Assessing the effect of Maillard reaction products on the functionality and antioxidant properties of Amaranth-red seaweed blends.

Plant-based proteins, represented by amaranth in our study, embrace a potential as an ingredient for the functional-food formulation. However, their efficacy is hindered by inherent limitations in solubility, emulsification, and antioxidant traits. The Maillard reaction, a complex chemical-process resulting in a diverse array of products, including Maillard conjugates and Maillard reaction products (MRPs), can employ variable effects on these specific attributes. To elucidate the influence of this reaction and the MRPs on the aforementioned properties, we used a complex blend of dehydrated seaweed Gracilaria and amaranth protein to create a conjugate-MRP blend. Our investigations revealed that the resultant incorporation enhanced solubility, emulsification, and antioxidant properties, while the intermediates formed did not progress to advanced glycation stages. This change is likely attributed to the dual effect of conjugates that altered the secondary protein structure, while the generation and/or preservation of MRPs post ultrasonication and spray drying enhanced its antioxidant potential.

Quantifying the effects of pre-roasting on structural and functional properties of yellow pea proteins.

Roasting could modify the protein structure/conformation, contributing to changes in functional properties. Here we investigated the effects of pre-roasting on the extraction efficiency, structural and functional properties of pea protein concentrates and isolates (PPC and PPI) produced from yellow split peas. The shorter roasting times (150 °C, 10 and 20 min) had little effect on protein yields and could increase the solubility of PPC or PPI by âˆ¼ 12% at pH 7 and enhance the solubility of PPI by âˆ¼ 12% (10-min roasting) and âˆ¼ 24% (20-min roasting) at pH 3. However, a longer duration of pre-roasting (150 °C, 30 min) significantly reduced the extraction efficiency of PPC and PPI by âˆ¼ 30% and âˆ¼ 61%, respectively. Meanwhile, pre-roasting had minor effects on SDS-PAGE profiles and the secondary structures of pea proteins but significantly altered tertiary structures by reducing free sulfhydryl groups, increasing disulfide bonds and surface hydrophobicity. As for the emulsifying properties, pre-roasting improved the emulsion ability index (EAI) of PPC and PPI but decreased the emulsion stability index (ESI) of PPC and had no significant effect on PPI. Moreover, PPC and PPI with shorter pre-roasting duration (10 and 20 min) had endothermic peaks and showed a slight decrease in the denaturation temperature (Td) and the onset temperature (To), respectively. Overall, the study demonstrated that controlled pre-roasting at 150 °C for 10 min and 20 min altered protein structures (mainly tertiary structures), improving the solubility and EAI of pea proteins at pH 7, while retaining their thermal properties in comparison to unroasted samples.

On surface composition and stability of ß-carotene microcapsules comprising pea/whey protein complexes by synchrotron-FTIR microspectroscopy.

This study aims to elucidate the stability of spray dried ß-carotene microcapsules by identifying their surface composition using synchrotron-Fourier transform infrared (FTIR) microspectroscopy. To investigate the impact of enzymatic cross-linking and polysaccharide addition on heteroprotein, three wall materials were prepared: pea/whey protein blends (Con), cross-linked pea/whey protein blends (TG), and cross-linked pea/whey protein blends-maltodextrin complex (TG-MD). The TG-MD exhibited the highest encapsulation efficiency (>90 %) after 8 weeks of storage followed by TG and Con. Chemical images obtained using synchrotron-FTIR microspectroscopy confirmed that the TG-MD displayed the least amount of surface oil, followed by TG and Con, due to increasing amphiphilic ß-sheet structure of the proteins led by cross-linking and maltodextrin addition. Both enzymatic cross-linking and polysaccharide addition improved the stability of ß-carotene microcapsules, demonstrating that pea/whey protein blends with maltodextrin can be utilised as a hybrid wall material for enhancing the encapsulation efficiency of lipophilic bioactive compounds in foods.

Interferon-gamma FlowSpot assay for the measurement of the T-cell response to cytomegalovirus.

