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Search Results (884)

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17 pages, 716 KiB  
Article
Fabrication and Evaluation of Polyhydroxyalkanoate-Based Nanoparticles for Curcumin Delivery in Biomedical Applications
by Fawzia Sha’at, Dana Miu, Mihaela Carmen Eremia, Georgeta Neagu, Adrian Albulescu, Radu Albulescu, Mihaela Deaconu, Mariana Gratiela Vladu and Ramona-Daniela Pavaloiu
Molecules 2025, 30(6), 1216; https://doi.org/10.3390/molecules30061216 (registering DOI) - 8 Mar 2025
Abstract
This study investigates the fabrication and characterization of polymeric nanoparticles based on polyhydroxyalkanoates (PHAs) loaded with curcumin for biomedical applications. PHAs, biodegradable and biocompatible polymers, were synthesized via bacterial fermentation and used to encapsulate curcumin using the nanoprecipitation method. The resulting nanoparticles were [...] Read more.
This study investigates the fabrication and characterization of polymeric nanoparticles based on polyhydroxyalkanoates (PHAs) loaded with curcumin for biomedical applications. PHAs, biodegradable and biocompatible polymers, were synthesized via bacterial fermentation and used to encapsulate curcumin using the nanoprecipitation method. The resulting nanoparticles were characterized for their particle size, polydispersity index, and encapsulation efficiency, achieving high entrapment rates (above 80%) and nanometric size distribution. Stability assessments demonstrated prolonged structural integrity under storage conditions. In vitro release studies conducted in phosphate-buffered saline at pH 5 and 7.4 revealed sustained drug release profiles. Biocompatibility and cytotoxicity assays using human astrocytes and fibroblasts confirmed the nanoparticles’ safety, while antiproliferative tests on glioblastoma and colon cancer cell lines indicated potential therapeutic efficacy. Additionally, skin irritation and corrosion tests using the EpiDerm™ model classified the formulations as non-irritant and non-corrosive. These findings suggest that PHA-based nanoparticles offer a promising nanocarrier system for curcumin delivery, with potential applications in cancer treatment and regenerative medicine. Future research should focus on optimizing the formulation and evaluating in vivo therapeutic effects. Full article
(This article belongs to the Special Issue Molecular Approaches to Drug Discovery and Development)
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<p>The release profiles of curcumin from nanoparticles based on polyhydroxyalkanoates in PBS 0.1 M pH 5 (<b>A</b>) compared to PBS 0.1 M pH 7.4 (<b>B</b>). The release followed the trend PHH &lt; PHO &lt; PHN in both media, indicating a composition-dependent effect on drug release.</p>
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<p>Antiproliferative effects of curcumin and PHA-based nanoparticles loaded with curcumin on U87 cells (<b>A</b>,<b>B</b>) and Caco-2 cells (<b>C</b>,<b>D</b>) after 24 and 48 h. Control is referring to untreated cells. Data are presented as the mean ± standard deviation. The results are considered statistically significant at <span class="html-italic">p</span> &lt; 0.05.</p>
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14 pages, 1779 KiB  
Article
Role of Bioimpedance Phase Angle and Hand Grip Strength in Predicting 12-Month Mortality in Patients Admitted with Haematologic Cancer
by Lara Dalla Rovere, Rocio Fernández-Jiménez, Alessandro Guerrini, María García-Olivares, Cristina Herola-Cobos, Carmen Hardy-Añón, Rahinatu Awol-Tanko, Agustín Hernandez-Sanchez and José Manuel García-Almeida
Cancers 2025, 17(5), 886; https://doi.org/10.3390/cancers17050886 - 5 Mar 2025
Viewed by 85
Abstract
Background/Objectives: Haematologic cancers, such acute leukaemia, lymphoma, and multiple myeloma, are associated with high morbidity and mortality rates, often exacerbated by malnutrition and functional decline. This study aims to evaluate the prognostic value of bioimpedance phase angle (PhA) and hand grip strength [...] Read more.
Background/Objectives: Haematologic cancers, such acute leukaemia, lymphoma, and multiple myeloma, are associated with high morbidity and mortality rates, often exacerbated by malnutrition and functional decline. This study aims to evaluate the prognostic value of bioimpedance phase angle (PhA) and hand grip strength (HGS) as nutritional and clinical markers for predicting 12-month mortality in hospitalized patients with haematologic cancers. Methods: A retrospective observational study was conducted on 121 patients admitted to Hospital Quironsalud Málaga between January 2019 and June 2021. PhA was measured using bioelectrical impedance analysis (BIA) and HGS was assessed using a dynamometer. Nutritional status was evaluated through Subjective Global Assessment (SGA) and the Global Leadership Initiative on Malnutrition (GLIM) criteria. The primary outcome was 12-month mortality, analysed using ROC curves, Kaplan–Meier survival estimates, and multivariate logistic regression models. Results: Lower PhA (<3.8° for females, <5.4° for males) and reduced HGS (<17 kg for females, <28 kg for males) were significantly associated with higher 12-month mortality (p < 0.001). The optimal PhA cut-off showed high sensitivity (85.5%) and specificity (62.2%). Multivariate analysis confirmed PhA as an independent predictor of mortality (OR = 0.417, p = 0.023). Patients with lower PhA and HGS values exhibited reduced survival rates, emphasizing the importance of these markers in clinical practice. Conclusions: PhA and HGS are reliable, non-invasive tools for assessing prognosis in haematologic cancer patients. Incorporating these markers into routine care could improve risk stratification, guide nutritional interventions, and enhance patient outcomes. Full article
(This article belongs to the Section Cancer Epidemiology and Prevention)
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<p>Clinical outcomes related to phase angle in haematological cancers patients, with red points indicating mortality cases. The ellipses represent the 50th, 75th, and 95th percentiles of normality, indicating reference ranges for bioimpedance measurements. R: resistance; H: height; Xc: reactance.</p>
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<p>ROC curve analyses for PhA and SPhA variables to predict 12-month mortality. (<b>a</b>) Overall and gender ROC curve analysis for PhA. (<b>b</b>) ROC curve analysis for SPhA.</p>
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<p>Kaplan–Meier survival curves of patients stratified by PhA and SPhA. (<b>a</b>) Survival curves for patients with PhA ≥ 3.8° (solid blue line) and PhA &lt; 3.8° (dotted red line). (<b>b</b>) Survival curves for patients with SPhA ≥ −1.5 (solid blue line) and SPhA &lt; −1.5 (dotted red line).</p>
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<p>ROC curve analyses for HGS variables to predict 12-month mortality.</p>
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<p>Kaplan–Meier survival curves of patients stratified by HGS. The solid blue line represents patients with HGS ≥ 28 kg for men and ≥17 kg for women, while the dotted red line represents patients with HGS &lt; 28 kg for men and &lt;17 kg for women.</p>
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19 pages, 9203 KiB  
Article
Anti-Inflammatory and Immunomodulatory Properties of Inorganic Fullerene-Like Tungsten Disulfide Nanoparticles in the Culture of Human Peripheral Blood Mononuclear Cells
by Snežana Zečević, Darinka Popović, Sergej Tomić, Marina Bekić, Sara Rakočević, Maja Kosanović, Dušica Stojanović, Petar Uskoković, Milan Marković, Dejan Bokonjić and Miodrag Čolić
Nanomaterials 2025, 15(5), 322; https://doi.org/10.3390/nano15050322 - 20 Feb 2025
Viewed by 371
Abstract
Tungsten disulfide (WS2) nanoparticles have emerged in the biomedical field as potential theranostic agents due to their unique properties, including biocompatibility. However, their impact on the immune response remains unexplored. This study aimed to evaluate the effects of inorganic fullerene-like WS [...] Read more.
