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14 pages, 911 KiB  
Review
Emerging Strategies in Cartilage Repair and Joint Preservation
by Mircea Adrian Focsa, Sorin Florescu and Armand Gogulescu
Medicina 2025, 61(1), 24; https://doi.org/10.3390/medicina61010024 (registering DOI) - 27 Dec 2024
Viewed by 291
Abstract
Background and Objectives: Cartilage repair remains a critical challenge in orthopaedic medicine due to the tissue’s limited self-healing ability, contributing to degenerative joint conditions such as osteoarthritis (OA). In response, regenerative medicine has developed advanced therapeutic strategies, including cell-based therapies, gene editing, and [...] Read more.
Background and Objectives: Cartilage repair remains a critical challenge in orthopaedic medicine due to the tissue’s limited self-healing ability, contributing to degenerative joint conditions such as osteoarthritis (OA). In response, regenerative medicine has developed advanced therapeutic strategies, including cell-based therapies, gene editing, and bioengineered scaffolds, to promote cartilage regeneration and restore joint function. This narrative review aims to explore the latest developments in cartilage repair techniques, focusing on mesenchymal stem cell (MSC) therapy, gene-based interventions, and biomaterial innovations. It also discusses the impact of patient-specific factors, such as age, defect size, and cost efficiency, on treatment selection and outcomes. Materials and Methods: This review synthesises findings from recent clinical and preclinical studies published within the last five years, retrieved from the PubMed, Scopus, and Web of Science databases. The search targeted key terms such as “cartilage repair”, “stem cell therapy”, “gene editing”, “biomaterials”, and “tissue engineering”. Results: Advances in MSC-based therapies, including autologous chondrocyte implantation (ACI) and platelet-rich plasma (PRP), have demonstrated promising regenerative potential. Gene-editing tools like CRISPR/Cas9 have facilitated targeted cellular modifications, while novel biomaterials such as hydrogels, biodegradable scaffolds, and 3D-printed constructs have improved mechanical support and tissue integration. Additionally, biophysical stimuli like low-intensity pulsed ultrasound (LIPUS) and electromagnetic fields (EMFs) have enhanced chondrogenic differentiation and matrix production. Treatment decisions are influenced by patient age, cartilage defect size, and financial considerations, highlighting the need for personalised and multimodal approaches. Conclusions: Combining regenerative techniques, including cell-based therapies, gene modifications, and advanced scaffolding, offers a promising pathway towards durable cartilage repair and joint preservation. Future research should focus on refining integrated therapeutic protocols, conducting long-term clinical evaluations, and embracing personalised treatment models driven by artificial intelligence and predictive algorithms. Full article
(This article belongs to the Section Orthopedics)
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<p>Overview of key regenerative techniques in cartilage repair.</p>
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<p>Poly (lactic-co-glycolic acid) (PLGA) porous scaffold for tissue engineering. Enlargement of a pore. The average pore size is 350–550 mm, and the porosity is estimated at 35–45%. © 2004–2024 University of Cambridge <a href="https://creativecommons.org/licenses/by-nc-sa/4.0/" target="_blank">https://creativecommons.org/licenses/by-nc-sa/4.0/</a> (accessed on 15 December 2024).</p>
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16 pages, 1858 KiB  
Article
Antibacterial Effect of Canine Leucocyte Platelet-Rich Plasma (L-PRP) and Canine Platelet-Poor Plasma (PPP) Against Methicillin-Sensitive and Methicillin-Resistant Staphylococcus pseudintermedius
by Roberta Perego, Gabriele Meroni, Piera Anna Martino, Eva Spada, Luciana Baggiani and Daniela Proverbio
Vet. Sci. 2024, 11(12), 670; https://doi.org/10.3390/vetsci11120670 - 20 Dec 2024
Viewed by 518
Abstract
Staphylococcus pseudintermedius (SP) is a commensal and opportunistic pathogen of skin and mucosal surfaces, isolated from healthy dogs and from canine pyoderma cases. It has recently gained attention due to its increasing antibiotic resistance. Platelet-rich plasma (PRP) is a biological product, obtained through [...] Read more.