Objectives: We describe a new method, FlowSpot, to assess CMV-specific T-cell response by quantification of interferon-gamma (IFN-γ). CMV-specific, T-cell-released IFN-γ was captured by flow beads and measured via flow cytometry. In the present study, we used FlowSpot to assess CMV-specific T-cell response in healthy individuals. The FlowSpot results were compared with those of serological analysis and enzyme-linked immunospot (ELISpot) assay. Methods: Experimental results and parameter analysis were investigated by using serological, ELISpot, and FlowSpot assays. Results: The levels of IFN-γ, which is released from CMV-specific T-cells, were measured, and the results and parameter analysis showed a good correlation between FlowSpot and ELISpot. However, FlowSpot was more sensitive and better reflected the strength of IFN-γ secretion than did ELISpot. Conclusions: Compared to ELISpot, FlowSpot has a high sensitivity and is cost and time effective. Thus, this method can be used in wider clinical and scientific applications.

Visual Monitoring of Disintegration of Solid Oral Dosage Forms in Simulated Gastric Fluids Using Low-Field NMR Imaging.

Compared to traditional drug release monitoring with manual sampling and testing procedures, low-field nuclear magnetic resonance (LF-NMR) imaging is a one-step, visual, non-destructive, and non-invasive measurement method. Here, we reported the application of LF-NMR to image the morphology, component, sub-diffusion, and spatial distribution of a solid oral formulation, Biyankang tablets, during dissolution in vitro. The drug ingredients with characteristic relaxation times were distinguished and localized based on the signal of standards, such as patchouli oil, Xanthium strumarium extract, and starch. The hydration, swelling, disintegration, and sub-diffusion of tablets in simulated gastric fluids (SGF) were visualized statically. All tablets showed similar expansion (37.4-42.0%) along the direction of thickness at 25 min and reached a full disintegration at 145 min, at pH 1.80-6.15, indicating pH-independent swelling and disintegration. Compared to that static immersion within 20 mL SGF, the tablet disintegration time was shortened by ~ 11% in 30 mL SGF. The application of shear reduced the time by ~ 28%, suggesting a major role of hydrodynamic condition in tablet dissolution. The ability to simultaneously visualize, distinguish, and localize drug ingredients using LF-NMR is expected to provide valuable information to develop drug release monitoring systems in vitro and potentially in vivo using small animal studies.

Serum Soluble B Cell-Activating Factor Is a Non-Invasive Biomarker of Antibody-Mediated Rejection in Kidney Allograft With Satisfactory Risk Stratification Performance But Negligible Diagnostic Value.

Objectives: B cell-activating factor (BAFF), which is critical in the activation and differentiation of B cells, is a candidate diagnostic and predictive biomarker for antibody-mediated rejection (ABMR). We aimed to investigate the value of serum soluble BAFF (sBAFF) for the diagnosis and risk stratification of ABMR after kidney transplantation. Methods: In the diagnostic study, sBAFF level among ABMR (n = 25), T cell-mediated rejection (TCMR) (n = 14), 4 other pathological lesions (n = 21), and stable allograft function group (n = 15) were compared. In the nested case-control study, kidney allograft recipients with de novo donor-specific antibody (DSA) or ABMR (n = 16) vs. stable allograft function (n = 7) were enrolled, and sBAFF was measured preoperatively, at D7, M1, M3, M6, M9, M12, M18 posttransplant and at allograft biopsy. Results: There was no significant difference in sBAFF level at biopsy between ABMR and non-ABMR groups. Longitudinal study showed that the sBAFF levels decreased dramatically at D7 in both groups. The sBAFF level in the DSA group started to increase within M1, while in the stable group, it maintained a low level until M3 and M6. The sBAFF levels of the DSA group were significantly higher than that of the stable group at M1 [1,013.23 (633.97, 1,277.38) pg/ml vs. 462.69 (438.77, 586.48) pg/ml, P = 0.005], M3 [1,472.07 (912.79, 1,922.08) pg/ml vs. 561.63 (489.77, 630.00) pg/ml, P = 0.002], and M6 [1,217.95 (965.25, 1,321.43) pg/ml vs. 726.93 (604.77, 924.60) pg/ml, P = 0.027]. sBAFF levels at M3 had the best predictive value for the DSA/ABMR with the area under the receiver operating characteristic (AUROC) curve value of 0.908. The predictive performance of the maximum (max) change rate from D7 to the peak within M3 was also excellent (AUROC 0.949, P = 0.580). Conclusion: We clarified by a diagnostic study that sBAFF is not a diagnostic biomarker for ABMR in kidney transplantation and revealed by a nested case-control study that sBAFF values at M3 posttransplant and dynamic changes in sBAFF within M3 posttransplant have a good predictive value for the DSA/ABMR. It provides a useful tool for early screening of low-risk patients with negative preoperative DSA for the risk of developing postoperative DSA in kidney allograft recipients.