Tungsten disulfide (WS2) nanoparticles have emerged in the biomedical field as potential theranostic agents due to their unique properties, including biocompatibility. However, their impact on the immune response remains unexplored. This study aimed to evaluate the effects of inorganic fullerene-like WS2 (IF-WS2) nanostructures on human peripheral blood mononuclear cells (PBMCs) in vitro. The study investigated several parameters to evaluate the effects of IF-WS2 nanoparticles. Cytotoxicity was assessed by measuring cell viability, apoptosis, and necrosis. Internalization of IF-WS2 by PBMCs was analyzed using morphological and flow cytometric techniques. Proliferation was studied in CellTrace Far Red-prestained total PBMCs stimulated with phytohemagglutinin (PHA) and in isolated T cell cultures stimulated with CD3/CD28-coated beads. Additionally, the production of cytokines and chemokines was measured in culture supernatants of total PBMCs and T cells. IF-WS2 nanoparticles were non-cytotoxic up to a concentration of 200 µg/mL. Concentrations ≥25 µg/mL inhibited PHA-stimulated PBMC proliferation but did not affect T cell proliferation. Morphological and flow cytometric analysis demonstrated dose- and time-dependent internalization of IF-WS2 by macrophages. Additionally, IF-WS2 significantly reduced the production of pro-inflammatory cytokines (IL-1β, TNF-α, IL-8, MCP-1, and GRO-α) in PHA-stimulated PBMCs. Th1, Th17, and Th21 cytokines were downregulated, while Th2, Th9, and T regulatory cytokines were upregulated. In conclusion, this study demonstrated for the first time that pristine IF-WS2 nanoparticles, at non-cytotoxic concentrations, exhibit notable anti-inflammatory and immunomodulatory properties on activated PBMCs in vitro. Full article
(This article belongs to the Section Biology and Medicines)
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<p>NTA measurements of IF-WS<sub>2</sub> size distribution and Z-potential. (<b>A</b>) Size distribution profiles from three independent measurements are shown on a normalized <span class="html-italic">Y</span>-axis, with the summarized graph presented beside. (<b>B</b>) Z-potential, as measured by NTA, is shown as mean mV ± SD (<span class="html-italic">n</span> = 3). (<b>C</b>) Concentration distribution according to IF-WS<sub>2</sub> size is shown, with each point displayed as mean ± SD (<span class="html-italic">n</span> = 3), and the summarized graph beside shows the mean concentration (number/mL) ± SD. * <span class="html-italic">p</span> &lt; 0.05, ** <span class="html-italic">p</span> &lt; 0.01 as indicated (one-way ANOVA).</p>
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<p>The effect of different concentrations of IF-WS<sub>2</sub> nanoparticles on viability (<b>A</b>), apoptosis (<b>B</b>,<b>D</b>), and necrosis (<b>C</b>,<b>D</b>) in PBMC cultures. Values are presented as mean percentages (%) ± SD (<span class="html-italic">n</span> = 6) or as flow cytometric data of one representative experiment (<b>D</b>). * <span class="html-italic">p</span> &lt; 0.05 compared to the control (one-way ANOVA).</p>
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<p>Internalization of IF-WS<sub>2</sub> nanoparticles in PBMC cultures. (<b>A</b>) Heterogeneity of IF-WS<sub>2</sub> uptake by macrophages after 48 h, where phagocytic indexes (PhI) are indicated above the cells. (<b>B</b>) Internalization of IF-WS<sub>2</sub> by macrophages, but not lymphocytes, analyzed after 48 h using cytospins. (<b>C</b>) The dose- and time-dependent effect of IF-WS<sub>2</sub> internalization by macrophages is shown. Values are expressed as mean ± SD of PhI (<span class="html-italic">n</span> = 3), determined by calculating PhI for 500 cells on each cytospin. * <span class="html-italic">p</span> &lt; 0.05 compared to the lowest concentration (12.5 µg/mL) or as indicated by bars and determined by two-way ANOVA followed by Tukey’s multiple comparison test.</p>
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<p>Internalization of IF-WS<sub>2</sub> by PBMCs in culture. PBMCs were cultured with IF-WS<sub>2</sub> for 24 h, followed by analysis by flow cytometry. (<b>A</b>) Gating strategy showing the population of monocytes/macrophages (above) and lymphocytes (below). (<b>B</b>) Mean values of size scatter for each population (monocytes/macrophages above, lymphocytes below) are presented. Data are shown as mean ± SD (<span class="html-italic">n</span> = 3), with representative histograms included. * <span class="html-italic">p</span> &lt; 0.05; ** <span class="html-italic">p</span> &lt; 0.01 as indicated on the horizontal bars (one-way ANOVA).</p>
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<p>The effect of IF-WS<sub>2</sub> nanoparticles on the proliferation of PHA-stimulated PBMCs. (<b>A</b>) Cell proliferation parameters are presented as relative values (%) ± SD (<span class="html-italic">n</span> = 3), with control cultures used as 100%. * <span class="html-italic">p</span> &lt; 0.05; ** <span class="html-italic">p</span> &lt; 0.01; *** <span class="html-italic">p</span> &lt; 0.005 compared to the control (one-way ANOVA). (<b>B</b>) Gating strategy. (<b>C</b>) Representative histograms from one experiment. The percentages above the bars represent the proportion of non-dividing cells. PI—proliferating index.</p>
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<p>The effect of IF-WS<sub>2</sub> nanoparticles on the production of pro-inflammatory cytokines and chemokines in PHA-stimulated PBMC cultures. Values are presented as mean ± SD (<span class="html-italic">n</span> = 3). * <span class="html-italic">p</span> &lt; 0.05; ** <span class="html-italic">p</span> &lt; 0.01; *** <span class="html-italic">p</span> &lt; 0.005 compared to the control (one-way ANOVA).</p>
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<p>The effect of IF-WS<sub>2</sub> nanoparticles on the production of Th1, Th2, and Th9 cytokines in PHA-stimulated PBMC cultures. Values are given as mean ± SD (<span class="html-italic">n</span> = 3). * <span class="html-italic">p</span> &lt; 0.05; ** <span class="html-italic">p</span> &lt; 0.01; *** <span class="html-italic">p</span> &lt; 0.005 compared to the control (one-way ANOVA).</p>
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<p>The effect of IF-WS<sub>2</sub> nanoparticles on the production of Th17, Th21, and Treg cytokines in PHA-stimulated PBMC cultures. Values are presented as mean ± SD (<span class="html-italic">n</span> = 3). * <span class="html-italic">p</span> &lt; 0.05; ** <span class="html-italic">p</span> &lt; 0.01; *** <span class="html-italic">p</span> &lt; 0.005 compared to the control (one-way ANOVA).</p>
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18 pages, 4108 KiB  
Article
Mining the Candidate Transcription Factors Modulating Tanshinones’ and Phenolic Acids’ Biosynthesis Under Low Nitrogen Stress in Salvia miltiorrhiza
by Yating Cheng, Siqi Gui, Siyu Hao, Xiujuan Li, Chao Zhuang, Yifei Shi, Wei Zhou and Guoyin Kai
Int. J. Mol. Sci. 2025, 26(4), 1774; https://doi.org/10.3390/ijms26041774 - 19 Feb 2025
Viewed by 215
Abstract
Mining valuable genes is helpful to breed high-quality Salvia miltiorrhiza exhibiting efficient nitrogen fertilizer utilization efficiency. In the present study, transcriptome sequencing was introduced to select the candidate transcription factors (TFs) involved in tanshinones’ (TAs) and phenolic acids’ (PHAs) biosynthesis as well as [...] Read more.
Mining valuable genes is helpful to breed high-quality Salvia miltiorrhiza exhibiting efficient nitrogen fertilizer utilization efficiency. In the present study, transcriptome sequencing was introduced to select the candidate transcription factors (TFs) involved in tanshinones’ (TAs) and phenolic acids’ (PHAs) biosynthesis as well as low nitrogen (LN) stress. In totally, 97.71 Gb clean data was obtained from fifteen sequencing samples and 30,975 unigenes were assembled. Among of them, 27,843 unigenes were successfully annotated. Overall, 8663 differential expression genes (DEGs) were identified, among of which 5034 unigenes were up-regulated, and 3629 unigenes were down-regulated. By enrichment of DEGs together with gene co-expression network construction, 10 candidate TFs including HSFB2b, LBD12, ERF1A, ERF98, LBD25, HSF24, RAM1, HSFA4B, TCP8, and WRKY24 were finally retrieved, which are predicted to participate in modulating TA and PHA biosynthesis under LN stress. Quantitative real-time polymerase chain reaction (qRT-PCR) detection was introduced to further detect the expression profile of candidate TFs under LN stress. These findings offer a valuable resource for in-depth study of TAs ‘and PHAs’ biosynthesis under LN stress in S. miltiorrhiza. Full article
(This article belongs to the Section Molecular Plant Sciences)
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<p>Content of TAs (<b>a</b>) and PHAs (<b>b</b>) in <span class="html-italic">S. miltiorrhiza</span> hairy roots under LN stress. DT, Dihydrotanshinone; CT, cryptotanshinone; TI, tanshinone I; TIIA, tanshinone IIA; TT, total of TAs. CA, caffeic acid; RA, rosmarinic acid; SalB, salvianolic acid B; TPA, total of PHAs. Error bars represent SD (n = 3), ** <span class="html-italic">p</span> &lt; 0.01.</p>
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<p>Annotation of unigenes. The bar chart on the left denotes the total number of unigenes; the connecting line represents annotated unigenes; and the number above indicates the number of unigenes annotated in the databases.</p>
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<p>DEGs in response to LN stress. (<b>a</b>) Venn diagram for DEGs compared with each of the LN-induced time points (1, 4, 12 and 48 h) and the Mock (0 h), respectively. (<b>b</b>) The gene expression level under LN treatment at 1, 4, 12 and 48 h compared with the Mock (0 h), respectively.</p>
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<p>Functional enrichment analysis of DEGs. (<b>a</b>) GO enrichment analysis of DEG. The y-axis represents the GO terms, and the x-axis represents the level of significance of enrichment. The larger the log<sub>10</sub> (<span class="html-italic">p</span>-adjust) value, the more significantly enriched the GO term is. (<b>b</b>) KEGG enrichment analysis of DEGs. The y-axis represents the KEGG pathway and the x-axis represents rich factor values; the size of the circles denotes the gene number. DEGs were mined using DESeq2 (version 1.24.0), with |log<sub>2</sub>FC| ≥ 1 and <span class="html-italic">p</span>-adjust &lt; 0.05 (DESeq2) as the cutoff.</p>
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<p>Gene expression and classification of transcription factors. (<b>a</b>) Expression profiles of TAs’ biosynthetic genes. (<b>b</b>) Expression profiles of PHAs’ biosynthetic genes. (<b>c</b>) The top 10 TF families with more than 65 members are listed. Solid arrows represent the confirmed synthetic pathways, while dashed arrows indicate intermediate processes that are uncertain and have not yet been resolved in the synthesis pathway.</p>
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<p>DEGs involved in LN stress. (<b>a</b>) Expression profiles of marker genes related to nitrogen uptake. (<b>b</b>) Co-expression network between differentially expressed marker genes involved in nitrogen uptake and candidate TFs. The light purple circles represent transcription factors with a co-expression coefficient greater than 0.9; the orange ones are the final ten selected transcription factors; and the rose-pink circles represent nitrogen-related genes.</p>
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<p>Co-expression relationship of candidate TFs and synthetase genes involved in TAs’ and PHAs’ biosynthesis. (<b>a</b>) Co-expression relationship of candidate TFs with synthetase genes related to TAs’ biosynthesis. (<b>b</b>) Co-expression relationship of candidate TFs with synthetase genes related to PHAs’ biosynthesis.The light purple circles represent transcription factors with a co-expression coefficient greater than 0.9; the orange ones are the final ten selected transcription factors; and the rose-pink circles represent differentially expressed synthases.</p>
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<p>qRT-PCR validation of DEGs in RNA-seq dataset. Expression of 6 TFs related to TAs’ and PHAs’ biosynthesis in response to LN induction using qRT-PCR detection and comparison of their expression with RNA-seq dataset. Error bars denote standard error (±SD). All the detections were triply repeated, * <span class="html-italic">p</span> &lt; 0.05; ** <span class="html-italic">p</span> &lt; 0.01.</p>
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13 pages, 2884 KiB  
Article
The Production and Characterization of an Aminolyzed Polyhydroxyalkanoate Membrane and Its Cytocompatibility with Osteoblasts
by Qiulan Luo, Fuming Zou, Dongjuan Yang, Yongping Huang, Dajie Xian, Ying Nie, Zhenxia Zhang, Yuzhong Zheng, Yaqun Liu, Fei Zhou, Peikui Yang, Yuting Jiang, Xianjing Huang and Xianghui Zou
Molecules 2025, 30(4), 950; https://doi.org/10.3390/molecules30040950 - 18 Feb 2025
Viewed by 193
Abstract
Polyhydroxyalkanoates (PHAs), recognized as a medical biomaterial, have been proven to promote cell proliferation and tissue repair. PHA has a variety of forms: PHB, PHV, PHHx, and PHBHHx, etc. In this study, PHBHHx was selected as the precursor to fabricate biopolyester films. Specifically, [...] Read more.