Staphylococcus pseudintermedius (SP) is a commensal and opportunistic pathogen of skin and mucosal surfaces, isolated from healthy dogs and from canine pyoderma cases. It has recently gained attention due to its increasing antibiotic resistance. Platelet-rich plasma (PRP) is a biological product, obtained through a blood centrifugation process, which has antibacterial properties evidenced by in vitro and in vivo studies conducted in both the human and veterinary field. This in vitro study evaluated the antimicrobial effect of canine non-activated and activated leucocyte-rich PRP (L-PRP) and platelet-poor plasma (PPP) against two strains of SP isolated from dogs with pyoderma: one a multidrug-resistant strain (MDR) and one a non-MDR strain. Twenty healthy un-sedated adult blood donor dogs were enrolled for L-PRP and PPP production via a closed semi-automatic system for veterinary use. The evaluation of antimicrobial effect was performed using the micro-inhibition in broth method, exposing SP strains to 10 L-PRP, 10 activated L-PRP and 10 PPP samples, respectively. Bacterial growth was evaluated using CFU count at three timepoints (immediately after incubation T0, after 1 h T1 and after 2 h T2). L-PRP and PPP had a significant antimicrobial effect at all three timepoints which was similar against both non-MDR and MDR SP strains. Activation appeared to reduce the duration of the antimicrobial effect in L-PRP. More studies are necessary to confirm these preliminary results. Full article
(This article belongs to the Section Veterinary Internal Medicine)
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<p>Antibacterial effect of L-PRP against non-MDR and MDR SP strains. * statistically significant.</p>
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<p>Antibacterial effect of PPP against non-MDR and MDR SP strains. * statistically significant.</p>
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<p>Effect of animal sex on the antibacterial activity of L-PRP (<b>A</b>,<b>B</b>) and PPP (<b>C</b>,<b>D</b>) against SP strains. * statistically significant.</p>
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<p>Activated L- PRP antibacterial action over time for SP40 (<b>A</b>) and SP67 (<b>B</b>) strains.</p>
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13 pages, 1052 KiB  
Article
The Combined Use of Triamcinolone and Platelet-Rich Plasma in Equine Metacarpophalangeal Joint Osteoarthritis Treatments: An In Vivo and In Vitro Study
by Kübra Guidoni, Elisabetta Chiaradia, Marco Pepe, Antonio Di Meo, Alessia Tognoloni, Matteo Seccaroni and Francesca Beccati
Animals 2024, 14(24), 3645; https://doi.org/10.3390/ani14243645 - 17 Dec 2024
Viewed by 345
Abstract
Intra-articular corticosteroids, such as triamcinolone acetonide (TA), help reduce pain related to osteoarthritis (OA), but they may impair cartilage metabolism. In contrast, platelet-rich plasma (PRP) therapy, a regenerative therapy, has shown potential to promote healing and regeneration of articular cartilage. This study investigates [...] Read more.
Intra-articular corticosteroids, such as triamcinolone acetonide (TA), help reduce pain related to osteoarthritis (OA), but they may impair cartilage metabolism. In contrast, platelet-rich plasma (PRP) therapy, a regenerative therapy, has shown potential to promote healing and regeneration of articular cartilage. This study investigates the effects of combining PRP with TA to treat osteoarthritis in racehorses. The study proposes that PRP injection following TA treatment could reduce side effects and improve treatment outcomes. Firstly, in the in vitro study, chondrocytes were exposed to different TA concentrations, with or without PRP. TA dramatically reduced chondrocyte viability. However, this was prevented by the addition of PRP, which also increased cell proliferation. In the in vivo study, 32 racehorses with metacarpophalangeal (MCP) joint OA were separated into two groups: one received only TA, while the other received TA followed by PRP. For both groups, there were improved flexion assessments one week following the last treatment, but by two weeks following the last treatment, only TA+PRP had improved flexion assessments. TA+PRP also had improved lameness scores two weeks after the last treatment. In conclusion, combining PRP with TA could enhance chondrocyte viability and provide a better long-term therapeutic option for treating OA in racehorses. Further trials are required to thoroughly assess this technique’s safety and efficacy. Full article
(This article belongs to the Section Equids)
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<p>Flowchart of the experimental schedules for horse groups (group TA and group TA+PRP) included in the study.</p>
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<p>Cell viability after the addition of platelet-rich plasma (PRP) to the triamcinolone acetonide (TA) after 24 h at the treatment dose 0, 0.25, 0.50, 1.0, 2.0 and 4.0 mg/mL, as compared with triamcinolone acetonide alone. Data are the mean ± SD of four independent experiments performed in triplicates. ** <span class="html-italic">p</span> &lt; 0.01; *** <span class="html-italic">p</span> &lt; 0.001; **** <span class="html-italic">p</span> &lt; 0.0001; # <span class="html-italic">p</span> &lt; 0.05; ## <span class="html-italic">p</span> &lt; 0.001 vs. control (CTRL).</p>
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<p>Reasons for re-admission to the hospital following the treatments in the TA and TA+PRP groups. The chart demonstrates the frequency of recurring clinical signs observed during re-admission to the hospital, revealing variations in effusion, flexion and lameness complaints between the TA and TA+PRP groups.</p>
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21 pages, 1996 KiB  
Article
Assessment of Extracellular Particles Directly in Diluted Plasma and Blood by Interferometric Light Microscopy. A Study of 613 Human and 163 Canine Samples
by Boštjan Korenjak, Armando Tratenšek, Matevž Arko, Anna Romolo, Matej Hočevar, Matic Kisovec, Maxence Berry, Apolonija Bedina Zavec, David Drobne, Tomaž Vovk, Aleš Iglič, Alenka Nemec Svete, Vladimira Erjavec and Veronika Kralj-Iglič
Cells 2024, 13(24), 2054; https://doi.org/10.3390/cells13242054 (registering DOI) - 12 Dec 2024
Viewed by 701
Abstract
Extracellular nanoparticles (EPs) are a subject of increasing interest for their biological role as mediators in cell–cell communication; however, their harvesting and assessment from bodily fluids are challenging, as processing can significantly affect samples. With the aim of minimizing processing artifacts, we assessed [...] Read more.