Digestion of curcumin-fortified yogurt in short/long gastric residence times using a near-real dynamic in vitro human stomach.

A dynamic in vitro human stomach (DIVHS), simulating the anatomical structures, peristalsis, and biochemical environments of a real stomach as practically as possible, was applied to mimic the gastric pH and emptying during yogurt digestion in short/long gastric residence times. The influences of peristalsis, dilution, and proteolysis on digesta viscosity were quantified respectively, indicating the dominant role of proteolysis and dilution. After incorporating curcumin-whey protein microparticles with targeted-release formula in yogurt, the peak curcumin release during intestinal digestion reached 43% at 120 min in the short gastric residence time and 16% at 180 min in the long gastric residence time. The change in the maximum curcumin release depended on the gastric emptying kinetics in each residence time. This emptying-kinetics dependence was reflected by the slower microparticle disintegration and proteolysis in the long gastric residence time. The dynamic reproduction of realistic gastric conditions using DIVHS helps revealing controlled release from foods.

Atractylenolide-I Sensitizes Triple-Negative Breast Cancer Cells to Paclitaxel by Blocking CTGF Expression and Fibroblast Activation.

This investigation was conducted to elucidate whether atractylenolide-I (ATL-1), which is the main component of Atractylodes macrocephala Koidz, can sensitize triple-negative breast cancer (TNBC) cells to paclitaxel and investigate the possible mechanism involved. We discovered that ATL-1 could inhibit tumor cell migration and increase the sensitivity of tumor cells to paclitaxel. ATL-1 downregulated the expression and secretion of CTGF in TNBC cells. Apart from inhibiting TNBC cell migration via CTGF, ATL-1 downregulated the expression of CTGF in fibroblasts and decreased the ability of breast cancer cells to transform fibroblasts into cancer-associated fibroblasts (CAFs), which in turn increased the sensitivity of TNBC cells to paclitaxel. In a mouse model, we found that ATL-1 treatments could enhance the chemotherapeutic effect of paclitaxel on tumors and reduce tumor metastasis to the lungs and liver. Primary cultured fibroblasts derived from inoculated tumors in mice treated with ATL-1 combined with paclitaxel expressed relatively low levels of CAF markers. Collectively, our data indicate that ATL-1 can sensitize TNBC cells to paclitaxel by blocking CTGF expression and fibroblast activation and could be helpful in future research to determine the value of ATL-1 in the clinical setting.

Efficacy and Safety of Bone Marrow-Derived Mesenchymal Stem Cells for Chronic Antibody-Mediated Rejection After Kidney Transplantation- A Single-Arm, Two-Dosing-Regimen, Phase I/II Study.

Objective: To investigate the efficacy and safety of bone marrow-derived mesenchymal stem cells (BM-MSCs) on chronic active antibody-mediated rejection (cABMR) in the kidney allograft. Methods: Kidney recipients with biopsy-proven cABMR were treated with allogeneic third-party BM-MSCs in this open-label, single-arm, single-center, two-dosing-regimen phase I/II clinical trial. In Regimen 1 (n=8), BM-MSCs were administered intravenously at a dose of 1.0×106 cells/kg monthly for four consecutive months, while in Regimen 2 (n=15), the BM-MSCs dose was 1.0×106 cells/kg weekly during four consecutive weeks. The primary endpoints were the absolute change of estimated glomerular filtration rate (eGFR) from baseline (delta eGFR) and the incidence of adverse events associated with BM-MSCs administration 24 months after the treatment. Contemporaneous cABMR patients who did not receive BM-MSCs were retrospectively analyzed as the control group (n =30). Results: Twenty-three recipients with cABMR received BM-MSCs. The median delta eGFR of the total BM-MSCs treated patients was -4.3 ml/min per 1.73m2 (interquartile range, IQR -11.2 to 1.2) 2 years after BM-MSCs treatment (P=0.0233). The median delta maximum donor-specific antibody (maxDSA) was -4310 (IQR -9187 to 1129) at 2 years (P=0.0040). The median delta eGFR of the control group was -12.7 ml/min per 1.73 m2 (IQR -22.2 to -3.5) 2 years after the diagnosis, which was greater than that of the BM-MSCs treated group (P=0.0342). The incidence of hepatic enzyme elevation, BK polyomaviruses (BKV) infection, cytomegalovirus (CMV) infection was 17.4%, 17.4%, 8.7%, respectively. There was no fever, anaphylaxis, phlebitis or venous thrombosis, cardiovascular complications, or malignancy after BM-MSCs administration. Flow cytometry analysis showed a significant decreasing trend of CD27-IgD- double negative B cells subsets and trend towards the increase of CD3+CD4+PD-1+/lymphocyte population after MSCs therapy. Multiplex analysis found TNF-α, CXCL10, CCL4, CCL11 and RANTES decreased after MSCs treatment. Conclusion: Kidney allograft recipients with cABMR are tolerable to BM-MSCs. Immunosuppressive drugs combined with intravenous BM-MSCs can delay the deterioration of allograft function, probably by decreasing DSA level and reducing DSA-induced injury. The underlying mechanism may involve immunomodulatory effect of MSCs on peripheral B and T cells subsets.