Polyhydroxyalkanoates (PHAs), recognized as a medical biomaterial, have been proven to promote cell proliferation and tissue repair. PHA has a variety of forms: PHB, PHV, PHHx, and PHBHHx, etc. In this study, PHBHHx was selected as the precursor to fabricate biopolyester films. Specifically, a novel type of biopolyester film was generated through an ammonolysis cross-linking reaction in conjunction with polyamidoamine dendrimer G2.0 (PAMAM). The properties of the resultant biopolyester films were comprehensively evaluated, encompassing surface characteristics, amino group content, and water contact angle. The drug-loading properties and compatibility with osteoblasts of the biopolyester films were also determined. The findings revealed that following aminolysis, the biopolyester film surface exhibited enhanced roughness and an enlarged surface area. Moreover, as the aminolysis duration extended, the hydrophilicity and drug-loading efficiency were significantly augmented. Post-aminolysis, the PHBHHx membrane exhibited a more favorable environment for the adhesion and proliferation of osteoblasts. Overall, the biopolyester film developed in this study provides novel insights and materials for tissue engineering, especially bone tissue repair. Full article
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<p>Properties of PHBHHx aminolyzed biopolyester films. (<b>A</b>) Surface morphology of PHBHHx biopolyester films with different ammonolysis times, observed using an SEM at 5000 times magnification. (<b>B</b>) Standard curve of the NH<sub>2</sub> content using CEDA as a reference standard. (<b>C</b>) The density of NH<sub>2</sub> in PHBHHx aminolyzed biopolyester films with different ammonolysis times. Different letters indicate significance by one-way ANOVA with Tukey’s post hoc test (<span class="html-italic">p</span> &lt; 0.05).</p>
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<p>Ibuprofen release curve on PHBHHx aminolyzed biopolyester films.</p>
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<p>The cell numbers (<b>A</b>) and cell proliferation multiples (<b>B</b>) of osteoblasts on PHBHHx aminolyzed biopolyester films.</p>
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<p>Cell morphology of osteoblasts on PHBHHx aminolyzed biopolyester film. The red border highlights the adhesion of osteoblasts on a PHBHHx biopolyester film that has been aminolyzed for 24 h, and these osteoblasts are magnified 5000 times.</p>
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21 pages, 5063 KiB  
Article
Investigating Polyhydroxyalkanoate Synthesis for Insights into Drug Resistance in Xanthomonas oryzae pv. oryzae
by Qingbiao Xie, Guangshu Lao, Yukai Fang, Xue Gao, Zheng Tan, Weiguo Miao and Pengfei Jin
Int. J. Mol. Sci. 2025, 26(4), 1601; https://doi.org/10.3390/ijms26041601 - 13 Feb 2025
Viewed by 313
Abstract
Polyhydroxyalkanoates (PHAs), synthesized by Xanthomonas to endure adverse conditions, are primarily regulated by the critical genes phaC and phaZ. Poly-3-hydroxybutyrate (PHB), a common polyhydroxyalkanoate (PHA), has been implicated in metabolism, pathogenicity, and various physiological processes in Xanthomonas oryzae pv. oryzae (Xoo [...] Read more.
Polyhydroxyalkanoates (PHAs), synthesized by Xanthomonas to endure adverse conditions, are primarily regulated by the critical genes phaC and phaZ. Poly-3-hydroxybutyrate (PHB), a common polyhydroxyalkanoate (PHA), has been implicated in metabolism, pathogenicity, and various physiological processes in Xanthomonas oryzae pv. oryzae (Xoo). In this study, we investigated the effects of HN-2 using n-butanol extract (HN-2 n-butanol extract) derived from Bacillus velezensis on Xoo. The results showed that HN-2 n-butanol extract could induce PHB accumulation in Xoo, potentially via surfactin. Moreover, examination of drug resistance, pathogenicity, and morphological characteristics of Xoo revealed PHB played a significant role in the drug resistance, pathogenicity, membrane integrity, and growth rate of Xoo strains following the deletion of phaZ and phaC. The ∆phaZ strain was the most significant, with a growth rate reduced to 58.19% of the PXO99A at 36 h and an inhibition zone 57.46% larger than that of PXO99A by HN-2 n-butanol extract. Transmission electron microscopy further revealed blank spots in Xoo after treatment, with the fewest spots observed in ∆phaZ, indicating its impaired ability to repair and maintain membrane integrity. These findings offer valuable insights that could serve as a foundation for elucidating the mechanisms of drug resistance and future research on preventing Xoo-induced diseases. Full article
(This article belongs to the Section Molecular Plant Sciences)
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<p>Characteristics of PHA metabolism-related genes. The PHA metabolism-related genes cluster distribution in <span class="html-italic">PXO99<sup>A</sup></span> genome and phaC and phaZ protein structure predicted by AlphaFold2 (<b>A</b>). Content analysis of PHB from <span class="html-italic">phaC/phaZ</span> deficiency strains (<b>B</b>). The data are shown as the means with SD (±SD) with two-way ANOVA followed by Tukey’s multiple mean comparisons test method, the letters represent significance, <span class="html-italic">p</span> &lt; 0.05.</p>
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<p>Investigation of drug-resistance of <span class="html-italic">Xoo</span> to HN-2 n-butanol extract. Disk diffusion assays were conducted to evaluate the sensitivity of the wild-type strain <span class="html-italic">PXO99<sup>A</sup></span>, ∆<span class="html-italic">phaC</span>, ∆<span class="html-italic">phaZ</span>, ∆<span class="html-italic">phaC</span>/∆<span class="html-italic">phaZ</span>, C: ∆<span class="html-italic">phaC</span>, and C: ∆<span class="html-italic">phaZ</span> to the HN-2 n-butanol extract, using 6-mm diameter paper disks. Bacitracin was used as a control for comparison (<b>A</b>). The diameters of the inhibition zones for the HN-2 n-butanol extract were measured and statistically analyzed (<b>B</b>). The data are shown as the means with SD (±SD) with two-way ANOVA followed by Tukey’s multiple mean comparisons test method, the letters represent significance, <span class="html-italic">p</span> &lt; 0.05.</p>
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<p>Effect of HN-2 n-butanol extract on the virulence of <span class="html-italic">Xoo</span> strains during rice infection. The lesion morphology on rice variety IR24 was observed 14 days post inoculation (dpi) with wild-type <span class="html-italic">PXO99<sup>A</sup></span>, ∆<span class="html-italic">phaC</span>, ∆<span class="html-italic">phaZ</span>, and ∆<span class="html-italic">phaC</span>/∆<span class="html-italic">phaZ</span> (<b>A</b>). The measurements of lesion lengths were shown in sequence in (<b>B</b>). Additionally, bacterial colony-forming units (CFUs) used to quantify the multiplication of <span class="html-italic">Xoo</span> in rice leaves were presented in sequence in (<b>C</b>). Treatment conditions were as follows: CK (control): ddH<sub>2</sub>O; T1: <span class="html-italic">Xoo</span> inoculation without treatment; T2: treated with HN-2 n-butanol extract at one day post-<span class="html-italic">Xoo</span> inoculation; T3: treated with bacitracin at one day post-<span class="html-italic">Xoo</span> inoculation; T4: treated with HN-2 n-butanol extract one day before <span class="html-italic">Xoo</span> inoculation; and T5: treated with bacitracin one day before <span class="html-italic">Xoo</span> inoculation. The data are shown as the means with SD (±SD) with two-way ANOVA followed by Tukey’s multiple mean comparisons test method, the letters represent significance, <span class="html-italic">p</span> &lt; 0.05.</p>
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<p>Morphology of <span class="html-italic">Xoo</span> cells under HN-2 n-butanol extract treatment. The morphology of <span class="html-italic">PXO99<sup>A</sup></span> and ∆<span class="html-italic">phaC</span>, ∆<span class="html-italic">phaZ,</span> and ∆<span class="html-italic">phaC</span>/∆<span class="html-italic">phaZ,</span> cells was observed using transmission electron microscopy (TEM). The effects of HN-2 n-butanol extract on <span class="html-italic">Xoo</span> morphology were also assessed. Red arrows indicate the presence of PHB.</p>
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<p>Influence of HN-2 n-butanol extract on the leakage of intracellular components in <span class="html-italic">Xoo</span>. The relative conductivity (<b>A</b>) and in vitro protein content (<b>B</b>) of the wild-type strain <span class="html-italic">PXO99<sup>A</sup></span>, ∆<span class="html-italic">phaC</span>, ∆<span class="html-italic">phaZ</span>, ∆<span class="html-italic">phaC</span>/∆<span class="html-italic">phaZ</span>, C: ∆<span class="html-italic">phaC</span>, and C: ∆<span class="html-italic">phaZ</span> were measured under HN-2 n-butanol extract treatment, with bacitracin used as a control. The data are shown as the means with SD (±SD) with two-way ANOVA followed by Tukey’s multiple mean comparisons test method, the letters represent significance, <span class="html-italic">p</span> &lt; 0.05.</p>
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<p>Determination of biofilm biosynthesis <span class="html-italic">Xoo</span> under HN-2 n-butanol extract treatment. The biofilm biosynthetic content of the wild-type strain <span class="html-italic">PXO99<sup>A</sup></span>, ∆<span class="html-italic">phaC</span>, ∆<span class="html-italic">phaZ</span>, ∆<span class="html-italic">phaC</span>/∆<span class="html-italic">phaZ</span>, C: ∆<span class="html-italic">phaC</span>, and C: ∆<span class="html-italic">phaZ</span> was assessed by measuring the optical density (OD) at 570 nm to evaluate the effect of HN-2 n-butanol extract on <span class="html-italic">Xoo</span> biofilm biosynthesis. The data are shown as the means with SD (±SD) with two-way ANOVA followed by Tukey’s multiple mean comparisons test method, the letters represent significance, <span class="html-italic">p</span> &lt; 0.05.</p>
