unstimulated cells. Consistently, the stimulated TMNCs secreted higher levels of nAbs against the ancestral Wuhan strain and the Omicron BA.1 variant compared to unstimulated cells by several folds. This increase was seen in all participants including children with no known history of infection, suggesting that these participants might have been previously exposed to SARS-CoV-2 and that not all asymptomatic cases necessarily could be detected by serum antibodies. Furthermore, nAb levels against both strains were significantly correlated in adults (r=0.8788; p = 0.0008) and children (r = 0.7521; p = 0.0076), and they strongly correlated with S1 and RBD-specific IgG antibodies. Conclusion: Our results provide evidence for persistent mucosal humoral memory in tonsils from previously infected and/or vaccinated adults and children against recent and old variants upon re-exposure. They also highlight the importance of targeting mucosal sites with vaccines to help control infection at the primary sites and prevent potential breakthrough infections.

Xylooligosaccharides Production from Xylan Hydrolysis Using Recyclable Strong Acidic Cationic Exchange Resin as Solid Acid Catalyst.

As the emerging functional food additives, xylooligosaccharides are receiving high commercial interest due to their excellent gut microbiota modulation capacity, and accumulating studies have suggested that acidic hydrolysis for xylooligosaccharides preparation is the most convenient and cost-effective approach, whereas liquid acids are still limited due to the challenges in acid catalysts separation and products recovery. In the present study, a strong acidic cationic resin (NKC-9), as a recyclable solid acid catalyst, was successfully applied to xylooligosaccharides production by acidic hydrolysis of xylan. Additionally, a central composite design with response surface methodology was employed to optimize the conditions for maximizing xylooligosaccharides yields. The results suggested that xylooligosaccharides with the desired degree of polymerization (2-6) could be prepared, and the maximum yield was reached 47.7% in the case of 5% solid acid loading at 131 °C for 42 min. Finally, the recyclability of the solid acid catalysts confirmed that it was a cost-effective strategy for xylooligosaccharides production.

Myosin Light Chain Kinase Modulates to Improve Myocardial Hypoxia/Reoxygenation Injury.

Objective: The aim of this study was to evaluate whether myosin light chain kinase (MLCK) knockdown attenuated H9C2 cell hypoxia/reoxygenation (H/R) injury and downstream signaling pathway. Methods: The MLCK expression in H/R injury model H9C2 cell was determined by western blot and qRT-PCR. H/R cells were transfected with si-MLCK in the presence of P38 inhibitor (SB203580) or ERK inhibitor (U0126). Then, cell apoptosis was verified by flow cytometry. Apoptosis-related proteins were detected by western blot. The contents of reactive oxygen species (ROS), lactate dehydrogenase (LDH), superoxide dismutase (SOD), interleukin-6 (IL-6), interleukin (IL)-1ß (IL-1ß), and tumor necrosis factor-α (TNF-α) were measured using flow cytometry and colorimetric assays, respectively. Results: MLCK expression was higher in H/R cells. Knockdown of MLCK diminished the amounts of ROS, LDH, IL-6, IL-1ß, and TNF-α and elevated the release of SOD in H/R model H9C2 cells. Additionally, H/R injury induced the cumulative expression and phosphorylation of ERK and the phosphorylation of P38, whereas MLCK siRNA-treated cells showed decreased ERK1/2 and P38 activation. Inversely, P38 inhibitor (SB203580) and ERK inhibitor (U0126) could reverse the cardioprotective effects induced by si-MLCK. Conclusion: MLCK knockdown attenuated H/R injury in H9C2 cells via regulating the ERK/P38 signaling pathway. MLCK/ERK/p38 axis may provide novel insight into therapeutic targets to restrain I/R injury caused by revascularization therapy after acute myocardial infarction.

Biorefinery Cascade Processing for Converting Corncob to Xylooligosaccharides and Glucose by Maleic Acid Pretreatment.

Corncob as an abundant and low-cost waste resource has received increasing attention to produce value-added chemicals, it is rich in xylan and regarded as the most preferable feedstock for preparing high value added xylooligosaccharides. The use of xylooligosaccharides as core products can cut costs and improve the economic efficiency in biorefinery. In this study, maleic acid, as a non-toxic and edible acidic catalyst, was employed to pretreat corncob and produce xylooligosaccharides. Firstly, the response surface methodology experimental procedure was employed to maximize the yield of the xylooligosaccharides; a yield of 52.9% (w/v) was achieved with 0.5% maleic acid (w/v) at 155 °C for 26 min. In addition, maleic acid pretreatment was also beneficial to enhance the enzymatic hydrolysis efficiency, resulting in an enzymatic glucose yield of 85.4% (w/v) with a total of 10% solids loading. Finally, a total of 160 g of xylooligosaccharides and 275 g glucose could be produced from 1000 g corncob starting from the maleic acid pretreatment. Overall, a cascade processing for converting corncob to xylooligosaccharides and glucose by sequential maleic acid pretreatment and enzymatic hydrolysis was successfully designed for the corncob wastes utilization.