On improving bioaccessibility and targeted release of curcumin-whey protein complex microparticles in food.

Curcumin is a bioactive food component, with poor bioaccessibility due to low water solubility and stability. Spray drying retained and in fact enhanced curcumin-whey protein isolate (WPI) complexation via desolvation, lowering the amount of unbound curcumin to <5% wt after drying, forming microparticles with better water solubility, stability, and bioaccessibility than raw curcumin. The desolvated microparticles encapsulated 3.47 ± 0.05 mg/g curcumin, almost one order of magnitude higher than the un-desolvated sample 0.37 ± 0.03 mg/g. After incorporation into yogurt, the rapid-release formula liberated 87% curcumin, whereas the targeted-release one discharged 44% before entering the simulated intestinal condition. Most of the yogurt sensory properties were not adversely affected, except for colour and curcumin flavour. This study proposed a strategy in which food ingredients containing hydrophobic bioactive small molecules can be incorporated into a food matrix to improve bioaccessibility and targeted release, without affecting their sensory properties.

Comparison of long-read methods for sequencing and assembly of a plant genome.

BACKGROUND: Sequencing technologies have advanced to the point where it is possible to generate high-accuracy, haplotype-resolved, chromosome-scale assemblies. Several long-read sequencing technologies are available, and a growing number of algorithms have been developed to assemble the reads generated by those technologies. When starting a new genome project, it is therefore challenging to select the most cost-effective sequencing technology, as well as the most appropriate software for assembly and polishing. It is thus important to benchmark different approaches applied to the same sample. RESULTS: Here, we report a comparison of 3 long-read sequencing technologies applied to the de novo assembly of a plant genome, Macadamia jansenii. We have generated sequencing data using Pacific Biosciences (Sequel I), Oxford Nanopore Technologies (PromethION), and BGI (single-tube Long Fragment Read) technologies for the same sample. Several assemblers were benchmarked in the assembly of Pacific Biosciences and Nanopore reads. Results obtained from combining long-read technologies or short-read and long-read technologies are also presented. The assemblies were compared for contiguity, base accuracy, and completeness, as well as sequencing costs and DNA material requirements. CONCLUSIONS: The 3 long-read technologies produced highly contiguous and complete genome assemblies of M. jansenii. At the time of sequencing, the cost associated with each method was significantly different, but continuous improvements in technologies have resulted in greater accuracy, increased throughput, and reduced costs. We propose updating this comparison regularly with reports on significant iterations of the sequencing technologies.

Mucosal-associated invariant T-cells are severely reduced and exhausted in humans with chronic HBV infection.

Chronic hepatitis B virus (CHBV) infection is a major cause of liver diseases. Mucosal-associated invariant T (MAIT) cells are important for antiviral immunity in the liver, but the distinction between intrasinusoidal and peripheral MAIT cells in patients with CHBV infections remains unclear. PBMCs were obtained from patients with CHBV infections (n = 29) and age-matched controls (n = 46). Liver-associated mononuclear cells (LMCs) were collected from healthy donors (n = 29) and explanted livers (n = 19) from patients and used for phenotypic, functional and TCR diversity analyses. The percentages of both peripheral and intrasinusoidal MAIT cells were significantly reduced in the CHBV infection group compared to the control group. Peripheral MAIT cells from CHBV-infected patients expressed higher levels of HLA-DR, CD69, CD38 and PD-1 than those of controls. We also confirmed that peripheral MAIT cells in HBV patients had elevated expression T-cell exhaustion genes. Except for a difference in the level of PD-1, no differences were observed between the liver MAIT cells of the two groups. The production of IFN-α in peripheral MAIT cells of CHBV infection patients was lower than in control patients, but no such difference was observed in liver MAIT cells. Additionally, a distinct TCR signature was found in CHBV patients. Hence, we found distinct activities and functions in liver and peripheral MAIT cells of patients with CHBV infections.