Extracellular nanoparticles (EPs) are a subject of increasing interest for their biological role as mediators in cell–cell communication; however, their harvesting and assessment from bodily fluids are challenging, as processing can significantly affect samples. With the aim of minimizing processing artifacts, we assessed the number density (n) and hydrodynamic diameter (Dh) of EPs directly in diluted plasma and blood using the following recently developed technique: interferometric light microscopy (ILM). We analyzed 613 blood and plasma samples from human patients with inflammatory bowel disease (IBD), collected in trisodium citrate and ethylenediaminetetraacetic acid (EDTA) anticoagulants, and 163 blood and plasma samples from canine patients with brachycephalic obstructive airway syndrome (BOAS). We found a highly statistically significant correlation between n in the plasma and n in the blood only in the human (i.e., but not canine) blood samples, between the samples with trisodium citrate and EDTA, and between the respective Dh for both species (all p < 10−3). In the human plasma, the average <Dh> was 139 ± 31 nm; in the human blood, <Dh> was 158 ± 11 nm; in the canine plasma, <Dh> was 155 ± 32 nm; and in the canine blood, <Dh> was 171 ± 33 nm. The differences within species were statistically significant (p < 10−2), with sufficient statistical power (P > 0.8). For <n>, we found no statistically significant differences between the human plasma and blood samples or between the samples with trisodium citrate and EDTA. Our results prove that measuring n and Dh of EPs in minimally processed fresh blood and in diluted fresh plasma by means of ILM is feasible for large populations of samples. Full article
(This article belongs to the Special Issue Advances in Biophysics of Cellular Membranes)
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Graphical abstract

Graphical abstract
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<p>(<b>a</b>) Hydrodynamic diameter of EPs <span class="html-italic">D</span><sub>h</sub> as a function of the number density of EPs <span class="html-italic">n</span> in human plasma; (<b>b</b>) <span class="html-italic">D</span><sub>h</sub> as a function of <span class="html-italic">n</span> in human blood; (<b>c</b>) <span class="html-italic">D</span><sub>h</sub> as a function of <span class="html-italic">n</span> in canine plasma and blood; (<b>d</b>) agreement between <span class="html-italic">D</span><sub>h</sub> of EPs in the samples collected in two anticoagulants (trisodium citrate and EDTA); (<b>e</b>) agreement between <span class="html-italic">D</span><sub>h</sub> of EPs in human plasma and in human blood; (<b>f</b>) agreement between <span class="html-italic">D</span><sub>h</sub> of EPs in canine plasma and in canine blood; (<b>g</b>) agreement between <span class="html-italic">n</span> in samples collected in two anticoagulants (trisodium citrate and EDTA); (<b>h</b>) agreement between <span class="html-italic">n</span> in human plasma and in human blood; (<b>i</b>) agreement between <span class="html-italic">n</span> in canine plasma and in canine blood.</p>
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<p>(<b>a</b>) Scanning electron micrograph of the canine plasma showing EPs (white arrow) and platelets (white triangle); (<b>b</b>) cryogenic transmission electron micrograph of EPs isolated from human plasma showing membrane-enclosed vesicles of various sizes (dashed white arrow points to a large vesicle, and black arrows point to smaller vesicles) and molecular complexes (white arrows); (<b>c</b>) correlation between <span class="html-italic">D</span><sub>h</sub> and the width of the distribution <span class="html-italic">W</span> for all analyzed samples (full circles: plasma; empty circles: blood); (<b>d</b>–<b>g</b>) examples of ILM results of human plasma and blood and of canine plasma and blood showing the distribution over <span class="html-italic">D</span><sub>h</sub> and displaying the detected particles (orange circles).</p>
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15 pages, 3251 KiB  
Article
Potential Use of Plasma Rich in Growth Factors in Age-Related Macular Degeneration: Evidence from a Mouse Model
by Eduardo Anitua, Francisco Muruzabal, Sergio Recalde, Patricia Fernandez-Robredo and Mohammad Hamdan Alkhraisat
Medicina 2024, 60(12), 2036; https://doi.org/10.3390/medicina60122036 - 10 Dec 2024
Viewed by 689
Abstract
Background and Objectives: Age-related macular degeneration (AMD) is the leading cause of low vision and legal blindness in adults in developed countries. Wet AMD can be successfully treated using vascular endothelial growth factor (VEGF) inhibitors; however, dry AMD currently has no effective [...] Read more.