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<p>Determination of the growth rate of <span class="html-italic">Xoo</span> under HN-2 n-butanol extract treatment. The impact of HN-2 n-butanol extract on the growth rate of the wild-type strain <span class="html-italic">PXO99<sup>A</sup></span>, ∆<span class="html-italic">phaC</span>, ∆<span class="html-italic">phaZ</span>, ∆<span class="html-italic">phaC</span>/∆<span class="html-italic">phaZ</span>, C: ∆<span class="html-italic">phaC</span>, and C: ∆<span class="html-italic">phaZ</span> was also determined by measuring the optical density (OD) at 600 nm. The data are shown as the means with SD (±SD).</p>
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14 pages, 1292 KiB  
Article
Differential Expression of miR-223-3p and miR-26-5p According to Different Stages of Mastitis in Dairy Cows
by Eleonora Dall’Olio, Fabio De Rensis, Eugenio Martignani, Silvia Miretti, Ugo Ala, Valeria Cavalli, Claudio Cipolat-Gotet, Melania Andrani, Mario Baratta and Roberta Saleri
Biomolecules 2025, 15(2), 235; https://doi.org/10.3390/biom15020235 - 6 Feb 2025
Viewed by 561
Abstract
Mastitis is the leading cause of economic losses in dairy farming, significantly impairing animal welfare and the quality and quantity of milk production. MicroRNAs are increasingly gaining attention, in both human and veterinary medicine, as biomarkers for various diseases. This study evaluated the [...] Read more.
Mastitis is the leading cause of economic losses in dairy farming, significantly impairing animal welfare and the quality and quantity of milk production. MicroRNAs are increasingly gaining attention, in both human and veterinary medicine, as biomarkers for various diseases. This study evaluated the diagnostic potential of four circulating microRNAs (miR-26-5p, miR-142-5p, miR-146a, and miR-223-3p) by examining changes in their expression in milk samples from dairy cows at different immune-cell subpopulations correlated to different stage of mastitis with a validated method. Additionally, this study has analyzed the possible source of these circulating microRNAs by the measurement of their secretion from activated immune cells (lymphocytes, monocytes, and neutrophils). miR-223-3p has been significantly expressed in an acute stage of mastitis (p < 0.01) but not in the chronic or susceptible stages. Conversely, mir-26-5p has been significantly reduced in acute, chronic, and susceptible groups of animals. In immune-cell cultures, miR-26 has been shown to be down-regulated in lipopolysaccharide (LPS)-stimulated neutrophils, while miR-223 has been shown to be up-regulated in phytohemagglutinin (PHA)-stimulated lymphocytes. The differential expression of miR-223-3p and miR-26-5p, combined with differential and total somatic cell count, could serve as a useful tool for identifying the evolutionary stage of mastitis-related inflammatory pathology. Full article
(This article belongs to the Special Issue The Role of Non-Coding RNAs in Health and Disease)
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<p>Scatter plot showing the distribution of dairy cows based on the two thresholds for somatic cell count (2 × 10<sup>5</sup> somatic cells/mL) and differential somatic cell count (68.5%).</p>
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<p>Relative expression levels reported as mean ± SD of (<b>A</b>) bta-miR-26a-5p and (<b>B</b>) bta-miR-223-3p in milk samples from each group. Data were analyzed using the 2<sup>−ΔΔCq</sup> method, in which the expression levels of the gene have been normalized to the expression of the reference genes bta-miR-148a-3p and UniSp2. * indicates a significant difference with a <span class="html-italic">p</span>-value &lt; 0.05; ** indicates a significant difference with a <span class="html-italic">p</span>-value &lt; 0.01. CTRL = control, AM = with acute mastitis, SU = susceptible, CM = with chronic mastitis.</p>
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<p>Correlation between bta-miR-223 and bta-miR-26. Data have been normalized to the expression of the reference genes bta-miR-148a-3p and UniSp2. R and <span class="html-italic">p</span>-value are displayed on the graphs.</p>
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<p>Relative expression levels reported as mean ± SD of bta-miR-26a-5p in (<b>A</b>) lymphocytes, (<b>B</b>) monocytes, (<b>C</b>) neutrophils, and of bta-miR-223-3p in (<b>D</b>) lymphocytes, (<b>E</b>) monocytes, and (<b>F</b>) neutrophils’ culture medium, unstimulated or stimulated for 4 h with PHA or LPS. Each group includes samples of 3 replicates from three separate experiments. Data were analyzed using the 2<sup>−ΔΔCq</sup> method, in which the expression levels of the gene have been normalized to the expression of the reference genes bta-miR-148a-3p and UniSp2. * indicates a significant difference with a <span class="html-italic">p</span>-value &lt; 0.05; ** indicates a significant difference with a <span class="html-italic">p</span>-value &lt; 0.01.</p>
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15 pages, 1239 KiB  
Article
Decreased Lower Limb Phase Angle in Older People Is an Indicator of Standing and Gait Function, Regardless of Age
by Daisuke Homma, Norio Imai, Dai Miyasaka, Moeko Yamato, Masafumi Ishisaki, Tsubasa Sugahara, Yoji Horigome, Hayato Suzuki, Yoichiro Dohmae, Naoto Endo, Izumi Minato and Hiroyuki Kawashima
J. Clin. Med. 2025, 14(3), 1023; https://doi.org/10.3390/jcm14031023 - 6 Feb 2025
Viewed by 546
Abstract
Background/Objectives: The phase angle (PhA), as measured using bioelectrical impedance analysis (BIA), indicates muscle mass and quality. However, its relationship with age-related muscle changes and motor function and effective BIA assessment/intervention sites remains unclear. Herein, we evaluated age-related changes in PhA, explored [...] Read more.
Background/Objectives: The phase angle (PhA), as measured using bioelectrical impedance analysis (BIA), indicates muscle mass and quality. However, its relationship with age-related muscle changes and motor function and effective BIA assessment/intervention sites remains unclear. Herein, we evaluated age-related changes in PhA, explored the relationship between PhA and muscle mass, and identified effective sites for BIA. Methods: We included 131 healthy community-dwelling adults divided into older (≥65 years) and control (≤65 years) groups. PhA and muscle mass were measured using BIA with a device determining electrical resistance by transmitting a weak alternating current <90 μA. Motor function was measured using ground reaction force index, gait function, and grip strength. The relationships between muscle mass, PhA, and motor function were analyzed. Results: All values, excluding upper limb muscle mass, were lower in the older group, as were the rates of change in muscle mass and PhA in the lower limbs vs. the upper limbs. Lower limb PhA showed negative and positive correlations with the Timed Up and Go test and standing function, respectively; it was significantly associated with all motor functions. Conclusions: Regardless of age, muscle mass and PhA measured by BIA were high and low in the lower limbs, respectively, and lower limb PhA was related to walking and standing function. The decrease in lower limb muscle mass and PhA may accelerate with age. Given global population aging, easy-to-assess lower limb PhA linked to the movements necessary for independent living may be an effective site for assessment and intervention in clinical practice. Full article
(This article belongs to the Section Clinical Rehabilitation)
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<p>Patient recruitment flowchart. Overall, 134 cases were included during the measurement period; 3 participants (2 with measurement errors and 1 with numbness in the lower limbs) were excluded. Thus, the data analyzed were from 131 participants. Participants aged 65 years or older were placed in the older group (<span class="html-italic">n</span> = 60), while those aged 65 years or younger were assigned to the control group (<span class="html-italic">n</span> = 71).</p>
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<p>The BIA measurement position. Prior to taking measurements, the participants’ electrodes and skin were cleansed with alcohol. All measurements were conducted under uniform conditions. During the procedure, participants stood with bare feet placed on the toe and heel electrodes while holding the hand grip, with their arms hanging a few centimeters from their bodies.</p>
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<p>Measurement of ground reaction force index when standing up. The sit-to-stand movement was performed three times under maximum effort. The graphs show F/w and RFD8.75/w as indices representing the ground reaction force during standing. F/w is calculated as the maximum ground reaction force divided by body weight, while RFD8.75/w measures the rate of change in ground reaction force over 87.5 milliseconds, which includes 37.5 milliseconds prior to and 12.5 milliseconds following the maximum recorded increase, then converted to per second and normalized by body weight. This figure was derived from prior studies [<a href="#B30-jcm-14-01023" class="html-bibr">30</a>,<a href="#B31-jcm-14-01023" class="html-bibr">31</a>] with some modifications.</p>
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17 pages, 11750 KiB  
Article
Preparation and Optimization of a Polyhydroxyoctanoate–Hydroxyapatite Composite Available to Scaffolds in Implantable Devices
by Dana-Maria Miu, Ramona Daniela Pavaloiu, Fawzia Sha’at, Mariana-Gratiela Vladu, Georgeta Neagu, Vasile-Sorin Manoiu and Mihaela-Carmen Eremia
Molecules 2025, 30(3), 730; https://doi.org/10.3390/molecules30030730 - 6 Feb 2025
Viewed by 410
Abstract
Biomaterials represent a distinct class of materials used in various medical applications, such as replicating the shape or function of damaged tissue caused by disease or trauma. The increasing focus on polyhydroxyalkanoate (PHA) research can be attributed to their properties, such as biodegradability, [...] Read more.