Modification of molecular conformation of spray-dried whey protein microparticles improving digestibility and release characteristics.

This study reports on the preparation of riboflavin-loaded whey protein isolate (WPI) microparticles, using desolvation and then spray drying. Ethanol desolvation led to the exposure of embedded hydrophobic amino acids of WPI to riboflavin, facilitating the formation of riboflavin-WPI complexes. The extent of desolvation and cross-linking influenced the morphology of the spray-dried microparticles, while the moisture content of microparticles decreased with desolvation and increased with crosslinking. The modification of WPI conformation upon desolvation could be retained in the dry state via spray drying. The gastric resistance, release site and release characteristics of microparticles were readily adjusted by varying the ethanol and calcium ion contents from 0 to 50% v/v and from 0 to 2 mM, respectively. The sample prepared from 30% v/v ethanol without calcium crosslinking displayed rapid peptic digestion in less than 30 min. The samples from 30% v/v ethanol at 1 and 2 mM Ca2+ exhibited excellent gastric resistance and intestinal release.

Ultrasonically synthesized organic liquid-filled chitosan microcapsules: part 2: characterization using AFM (atomic force microscopy) and combined AFM-confocal laser scanning fluorescence microscopy.

Atomic Force Microscopy (AFM) is used to measure the stiffness and Young's modulus of individual microcapsules that have a chitosan cross-linked shell encapsulating tetradecane. The oil filled microcapsules were prepared using a one pot synthesis via ultrasonic emulsification of tetradecane and crosslinking of the chitosan shell in aqueous solutions of acetic acid. The concentration of acetic acid in aqueous solutions of chitosan was varied from 0.2% to 25% v/v. The effect of acetic acid concentration and size of the individual microcapsules on the strength was probed. The deformations and forces required to rupture the microcapsules were also measured. Three dimensional deformations of microcapsules under large applied loads were obtained by the combination of Laser Scanning Confocal Microscopy (LSCM) with Atomic Force Microscopy (AFM). The stiffness, and hence the modulus, of the microcapsules was found to decrease with an increase in size with the average stiffness ranging from 82 to 111 mN m-1 and average Young's modulus ranging from 0.4 to 6.5 MPa. The forces required to rupture the microcapsules varied from 150 to 250 nN with deformations of the microcapsules up to 62 to 110% relative to their radius, respectively. Three dimensional images obtained using laser scanning confocal microscopy showed that the microcapsules retained their structure and shape after being subjected to large deformations and subsequent removal of the loads. Based on the above observations, the oil filled chitosan crosslinked microcapsules are an ideal choice for use in the food and pharmaceutical industries as they would be able to withstand the process conditions encountered.

Ultrasonically synthesized organic liquid-filled chitosan microcapsules: part 1: tuning physical & functional properties.

This study reports the synthesis of tetradecane-filled chitosan microcapsules in acetic acid aqueous solutions using high intensity ultrasound at 20 kHz. The size, size distribution, and stability of microcapsules were tuned by varying the concentration of acetic acid from 0.2% to 25% v/v. After long-time storage at room temperature (more than 3 months), the microcapsules maintained their shell-core structure where the volume of the microcapsules at 0.2% acetic acid concentration increased by 8.3% due to leaking and coalescence. Microcapsules were consistently spherical and had a smooth shell surface, however, their shell thickness varied with acetic acid concentration. The relaxation behavior of individual microcapsules to an applied constant stress was measured by atomic force microscopy (AFM) to probe the shell strength and extent of crosslinking. The effect of acetic acid on the relative viscosity of chitosan aqueous solutions played a major role in microcapsule size control at low acid concentrations. With constant addition of acetic acid, amino groups in chitosan chains were acetylated partially under ultrasonic irradiation. This reduced the amphiphilicity of the shell material and therefore influenced the size, size distribution, stability and mechanical strength of the microcapsules. Apart from the acetylation effect, the counter-ion effect and the formation of covalent bond crosslinks also made contributions to the formation of stable chitosan microcapsules.