Background and Objectives: Age-related macular degeneration (AMD) is the leading cause of low vision and legal blindness in adults in developed countries. Wet AMD can be successfully treated using vascular endothelial growth factor (VEGF) inhibitors; however, dry AMD currently has no effective treatment. The purpose of this study is to analyze the efficacy of intraocular injection of plasma rich in growth factors (PRGF) in an AMD mouse model induced by intraperitoneal administration of sodium iodate. Materials and Methods: Intravitreal application of PRGF (experimental group) and saline (control group) was performed immediately after intraperitoneal injection of sodium iodate. Retinographies were performed at 2 and 7 days after treatment administration. The eyes were retrieved for histological and immunohistological analysis. Statistical analysis was performed to compare the outcomes between the study groups. Results: In comparison to saline solution, PRGF significantly decreased the depigmentation of the RPE, showing a more reddened retina. PRGF intravitreal treatment significantly reduced the glial fibrillary acidic protein (GFAP) stained processes, suggesting a significant reduction in the risk of scar formation. Moreover, the myofibroblast invasion into the RPE cell layer was significantly reduced in the PRGF-treated group of mice. There was a tendency for better preservation of the photoreceptors in the PRGF group. Conclusions: Within the limitations of this study, intravitreal injection of PRGF provided significant protection against the degeneration of the photoreceptors and the RPE induced by the systemic administration of NaIO3. Full article
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<p>Representative macroscopic images of retinas on the same day of treatment (t0) and after 2 and 7 days of treatment administration from female and male mice of the different groups (control, NaIO<sub>3</sub> -+ SS, and NaIO<sub>3</sub> + PRGF). The control group showed the typical retinal redness throughout the study period, while the disease group (NaIO<sub>3</sub> -+ SS) showed severe depigmentation of the retinal fundus at 2 days after intraperitoneal injection, which recovered at 7 days after treatment, but not to levels similar to the control group. The group of mice treated with PRGF (NaIO<sub>3</sub> + PRGF) showed significantly less retinal depigmentation than the disease group on day 2, with a more reddened retina. At day 7, retinal depigmentation was significantly reduced, showing a white patch throughout the retina, but this patch was smaller than in the disease group (NaIO<sub>3</sub> + SS). Macroscopic images show the different effects of NaIO<sub>3</sub> depending on the sex of the mice, showing a significant reduction effect on the retina of female mice compared to the retina of male mice.</p>
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<p>Representative histological images of female and male mice from the different treatment groups (control, NaIO<sub>3</sub> -+ SS, and NaIO<sub>3</sub> + PRGF) at two different magnifications (2× and 20×). Images at 20× magnification correspond to the black boxed area drawn in the lower magnification (2×) upper image. The histological images show remarkable retinal disorganization in the disease group of mice compared to the control group and the PRGF group, while the retinas of the PRGF-treated mice showed similar histological morphology to the control group.</p>
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<p>Immunohistochemical analysis of GFAP protein expression. The results showed that GFAP expression increased after intraperitoneal administration of NaIO<sub>3</sub> compared with the control group. On the other hand, intravitreal administration of PRGF reduced GFAP expression. However, no significant differences were observed between the different groups. Representative GFAP immunohistochemical images show an increased number of GFAP-positive processes throughout the retina of mice in the disease group (black arrowheads) compared with the control and PRGF groups. Noteworthy differences in GFAP protein expression were observed in relation to the mice’s genders.</p>