Biomaterials represent a distinct class of materials used in various medical applications, such as replicating the shape or function of damaged tissue caused by disease or trauma. The increasing focus on polyhydroxyalkanoate (PHA) research can be attributed to their properties, such as biodegradability, biocompatibility, and bioresorbability. PHAs can be incorporated into polymeric complexes or combined with bioceramics or bioactive substances. Films of PHO-HAp-Curcumin were prepared, and optimization studies were conducted using Design-Expert software (Stat-Ease 360-Trial Version). The effects of independent variables (amount of PHO, HAp, and curcumin) on biodegradability, film thickness, and curcumin release were studied. Statistical modeling revealed significant interactions among the components, with the 2FI and quadratic models providing strong predictive accuracy. The interaction of HAp and PHO amounts (X2X3) has a significant effect on biodegradability (Y1) and film thickness (Y3). For the degree of the cumulative release of curcumin (CDR), there was no significant interaction between the independent variables (curcumin-X1, HAp-X2, and PHO-X3). Optimized film exhibited a maximum desirability of 0.777 with 1 mg of curcumin, 100 mg of HAp, and 172.31 mg of PHO. A morphological analysis of optimized film revealed a rough, particle-rich surface favorable for biomedical use. The findings highlight the promise of PHO-HAp-Curcumin composite films in advancing tissue engineering. Full article
(This article belongs to the Special Issue Synthesis and Applications of Natural Polymers and Their Derivatives)
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<p>Structure of polyhydroxyalkanoates (PHAs).</p>
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<p>The effect of the interaction on the degree of biodegradability (Y<sub>1</sub>)—(<b>A</b>) between the amount of curcumin and the amount of HAp (X<sub>1</sub>X<sub>2</sub>), (<b>B</b>) between the amount of curcumin and the amount of PHO (X<sub>1</sub>X<sub>3</sub>), and (<b>C</b>) between the amount of HAp and the amount of PHO (X<sub>2</sub>X<sub>3</sub>).</p>
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<p>The effect of the interaction on the film thickness (Y<sub>3</sub>)—(<b>A</b>) between the amount of curcumin and the amount of HAp (X<sub>1</sub>X<sub>2</sub>), (<b>B</b>) between the amount of curcumin and the amount of PHO (X<sub>1</sub>X<sub>3</sub>), and (<b>C</b>) between the amount of HAp and the amount of PHO (X<sub>2</sub>X<sub>3</sub>).</p>
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<p>The effect of the interaction on the CDR (Y<sub>4</sub>)—(<b>A</b>) between the amount of curcumin and the amount of HAp (X<sub>1</sub>X<sub>2</sub>), (<b>B</b>) between the amount of curcumin and the amount of PHO (X<sub>1</sub>X<sub>3</sub>), and (<b>C</b>) between the amount of HAp and the amount of PHO (X<sub>2</sub>X<sub>3</sub>).</p>
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<p>PHO-HAp-Curcumin composite polymer films.</p>
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<p>Desirability profiles—contour plots (2D) (<b>a</b>,<b>c</b>,<b>e</b>) and response surfaces (3D) (<b>b</b>,<b>d</b>,<b>f</b>).</p>
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<p>SEM images of the surface of the optimal PHO-HAp-Curcumin composite film.</p>
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<p>Appearance of L929 cell culture: (<b>a</b>) control, 24-well plate, (<b>b</b>) extraction method on optimum film, and (<b>c</b>) contact method on optimum film (Inverted microscope, Ob. 10×).</p>
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<p>Preparation scheme of PHO-Hap-Curcumin composite films.</p>
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39 pages, 5551 KiB  
Review
Natural Antioxidants: Advancing Stability and Performance in Sustainable Biobased and Biodegradable Plastics
by Norma Mallegni, Francesca Cicogna, Elisa Passaglia, Vito Gigante, Maria-Beatrice Coltelli and Serena Coiai
Compounds 2025, 5(1), 4; https://doi.org/10.3390/compounds5010004 - 3 Feb 2025
Viewed by 1822
Abstract
The depletion of fossil fuels and environmental concerns have driven the development of sustainable materials, including bio-based and biodegradable plastics, as alternatives to conventional plastics. Although these plastics aid in waste management and climate change mitigation, their vulnerability to oxidative degradation impacts their [...] Read more.
The depletion of fossil fuels and environmental concerns have driven the development of sustainable materials, including bio-based and biodegradable plastics, as alternatives to conventional plastics. Although these plastics aid in waste management and climate change mitigation, their vulnerability to oxidative degradation impacts their longevity, durability, and performance. Natural antioxidants such as tocopherols, flavonoids, and tannins, extracted from plants or agri-food waste, present a sustainable alternative to synthetic stabilizers by enhancing the oxidative thermal stability of polymers like poly(lactic acid) (PLA), poly(butylene succinate) (PBS), poly(butylene succinate-adipate) (PBSA), poly(butylene adipate-co-terephthalate) (PBAT), poly(hydroxyalkanoate) (PHA), and starch-based materials. This review highlights recent advances in bio-based plastics stabilized with natural antioxidants, their mechanisms of action, and their role in improving material properties for applications like packaging. Additionally, it explores their impact on recycling processes, advancements in composite production techniques, and future research directions. Bioplastics can achieve enhanced performance, reduce waste, and support a circular economy by incorporating natural antioxidants. Full article
(This article belongs to the Special Issue Feature Papers in Compounds (2024))
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<p>Radical oxidation mechanism of PLA at temperatures below its melting temperature (upper figure) and three possible β-scission pathways (<b>1</b>, <b>2</b>, and <b>3</b>) of the alkoxy radical (PO°) leading to the final products (lower figure). Reprinted (adapted) with permission from Ref. [<a href="#B36-compounds-05-00004" class="html-bibr">36</a>]. Copyright 2018 Elsevier.</p>
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<p>Radical oxidation process of PLA irradiated at 300 nm under air at 60 °C. Reprinted with permission from Ref. [<a href="#B45-compounds-05-00004" class="html-bibr">45</a>]. Copyright 2010 American Chemical Society.</p>
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<p>Thermal degradation of PBS under an inert atmosphere. Reprinted (adapted) with permission from Ref. [<a href="#B50-compounds-05-00004" class="html-bibr">50</a>]. Copyright 2022 Elsevier.</p>
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<p>Thermal-oxidation mechanism of PBS. Reprinted with permission from Ref. [<a href="#B49-compounds-05-00004" class="html-bibr">49</a>]. Copyright 2018 Elsevier.</p>
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<p>Photooxidation mechanism of PBS. Reprinted with permission from Ref. [<a href="#B56-compounds-05-00004" class="html-bibr">56</a>]. Copyright 2004 American Chemical Society.</p>
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<p>Classification of plant phenolic compounds [<a href="#B69-compounds-05-00004" class="html-bibr">69</a>].</p>
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<p>Structures of benzoic acid (<b>1</b>), common hydroxybenzoic acids (<b>2</b>–<b>5</b>), and common hydroxycinnamic acids (<b>6</b>–<b>10</b>). Reprinted with permission from Ref. [<a href="#B70-compounds-05-00004" class="html-bibr">70</a>]. Copyright 2020 Springer.</p>
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<p>The different categories of food sourced simple phenolic acids. Reprinted with permission from Ref. [<a href="#B71-compounds-05-00004" class="html-bibr">71</a>]. Copyright 2023 Springer.</p>
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<p>Rosmarinic acid.</p>
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<p>Chemical structure of a series of flavonols: myricetin, quercetin, kaempferol, and galangin. Their free radical scavenging ability follows this order: myricetin &gt; quercetin &gt; kaempferol &gt; galangin.</p>
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<p>(<b>a</b>) Weight average molecular weight (MW) of PBS (PBS Proc) and PBS samples containing GPext, T, and Irganox 1010 (PBS GPext, PBS T, and PBS Irg) as a function of the number of processes in a mechanical mixer. (<b>b</b>) MW and (<b>c</b>) MFR of PBS Proc, PBS GPext, PBS T, and PBS Irg samples as a function of oven aging time. Reprinted (adapted) with permission from Ref. [<a href="#B180-compounds-05-00004" class="html-bibr">180</a>]. Copyright 2020 Elsevier.</p>
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<p>Schematic representation of a generic flavonoid subjected to auto-oxidation under aging conditions. Reprinted with permission from Ref. [<a href="#B180-compounds-05-00004" class="html-bibr">180</a>]. Copyright 2020 Elsevier.</p>
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<p>TGA curves of neat PBSA samples subjected to increasing extrusion cycles up to 6 cycles (PX: from P0 to P6) and PBSA blended with 0.5 wt.% LG treated up to 12 cycles (PLGX: from PLG1 to PLG12). The analyses were performed under a nitrogen atmosphere with a heating rate of 1 °C/min. Figure (<b>a</b>) compares neat PBSA (P0) with PBSA treated up to 6 cycles; (<b>b</b>) shows the comparison between P0, P1, and PLG1; (<b>c</b>) shows the comparison after 3 extrusion cycles with and without LG, always compared to neat PBSA; (<b>d</b>) compares the extreme treatments with and without LG, relative to neat PBSA. Reprinted (adapted) with permission from Ref. [<a href="#B182-compounds-05-00004" class="html-bibr">182</a>]. Copyright 2024 Springer.</p>
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<p>(<b>a</b>) Possible radical reactions during thermo-oxidation, (<b>b</b>) thermal degradation via random non-radical chain scission (a widely accepted mechanism), and (<b>c</b>) polyphenols as crosslinking agents within PHB. Reprinted with permission from Ref. [<a href="#B204-compounds-05-00004" class="html-bibr">204</a>]. Copyright 2020 Wiley.</p>
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20 pages, 3739 KiB  
Article
Production and Bioseparation Applications of Polyhydroxyalkanoate Nano-Granules Functionalized with Streptavidin
by Yuyan Zhang, Jiping Zhao, Hui Guo, Xiaoyun Lu and Dan Tan
Microorganisms 2025, 13(2), 312; https://doi.org/10.3390/microorganisms13020312 - 1 Feb 2025
Viewed by 351
Abstract
Rapidly growing industrial biotechnology and bio-manufacturing require simple and cost-effective bioseparation tools. A novel strategy of bioseparation based on the streptavidin-decorated polyhydroxyalkanoate (PHA) nano-granules was developed in this study. By fusing to the N-terminus of PHA-associated phasin protein, the streptavidin was one-step immobilized [...] Read more.