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<p>Smooth muscle alpha actin (SMA) expression in the mice retina of the different treatment groups. The squared area in the retina in each image has been enlarged in the upper left corner of the same image. SMA expression was significantly increased in the retinal choriocapillary area in mice in the disease group compared to the control group. PRGF treatment reduced SMA expression in the disease group. On the other hand, a clear difference in SMA expression was observed between female and male retinas.</p>
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<p>Immunohistochemical analysis of rhodopsin in the retina of treated mice. The results showed that rhodopsin was significantly reduced in the retina of mice in which NaIO<sub>3</sub> was administered intraperitoneally compared with the control group. On the other hand, PRGF increased rhodopsin expression with respect to the disease group, though without reaching statistical significance. Rhodopsin immunohistochemical imaging shows that, although the rhodopsin-positive area was markedly higher in the PRGF group than in the disease group, no significant differences were observed between them. The image in the upper left corner of each image corresponds to the magnification of the white-boxed retinal area of the same image. The enlarged image shows that the length of the photoreceptor outer segments, delimited between the black arrow and the white arrow, is significantly shorter in the retina of the disease group compared with the control group and the PRGF group. *: statistically significant (<span class="html-italic">p</span> &lt; 0.05).</p>
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21 pages, 4591 KiB  
Article
Impact of the COL1A1 Gene Polymorphisms on Pain Perception in Tennis Elbow Patients: A Two-Year Prospective Cohort Study
by Paweł Niemiec, Alicja Jarosz, Tomasz Nowak, Anna Balcerzyk-Matić, Tomasz Iwanicki, Joanna Iwanicka, Katarzyna Gawron, Marcin Kalita, Sylwia Górczyńska-Kosiorz, Wojciech Kania and Karol Szyluk
Int. J. Mol. Sci. 2024, 25(23), 13221; https://doi.org/10.3390/ijms252313221 - 9 Dec 2024
Viewed by 585
Abstract
The COL1A1 gene encodes the α1 chain of type I collagen, and the data reported so far demonstrate that its polymorphic variants may affect biomechanical properties of bones, muscles, and tendons, and contribute to musculoskeletal disorders. Given, however, limited research on these variants [...] Read more.
The COL1A1 gene encodes the α1 chain of type I collagen, and the data reported so far demonstrate that its polymorphic variants may affect biomechanical properties of bones, muscles, and tendons, and contribute to musculoskeletal disorders. Given, however, limited research on these variants in tendon pathology, we analyzed the impact of COL1A1 polymorphisms on the tendinopathy phenotype and the effectiveness of platelet-rich plasma (PRP) treatment for tennis elbow. Pain perception and therapy outcomes were analyzed from baseline, i.e., before PRP injection to two years post-PRP injection in a cohort of 107 patients. The study focused on seven COL1A1 variants: rs2249492 (C/T), rs2586488 (A/G), rs2075558 (A/C), rs2253369 (C/T), rs35231764 (A/G), rs1800012 (C/A), and rs9898186 (C/T). We demonstrated that carriers of the TT/CT (rs2249492), AA/AC (rs1800012), and TT/CT (rs9898186) genotypes reported pain related to injury more frequently than subjects with other COL1A1 variants, also in the context of performing specific activities and other pain characteristics. These polymorphisms did not significantly influence therapy effectiveness, although rs35231764 showed a moderate effect. In conclusion, the T (rs2249492), A (rs1800012), and T (rs9898186) alleles of COL1A1 gene are risk factors for pain perception in tennis elbow patients, but do not appear to substantially impact PRP treatment outcomes. Full article
(This article belongs to the Section Molecular Genetics and Genomics)
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<p>Crucial processes observed in tendinopathy (based on two figures from Servier Medical Art resource [<a href="#B13-ijms-25-13221" class="html-bibr">13</a>]: Tendon anatomy and Tendonitis, both licensed under CC BY 4.0., modified by P. Niemiec). Legend: ECM, extracellular matrix; MMPs, matrix metalloproteinases; VEGF, vascular endothelial growth factor.</p>
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<p>Location of the <span class="html-italic">COL1A1</span> gene single nucleotide polymorphisms (SNPs). The figure was created on the basis of data from LDmatrix Tool [<a href="#B50-ijms-25-13221" class="html-bibr">50</a>]. Legend: GRCh38, Genome Reference Consortium Human Build 38 Organism: <span class="html-italic">Homo sapiens</span>; SNPs, single nucleotide polymorphisms.</p>
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<p>Haplotype analysis of the <span class="html-italic">COL1A1</span> gene polymorphisms in the study group (<b>A</b>) and CEU population (<b>B</b>). The darker the color, the higher the D′ or R<sup>2</sup> values.</p>
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<p><span class="html-italic">COL1A1</span> gene expression in skeletal muscle tissue, depending on the genotypes of the rs2586488 and rs2253369 polymorphisms. Based on the data from GTEx Portal [<a href="#B51-ijms-25-13221" class="html-bibr">51</a>].</p>
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<p>Medians (±QD) of VAS and QDASH values in respect to genotype variants of the <span class="html-italic">COL1A1</span> gene rs35231764 polymorphism (recessive–dominant model). Legend: QD, quartile deviation; VAS, visual analog scale; QDASH, quick version of disabilities of the arm, shoulder and hand score; *, differences remaining significant after Hochberg correction for multiple comparisons (threshold of significance for recessive–dominant model: <span class="html-italic">p</span> ≤ 0.009).</p>
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<p>Achievement of minimal clinically important difference for VAS, QDASH and PRTEE in the context of respective polymorphic variants of the <span class="html-italic">COL1A1</span> gene: rs2249492 (<b>A</b>), rs2586488 (<b>B</b>), rs2075558 (<b>C</b>), rs2253369 (<b>D</b>), rs35231764 (<b>E</b>), and rs1800012 (<b>F</b>). Legend: PRTEE, Patient-Rated Tennis Elbow Evaluation; QD, Quartile Deviation; QDASH, quick version of Disabilities of the Arm, Shoulder and Hand score; VAS, Visual Analog Scale; *, differences remaining significant after Hochberg correction for multiple comparisons (threshold of significance for MCID analysis: <span class="html-italic">p</span> ≤ 0.022); §, differences significant (<span class="html-italic">p</span> &lt; 0.050) in multivariate logistic regression analysis (adjusted for all SNPs, age, sex, comorbidities and additional forms of therapy during follow-up).</p>
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<p>Pathophysiological mechanism for the adverse effects of the A allele (rs1800012) on the musculoskeletal system. Based on [<a href="#B25-ijms-25-13221" class="html-bibr">25</a>,<a href="#B37-ijms-25-13221" class="html-bibr">37</a>,<a href="#B38-ijms-25-13221" class="html-bibr">38</a>,<a href="#B39-ijms-25-13221" class="html-bibr">39</a>,<a href="#B40-ijms-25-13221" class="html-bibr">40</a>,<a href="#B41-ijms-25-13221" class="html-bibr">41</a>,<a href="#B42-ijms-25-13221" class="html-bibr">42</a>,<a href="#B43-ijms-25-13221" class="html-bibr">43</a>,<a href="#B52-ijms-25-13221" class="html-bibr">52</a>,<a href="#B55-ijms-25-13221" class="html-bibr">55</a>,<a href="#B56-ijms-25-13221" class="html-bibr">56</a>,<a href="#B57-ijms-25-13221" class="html-bibr">57</a>,<a href="#B58-ijms-25-13221" class="html-bibr">58</a>]. Description in the text.</p>
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<p>Flowchart of the study selection.</p>
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15 pages, 2847 KiB  
Article
The VEGFB Gene Variants and the Effectiveness of Platelet-Rich Plasma Treatment of Lateral Elbow Tendinopathy: A Prospective Cohort Study with a Two-Year Follow-Up
by Alicja Jarosz, Tomasz Nowak, Karol Szyluk, Anna Balcerzyk-Matić, Tomasz Iwanicki, Joanna Iwanicka, Marcin Kalita, Katarzyna Gawron, Wojciech Kania and Paweł Niemiec
Int. J. Mol. Sci. 2024, 25(23), 13166; https://doi.org/10.3390/ijms252313166 - 7 Dec 2024
Viewed by 645
Abstract
Platelet-rich plasma (PRP) is an autologous preparation used to accelerate regeneration; however, this form of therapy is not always effective. Vascular endothelial growth factor B (VEGFB), which affects vessel survival, pathological angiogenesis, and muscle development may differentiate the risk and treatment [...] Read more.