Rapidly growing industrial biotechnology and bio-manufacturing require simple and cost-effective bioseparation tools. A novel strategy of bioseparation based on the streptavidin-decorated polyhydroxyalkanoate (PHA) nano-granules was developed in this study. By fusing to the N-terminus of PHA-associated phasin protein, the streptavidin was one-step immobilized on the surface of PHA nano-granules simultaneously with the accumulation of PHA in recombinant Escherichia coli. About 1.95 g/L of PHA nano-granules (54.51 wt% of cell dry weight) were produced after 48 h bacterial cultivation. The following qualitative and quantitative characterizations demonstrated that the streptavidin accounted for approximately 6.78% of the total weight of the purified PHA nano-granules and confirmed a considerable biotin affinity of 0.1 ng biotin/μg surface protein. As a proof of concept, the nano-granules were further functionalized with biotinylated oligo(dT) for mRNA isolation and about 1.26 μg of mRNA (occupied 2.59%) was purified from 48.45 μg of total RNA, achieving good integrity and high purity with few DNA and rRNA contaminations. Moreover, the nano-granules retained more than 80% of their initial mRNA recovery efficiency after ten cycles of repeated use. The PHA-SAP nano-granules were also functionalized with biotinylated magnetic beads, allowing magnetic recovery of the PHA nano-granules from cell lysates that still needs optimization. Our study provides a novel and expandable platform of PHA nano-granules that can be further functionalized with various biological groups for bioseparation applications. The functional PHA nano-granules have a great potential to serve as bioseparation resin for large-scale purification processes after suitable optimizations for “bench-to-factory” translation, contributing to scalable and sustainable bioprocessing. Full article
(This article belongs to the Section Microbial Biotechnology)
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<p>One-step preparation of streptavidin-decorated PHA nano-granules with their applications in mRNA isolation and magnetic recovery. SA: Streptavidin, PhaP: PHA phasin protein, PhaC: PHA synthetase, PhaA: NADPH-dependent acetyl-CoA acetyltransferase, PhaB: 3-Hydroxybutyryl-CoA dehydrogenase.</p>
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<p>Strain and plasmid construction for the production of PHA-SAP nano-granules. (<b>a</b>) Construction of pET28a-SAP plasmid for the expression of streptavidin-PhaP fusion protein. (<b>b</b>) Construction of recombinant <span class="html-italic">E. coli</span> strain harboring plasmids pET28a-SAP and pCDF-ABC that contained PHA synthetic gene cluster <span class="html-italic">pha</span>CAB. (<b>c</b>) Colony PCR verification of recombinant <span class="html-italic">E. coli</span> harboring two expression vectors. Lane 1S–4S: colony PCR results of <span class="html-italic">sap</span> fragment of four different clones, Lane 1C–4C: colony PCR results of <span class="html-italic">phaC</span> gene of the same four clones with 1S–4S. M: DNA ladder, top to bottom, was 10,000, 7000, 4000, 2000, 1000, 500, 250 (bp). (<b>d</b>) Protein profile of recombinant <span class="html-italic">E. coli</span> harboring two expression vectors after IPTG induction. Lane 1 and Lane 2: Protein profile of recombinant <span class="html-italic">E. coli</span> harboring pET28a-SAP and pCDF-ABC with (Lane 1) and without (Lane 2) IPTG induction. Lane 3 and Lane 4: Protein profile of recombinant <span class="html-italic">E. coli</span> harboring pET28a-SAP with (Lane 3) and without (Lane 4) IPTG induction. Arrows indicate the streptavidin-PhaP fusion protein (lower one) and the PhaC protein (upper one).</p>
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<p>Production and characterization of streptavidin decorated PHA-SAP nano-granules. (<b>a</b>) Fluorescence microscopy of PHA-producing <span class="html-italic">E. coli</span> harboring two expression plasmids at an excitation wavelength of 460 nm after Nile blue A staining. (<b>b</b>) TEM observation of PHA-containing <span class="html-italic">E. coli</span> harboring two expression plasmids, showing conspicuous white PHA nano-granules in the cells. (<b>c</b>) Purified white powder of PHA-SAP nano-granules.</p>
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<p>Qualitative and quantitative characterization of the streptavidin-PhaP fusion protein on the surface of PHA-SAP nano-granules. (<b>a</b>) SDS-PAGE analysis of the protein profile on the surface of the PHA-SAP nano-granules, with streptavidin-PhaP fusion protein being the dominant one (arrow). (<b>b</b>) Qualitative characterization of biotin-binding ability of PHA-SAP nano-granules via Western blot. Lane 1 and Lane 2: two repeated parallel tests of biotin affinity of PHA-SAP nano-granules. Lane 3 and Lane 4: two repeated parallel tests of biotin affinity of PHA blank granules without streptavidin on the surface. (<b>c</b>) Quantitative characterization of biotin affinity with PHA-SAP nano-granules after chromogenic reaction with TMB.</p>
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<p>mRNA isolation by PHA-SAP nano-granules further functionalized with oligo(dT). (<b>a</b>) The scheme of mRNA isolation procedure by PHA-SAP nano-granules. (<b>b</b>) The electrophoresis detection of purified mRNA and other intermediates. Lane 1: Total RNA of C6 cells. Lane 2: The supernatant after precipitation of mRNA with PHA-SAP-oligo(dT) granules (supernatant 0). Lane 3: The supernatant after first washing of precipitated mRNA. Lane 4: The supernatant after the third washing of precipitated mRNA (supernatant 3). Lane 5: Purified mRNA. M: DNA ladder, top to bottom, is 10,000, 7000, 4000, 2000, 1000, 500, and 250 (bp). (<b>c</b>) RT-qPCR amplification curve and Ct values of three genes (GADPH, S100B, and IL-10) using purified mRNA as the template.</p>
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<p>Reusability of PHA-SAP nano-granules. After one cycle of TMB chromogenic reaction (for PHA-SAP-biotin-HRP granules) or mRNA recovery (for PHA-SAP-oligo(dT) granules), the PHA-SAP nano-granules were recycled by centrifugation and subjected to the same procedure and repeated for ten rounds. The initial activity was set to 100%. Error bars indicate SD values from triplicate trials.</p>
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<p>The structures of the magnetic PHA-SAP nano-granules observed by fluorescence microscopy after Nile blue staining and TEM observation. (<b>a</b>,<b>d</b>): Fluorescence microscopy (<b>a</b>) and TEM observation (<b>d</b>) of the blank magnetic beads. (<b>b</b>,<b>e</b>): Fluorescence microscopy (<b>b</b>) and TEM observation. (<b>e</b>) of the complex of the magnetic beads and the PHA blank granules without streptavidin on the surface. (<b>c</b>,<b>f</b>): Fluorescence microscopy (<b>c</b>) and TEM observation (<b>f</b>) of the magnetic PHA-SAP nano-granules.</p>
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18 pages, 2956 KiB  
Article
Does Resveratrol Impact Oxidative Stress Markers in Patients with Head and Neck Cancer Receiving Home Enteral Nutrition?
by Michał Ławiński, Katarzyna Zadka, Natalia Ksepka, Maima Matin, Kamil Wysocki, Dominika Karkocha, Aleksandra Gradowska, Atanas G. Atanasov, Maciej Słodkowski, Agnieszka Wierzbicka and Artur Jóźwik
Nutrients 2025, 17(3), 504; https://doi.org/10.3390/nu17030504 - 30 Jan 2025
Viewed by 951
Abstract
Objectives: Resveratrol (RES) is well documented for its multiple health benefits, with a notable impact on cancer prevention and therapy. This study aimed to evaluate the effect of RES supplementation on oxidative stress in patients with head and neck cancer (HNC) receiving [...] Read more.