Platelet-rich plasma (PRP) is an autologous preparation used to accelerate regeneration; however, this form of therapy is not always effective. Vascular endothelial growth factor B (VEGFB), which affects vessel survival, pathological angiogenesis, and muscle development may differentiate the risk and treatment of lateral elbow tendinopathy (LET). In this study, we analyzed the influence of VEGFB gene polymorphisms on the effectiveness of LET treatment with PRP. Therapeutic effectiveness was analyzed in 107 patients (132 elbows) using patient-reported outcome measures (PROMs), specifically the visual analog scale (VAS); quick version of disabilities of the arm, shoulder, and hand score (QDASH); and patient-rated tennis elbow evaluation (PRTEE), for two years (weeks 2, 4, 8, 12, 24, 52, and 104). The polymorphisms selected for the study were rs72922019, rs12366035, rs4930152, rs594942, and rs595880, being in strong linkage disequilibrium. Patients with TT (rs72922019), TT (rs12366035), AA (rs4930152), CC (rs594942), and GG (rs595880) genotypes showed better treatment effectiveness. Statistically important differences were shown for rs72922019 VAS (week 2), QDASH (weeks 0–4), and PRTEE (week 2); rs12366035 and rs4930152 VAS (week 2), QDASH (week 2), and PRTEE (weeks 2 and 4); and rs594942 and rs595880 VAS (weeks 2 and 4), QDASH (week 2), and PRTEE (weeks 2, 52, and 104). The studied polymorphisms also showed an association with blood morphological parameters, including mean platelet volume, platelet distribution width, and eosinophil levels, as well as some comorbidities (heart failure). Genotyping due to patient selection for therapy may be considered for any of the rs72922019, rs12366035, or rs4930152 polymorphisms. Full article
(This article belongs to the Section Molecular Pathology, Diagnostics, and Therapeutics)
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<p>Growth factors from the VEGF family and their receptors. Selected growth factors bind to specific receptors and coreceptors. Binding to a particular receptor leads to a different biological effect [<a href="#B1-ijms-25-13166" class="html-bibr">1</a>,<a href="#B2-ijms-25-13166" class="html-bibr">2</a>,<a href="#B3-ijms-25-13166" class="html-bibr">3</a>,<a href="#B4-ijms-25-13166" class="html-bibr">4</a>,<a href="#B5-ijms-25-13166" class="html-bibr">5</a>,<a href="#B6-ijms-25-13166" class="html-bibr">6</a>]. A description is in the text. Legend: IG domain, immunoglobulin domain; TK domain, tyrosine kinase domain; VEGF, vascular endothelial growth factor; PlGF, placenta growth factor; VEGFR, vascular endothelial growth factor receptor; NRP, neuropilin.</p>
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<p>Locations of the studied <span class="html-italic">VEGFB</span> polymorphisms. The figure was created using data from the LDmatrix tool [<a href="#B24-ijms-25-13166" class="html-bibr">24</a>].</p>
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<p>Linkage analyses between studied polymorphisms. (<b>A</b>) Haplotype block with D’ value. (<b>B</b>) Haplotype block with R<sup>2</sup> value. (<b>C</b>) Haplotypes and their frequency. Colors highlight the degree of linkage disequilibrium. The darker the color, the greater the D’ or R2 values.</p>
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<p>Medians (±QD) of PROM values for carriers of different rs12366035 and rs4930152 genotypes. Results for (<b>A</b>) VAS, (<b>B</b>) QDASH, and (<b>C</b>) PRTEE. Legend: QD, quartile deviation; PROM, patient-reported outcome measure; VAS, visual analog scale; QDASH, quick version of disabilities of the arm, shoulder, and hand score; PRTEE, patient-rated tennis elbow evaluation; *, significant difference (<span class="html-italic">p</span> &lt; 0.050).</p>
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<p>In silico analysis of rs72922019 <span class="html-italic">VEGFB</span> gene polymorphism influence on expression level. Results for (<b>A</b>) skeletal muscle tissue and (<b>B</b>) whole blood. Based on GTEx Portal [<a href="#B24-ijms-25-13166" class="html-bibr">24</a>].</p>
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<p>Flow chart presenting selection of studied group.</p>
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12 pages, 4430 KiB  
Article
Pulpal Responses to Leukocyte- and Platelet-Rich Plasma Treatment in Mouse Models for Immediate and Intentionally Delayed Tooth Replantation
by Angela Quispe-Salcedo, Kiyoko Suzuki-Barrera, Mauricio Zapata-Sifuentes, Taisuke Watanabe, Tomoyuki Kawase and Hayato Ohshima
Appl. Sci. 2024, 14(23), 11358; https://doi.org/10.3390/app142311358 - 5 Dec 2024
Viewed by 551
Abstract
This study aimed to evaluate the effect of leukocyte and platelet-rich plasma (L-PRP) on the pulpal healing process following immediate and intentionally delayed tooth replantation in mice. After the maxillary first molars of 3-week-old mice were extracted, the teeth were immersed for 1 [...] Read more.