Objectives: Resveratrol (RES) is well documented for its multiple health benefits, with a notable impact on cancer prevention and therapy. This study aimed to evaluate the effect of RES supplementation on oxidative stress in patients with head and neck cancer (HNC) receiving home enteral nutrition (HEN). Methods: This randomized, single-center, open-label study involved 72 adult patients, with 40 completing the intervention. Participants in the intervention group received 400 mg of liposomal RES daily for 12 weeks alongside HEN, while the control group received HEN only. Body composition and oxidative stress markers—including total antioxidant capacity (TAC), malondialdehyde (MDA), superoxide dismutase (SOD), glutathione peroxidase (GPx), and reduced glutathione (GSH)—were measured at baseline and after 12 weeks. Results: Significant increases in TAC and SOD activity were observed in both groups. GPx activity increased significantly only in the RES group. MDA levels rose in both groups but were more pronounced in the RES group. GSH levels showed no significant changes. Phase angle (PhA) increased significantly in the RES group, while no significant change was observed in the control group. Conclusions: RES supplementation may enhance antioxidant defenses, as evidenced by increased GPx activity and improvements in TAC and SOD levels, supporting oxidative balance in patients with HNC receiving HEN. The higher MDA levels in the RES group may reflect RES’s dual antioxidant and pro-oxidant activities. Additionally, the observed increase in PhA suggests potential cellular health benefits. These findings highlight the potential of RES as a complementary antioxidant intervention in clinical oncology, warranting further investigation to clarify its therapeutic effects on oxidative stress and cellular health in cancer care. Full article
(This article belongs to the Section Phytochemicals and Human Health)
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<p>The chemical structures of trans isomer of resveratrol.</p>
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<p>Flow chart showing participant dropout. * IC—indirect calorimetry.</p>
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<p>Total antioxidant capacity (TAC) in the resveratrol and control groups measured at baseline and after 12 weeks of intervention. * The statistically significant change within the group: RES group <span class="html-italic">p</span> &lt; 0.001; control group <span class="html-italic">p</span> = 0.001. ** The statistically significant differences between the groups: start <span class="html-italic">p</span> &lt; 0.001; after 12 weeks <span class="html-italic">p</span> &lt; 0.001.</p>
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<p>Reduced malondialdehyde (MDA) in the resveratrol and control groups measured at baseline and after 12 weeks of intervention. * The statistically significant change within the group: RES group <span class="html-italic">p</span> &lt; 0.001; control group <span class="html-italic">p</span> = 0.007. ** The statistically significant differences between the groups: after 12 weeks <span class="html-italic">p</span> = 0.001.</p>
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<p>Superoxide dismutase (SOD) in the resveratrol and control groups measured at baseline and after 12 weeks of intervention. * The statistically significant change within the group: RES group <span class="html-italic">p</span> &lt; 0.001; control group <span class="html-italic">p</span> = 0.001.</p>
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<p>Glutathione peroxidase GPx in the resveratrol and control groups measured at baseline and after 12 weeks of intervention. * The statistically significant change within the group: RES group <span class="html-italic">p</span> = 0.02.</p>
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<p>Reduced glutathione (GSH) in the resveratrol and control groups measured at baseline and after 12 weeks of intervention.</p>
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17 pages, 3402 KiB  
Article
Decoding Cytokine Dynamics: Wharton’s Jelly Stromal Cells and Chondro-Differentiates in PHA-Stimulated Co-Culture
by Raja Sundari Meenakshi Sundaram, Secunda Rupert, Prasanna Srinivasan, Jeswanth Sathyanesan, Kavitha Govarthanan, Naveen Jeyaraman, Swaminathan Ramasubramanian, Madhan Jeyaraman, Ho Yun Chung, Prakash Gangadaran and Byeong-Cheol Ahn
Cells 2025, 14(3), 174; https://doi.org/10.3390/cells14030174 - 23 Jan 2025
Viewed by 709
Abstract
Introduction: Articular cartilage damage presents a significant clinical challenge, with limited options for effective regeneration. Mesenchymal stromal cells (MSCs) derived from Wharton’s jelly (WJ) are a promising cell source for cartilage repair due to their regenerative and immunomodulatory properties. While undifferentiated MSCs have [...] Read more.
Introduction: Articular cartilage damage presents a significant clinical challenge, with limited options for effective regeneration. Mesenchymal stromal cells (MSCs) derived from Wharton’s jelly (WJ) are a promising cell source for cartilage repair due to their regenerative and immunomodulatory properties. While undifferentiated MSCs have demonstrated potent immunoregulatory effects, the immunomodulatory potential of chondrocytes derived from WJ-MSCs remains underexplored, particularly under inflammatory conditions. This study investigates the differential cytokine expression profiles of WJ-MSC-derived chondrocytes and undifferentiated MSCs under inflammatory stimulation with phytohemagglutinin (PHA) to understand their immunomodulatory capacities. Materials and Methods: WJ-MSCs were differentiated into chondrocytes using a micromass culture system. Differentiated chondrocytes were then co-cultured with immune cells under PHA-induced inflammatory conditions. Control groups included co-cultured cells without PHA activation and chondrocytes activated with PHA in the absence of immune cell interaction. Cytokine expression profiles were analyzed using the RT2 Customized Gene Array to evaluate pro- and anti-inflammatory markers. Morphological changes were assessed microscopically. The immunomodulatory responses of chondrocytes were compared to those of undifferentiated MSCs under the same experimental conditions. Results: Chondrocytes co-cultured with immune cells under PHA activation exhibited downregulation of IDO, HLA-G, PDGF, IL-10, TNF-α, IL-6, and IFN-γ compared to undifferentiated MSCs in similar conditions. In non-PHA co-cultured conditions, chondrocytes showed increased expression of IL-6, IFN-γ, IL-4, VEGF, iNOS, PDGF, PTGS-2 and TGF-β, while TNF-α, IL-10, IDO and HLA-G were decreased. In contrast, chondrocytes activated with PHA without immune cell interaction displayed reduced expression of HLA-G and TNF-α, with no significant changes in IL-6, IFN-γ, IL-4, IL-10, VEGF, PDGF, PTGS-2, TGF-β, IDO, and iNOS compared to PHA-stimulated undifferentiated MSCs. Conclusion: This study demonstrates that chondrocytes derived from WJ-MSCs exhibit limited immunomodulatory potential compared to undifferentiated MSCs, particularly under PHA-induced inflammatory conditions. Undifferentiated MSCs showed superior regulation of key cytokines associated with immune modulation. These findings suggest that maintaining MSCs in an undifferentiated state may be advantageous for therapeutic applications targeting inflammatory conditions, such as osteoarthritis. Future research should explore strategies to enhance the immunomodulatory efficacy of chondrocytes, potentially through genetic modification or adjunctive therapies. Full article
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<p>Isolation and morphological characteristics of mesenchymal stromal cells (MSCs) from umbilical cord. (<b>A</b>,<b>B</b>) Isolation of mesenchymal stromal cells from the umbilical cord by explant method and (<b>C</b>) spindle cell formation after removal of explants at day 7. (<b>D</b>) MSCs showing characteristic spindle-shaped morphology under the phase contrast microscope at 10× magnification.</p>
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<p>Immunophenotypic characterization of mesenchymal stromal cells (MSCs) according to International Society for Cellular Therapy (ISCT) criteria. Mesenchymal stromal cells for the positive expression of CD 73, CD 90, and CD 105 (<b>A</b>–<b>C</b>) and lack of expression of CD14, CD34, and CD45 (<b>D</b>–<b>F</b>).</p>
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<p>Characterization of mesenchymal stromal cells by tri-lineage differentiation according to the minimal criteria proposed by ISCT. (<b>A</b>–<b>C</b>) Confirmation of adipogenic differentiation by Oil-Red O staining, osteogenic differentiation by Alizarin Red staining, and chondrogenic differentiation by Alcian Blue staining with their respective controls (<b>A1</b>–<b>C1</b>). All the images were taken at 10× magnification under a phase contrast microscope.</p>
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<p>Relative gene expression levels of chondrocyte-specific markers (SOX9 and COL2A1) and fibroblast-associated marker (COL1A1) in MSCs after chondrogenic differentiation. SOX9 and COL2A1 were significantly upregulated in differentiated cells compared to undifferentiated controls), confirming successful chondrogenic differentiation (**** <span class="html-italic">p</span> &lt; 0.0001). Expression of COL1A1 was negligible, indicating the absence of fibroblast contamination (* <span class="html-italic">p</span> &lt; 0.05). Data are presented as mean ± SEM (<span class="html-italic">n</span> = 3 biological replicates), normalized to GAPDH expression. Statistical significance was determined using multiple <span class="html-italic">t</span>-test analysis.</p>
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<p>Influence of PHA activation on the morphological changes during co-culture of WJ-MSC-derived chondrocytes with MNC. (<b>A</b>) Micrograph of chondrocytes showing the formation of micro-mass pellet after 21 days of chondrogenic differentiation (negative control). (<b>B</b>) Chondrocytes treated with PHA (positive control) showing an increase in size, indicating responsiveness to PHA. (<b>C</b>) Chondrocytes co-cultured with MNCs in the absence of PHA, showing no difference in their morphology. (<b>D</b>) Chondrocytes co-cultured with immune cells in the presence of PHA, showing a decrease in cell size compared to positive control, indicating responsiveness towards PHA. Cell size measured by Image J software (latest v. 1.54).</p>
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<p>(<b>A</b>) Pro-inflammatory Cytokines: Expression levels of IL-6, TNF-α, and IFN-γ,iNOS were compared under three conditions: co-culture of chondrocytes with immune cells under PHA stimulation, co-culture of chondrocytes without PHA stimulation, and PHA stimulation alone. Significant downregulation of IL-6, IFN-γ, and TNF-α was observed in the PHA-activated co-culture condition compared to other conditions (* <span class="html-italic">p</span> &lt; 0.05, ** <span class="html-italic">p</span> &lt; 0.0, *** <span class="html-italic">p</span> &lt; 0.001, and **** <span class="html-italic">p</span> &lt; 0.0001). (<b>B</b>) Anti-inflammatory cytokines: Expression levels of IL-10, TGF-β, and IL-4 were compared under three conditions: co-culture of chondrocytes with immune cells under PHA stimulation, co-culture of chondrocytes without PHA stimulation, and PHA stimulation alone. Significant downregulation in the level of IL-10, with no significant changes in the level of IL-4 and TGF-β was observed in the PHA-activated co-culture condition in differentiated chondrocytes. (<b>C</b>) Immunomodulatory molecules: HLA-G and IDO expression was reduced in differentiated chondrocytes across all conditions compared to undifferentiated MSCs. (<b>D</b>) Growth factors: VEGF and PDGF were significantly upregulated in non-PHA co-cultured conditions. Data were analyzed using GraphPad Prism 10.4 with one-way ANOVA and Sidak’s multiple comparisons test. Values are presented as mean ± SEM.</p>
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<p>(<b>A</b>) Pro-inflammatory Cytokines: Expression levels of IL-6, TNF-α, and IFN-γ,iNOS were compared under three conditions: co-culture of chondrocytes with immune cells under PHA stimulation, co-culture of chondrocytes without PHA stimulation, and PHA stimulation alone. Significant downregulation of IL-6, IFN-γ, and TNF-α was observed in the PHA-activated co-culture condition compared to other conditions (* <span class="html-italic">p</span> &lt; 0.05, ** <span class="html-italic">p</span> &lt; 0.0, *** <span class="html-italic">p</span> &lt; 0.001, and **** <span class="html-italic">p</span> &lt; 0.0001). (<b>B</b>) Anti-inflammatory cytokines: Expression levels of IL-10, TGF-β, and IL-4 were compared under three conditions: co-culture of chondrocytes with immune cells under PHA stimulation, co-culture of chondrocytes without PHA stimulation, and PHA stimulation alone. Significant downregulation in the level of IL-10, with no significant changes in the level of IL-4 and TGF-β was observed in the PHA-activated co-culture condition in differentiated chondrocytes. (<b>C</b>) Immunomodulatory molecules: HLA-G and IDO expression was reduced in differentiated chondrocytes across all conditions compared to undifferentiated MSCs. (<b>D</b>) Growth factors: VEGF and PDGF were significantly upregulated in non-PHA co-cultured conditions. Data were analyzed using GraphPad Prism 10.4 with one-way ANOVA and Sidak’s multiple comparisons test. Values are presented as mean ± SEM.</p>
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<p>Bar graph showing the quantitative intracellular protein levels by solid-phase sandwich ELISA for VEGF during the various culture conditions. VEGF expression was significantly higher in undifferentiated MSCs compared to chondro-differentiated cells in all the culture conditions (* <span class="html-italic">p</span> &lt; 0.05, ** <span class="html-italic">p</span> &lt; 0.01, *** <span class="html-italic">p</span> &lt; 0.001) Data were analyzed using GraphPad Prism 10.4 with one-way ANOVA and Sidak’s multiple comparisons test. Values are presented as mean ± SEM.</p>
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21 pages, 5189 KiB  
Article
Simulation Tool for the Techno-Economic Assessment of the Integrated Production of Polyhydroxyalkanoates as Value-Added Byproducts of a Wastewater Treatment Plant
by Laura Pozo-Morales, Antonio Rosales Martínez, Enrique Baquerizo and Germán del Valle Agulla
Processes 2025, 13(2), 295; https://doi.org/10.3390/pr13020295 - 21 Jan 2025
Viewed by 1318
Abstract
The polyhydroxyalkanoate (PHA) production process that uses mixed microbial cultures combined with main stream wastewater treatment plants (WWTPs) is a competitive integrated resource recovery process in which non-oxygen electron acceptors can be used to enrich the PHA producer. Trials carried out in operating [...] Read more.
The polyhydroxyalkanoate (PHA) production process that uses mixed microbial cultures combined with main stream wastewater treatment plants (WWTPs) is a competitive integrated resource recovery process in which non-oxygen electron acceptors can be used to enrich the PHA producer. Trials carried out in operating plants are very scarce, and there are no simulation tools available to analyse the feasibility of integrating the two processes. This research presents a novel analysis tool for a techno-economic assessment of value-added biopolymers. A general model for a conventional WWTP has been designed and eventually validated using the operating data collected in the database of a fully operational plant. In the model, a simulation of a PHA production line based on thickened primary sludge as a substrate has been integrated. The assembly has been treated as a closed-loop system with an accuracy level of 0.1% with a limit of 1000 iterations. Two strategies based on internal (ADF) or external (AN/AD) limitations of some nutrients have been contrasted for the selection of a biomass capable of feast–famine PHA synthesis. The ADF strategy was found to be the most favourable system, with a production of 0.226 kg of CODPHA·kg−1 COD. The calculated production cost was EUR 0.11·kg−1 CODPHA. The sludge production was reduced by 6%. Full article
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<p>Ground plan of WWTP El Copero.</p>
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<p>Process diagram—ADF process alternative.</p>
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<p>Process diagram—AN/AE process alternative.</p>
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<p>Mass balance at reference scenario.</p>
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<p>Mass balance in ADF process alternative.</p>
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<p>Mass balance in AN/AE process alternative.</p>
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<p>Comparison between both models’ energy consumption overall.</p>
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40 pages, 2483 KiB  
Review
Valorization of Agri-Food Waste into PHA and Bioplastics: From Waste Selection to Transformation
by Marta Arriaga, Francisco Javier Pinar, Irene Izarra, Jesús del Amo, Javier Vicente, Francisco Jesús Fernández-Morales and Javier Mena
Appl. Sci. 2025, 15(3), 1008; https://doi.org/10.3390/app15031008 - 21 Jan 2025
Viewed by 860
Abstract
The valorization of agri-food waste is a pivotal component in developing the circular economy, wherein waste is given a second life through various conversion technologies. This review aims to provide an overview of the current state of knowledge on the valorization of agri-food [...] Read more.
The valorization of agri-food waste is a pivotal component in developing the circular economy, wherein waste is given a second life through various conversion technologies. This review aims to provide an overview of the current state of knowledge on the valorization of agri-food waste, with a particular focus on volatile fatty acids (VFAs) and subsequent bioplastics production. To this end, a comprehensive literature search was conducted using specialist bibliographic databases. The study primarily focuses on reviewing the biological production of PHBV (poly(3-hydroxybutyrate-co-3-hydroxyvalerate)), a type of PHA, due to its relevance as a substitute for fossil-based plastics. Significant attention has been directed toward exploring the potential of agri-food wastes, such as whey, potato peelings, and brewery bagasse, as raw materials for their transformation into a tailor-made VFA solution, rich in valeric and propionic acids. Additionally, existing operational strategies and waste co-treatments have been reviewed. The produced VFAs present multiple applications, including single-cell oils, Omega-3-rich oils, and different types of PHA. Factors influencing the bio-polymerization of VFAs to PHBV, such as the type of substrate, operating conditions (pH and retention time), and the presence of specific microorganisms, have also been reviewed. Due to its physicochemical properties, PHBV has applications in sectors such as agriculture and packaging. This review concludes that transforming agri-food waste into PHBV has the potential to integrate environmental and economic benefits within a circular bioeconomy model, fostering technological innovation and the sustainable use of waste resources. Full article
(This article belongs to the Section Energy Science and Technology)
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<p>Schematic overview of the dark fermentation process for VFA synthesis. Adapted from Pandey et al. [<a href="#B35-applsci-15-01008" class="html-bibr">35</a>], Pilli et al. [<a href="#B36-applsci-15-01008" class="html-bibr">36</a>].</p>
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<p>Effect of organic loading rate on VFA distribution. HAc = acetic acid, HPr = propionic acid, HBu = butiric acid, and HVal = valeric acid. Modified from Calero et al. [<a href="#B77-applsci-15-01008" class="html-bibr">77</a>].</p>
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<p>Schematic overview of the dark fermentation process and production of VFAs that can be consumed by different microorganisms to produce different bioproducts. Adapted from Chalima et al. [<a href="#B32-applsci-15-01008" class="html-bibr">32</a>].</p>
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<p>Scl-PHA production pathway using glucose as the carbon source. PhaC is the PHA synthase enzyme in all pathways and catalyzes the polymerization of monomers to PHA polymers. Modified from Lu et al. [<a href="#B138-applsci-15-01008" class="html-bibr">138</a>].</p>
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<p>Bioplastic polymers, and their monomers, produced by microbial fermentation combined with chemical synthesis. Modified from Chen and Patel [<a href="#B162-applsci-15-01008" class="html-bibr">162</a>].</p>
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