This study aimed to evaluate the effect of leukocyte and platelet-rich plasma (L-PRP) on the pulpal healing process following immediate and intentionally delayed tooth replantation in mice. After the maxillary first molars of 3-week-old mice were extracted, the teeth were immersed for 1 min [immediate reimplantation (IR)] or 30 min [intentionally delayed reimplantation (IDR)] in phosphate-buffered saline (PBS) solution. The alveolar socket was filled with or without 1.5 μL of L-PRP [experimental or control groups (EG or CG)] followed by tooth replantation. Samples were collected from day 1 to week 4 after the operation, processed for histology, and evaluated by immunohistochemistry for Nestin and Ki-67 expression. Quantitative analysis revealed positive Nestin staining during pulpal healing in the EG at week 1 following IR and week 2 following IDR. Hard tissue deposition was significantly increased in the EG after IR at week 2. Cell proliferation was higher in the EG compared with that in the CG at week 1 and significantly decreased in the coronal pulp of the EG after the IDR at week 2. Our data suggest that treatment with L-PRP may have a positive effect on pulpal healing, even in teeth replanted after an extended extra-oral period. Full article
(This article belongs to the Special Issue Advances in Dental Materials and Their Applications)
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<p>Hematoxylin–eosin (H&amp;E) (<b>A</b>,<b>D</b>,<b>G</b>,<b>J</b>) and AZAN (<b>B</b>,<b>E</b>,<b>H</b>,<b>K</b>) staining, and immunohistochemistry for Nestin (<b>C</b>,<b>F</b>,<b>I</b>,<b>L</b>) in samples from the CG (<b>A</b>–<b>C</b>,<b>G</b>–<b>I</b>) and EG (<b>D</b>–<b>F</b>,<b>J</b>–<b>L</b>) one week after IR (<b>A</b>–<b>F</b>) and IDR (<b>G</b>–<b>L</b>). (<b>A</b>,<b>G</b>) The dental pulp of the CG after IR and IDR shows eosinophilic amorphous matrices and small hemorrhagic areas in the odontoblast layer (arrowheads in <b>G</b>). (<b>D</b>,<b>J</b>) Fewer inflammatory lesions and more blood vessels are observed in the dental pulp of samples from the EG after IR and IDR. (<b>J</b>,<b>K</b>) Some samples show early deposition of newly formed hard tissue (arrows) in certain areas of the pulpal floor after IDR. (<b>B</b>,<b>E</b>) Collagen-related blue areas are consistently detected by AZAN staining in the CG and EG after IR. (<b>H</b>,<b>K</b>) Weak AZAN staining is observed in the dental pulp of samples from the CG and EG after IDR. (<b>C</b>,<b>F</b>) Samples from the CG and EG show a positive reaction for Nestin in odontoblast-like cells, pulpal cells, and filamentous structures after IR. (<b>I</b>,<b>L</b>) Nestin immunoreaction is more intensely detected in the whole dental pulp in samples from the CG and EG after IDR. DP: Dental pulp. Scale bars: 100 µm (<b>A</b>–<b>L</b>).</p>
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<p>H&amp;E (<b>A</b>,<b>D</b>,<b>G</b>,<b>J</b>) and AZAN staining (<b>B</b>,<b>E</b>,<b>H</b>,<b>K</b>), and immunohistochemistry for Nestin (<b>C</b>,<b>F</b>,<b>I</b>,<b>L</b>) in samples from the CG (<b>A</b>–<b>C</b>,<b>G</b>–<b>I</b>) and EG (<b>D</b>–<b>F</b>,<b>J</b>–<b>L</b>) at week 2 after IR (<b>A</b>–<b>F</b>) and IDR (<b>G</b>–<b>L</b>). (<b>A</b>,<b>D</b>,<b>G</b>,<b>J</b>) Newly formed hard tissue is observed in the dental pulp of the CG and EG after IR and IDR. (<b>B</b>,<b>H</b>) Collagen-related blue areas in the hard tissue areas are evident following AZAN staining in samples from the CG. (<b>E</b>,<b>K</b>) AZAN-positive areas are more abundant in the pulpal tissue of the EG following IR and IDR. (<b>C</b>,<b>F</b>,<b>I</b>,<b>L</b>) Nestin expression is localized in the odontoblast-like cells aligned beneath the newly formed tertiary dentin in samples from the CG and EG after IR and IDR. (<b>F</b>,<b>I</b>) Bone-like tissue formation in some samples of the CG and EG lacks Nestin immunoreactivity following either IR or IDR. B: Bone-like tissue, DP: Dental pulp, TD: Tertiary dentin. Scale bars: 100 µm (<b>A</b>–<b>L</b>).</p>
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<p>Quantitative analysis of H&amp;E staining and immunohistochemistry for Nestin in the control and L-PRP teeth after IR and IDR. (<b>A</b>) Significant differences (<span class="html-italic">p</span> &lt; 0.002, <span class="html-italic">p</span> &lt; 0.012) in the percentage of Nestin-positive perimeter are evident between observation points in the CG and EG after IR. (<b>B</b>) No significant differences are noted between groups or observation periods at week 2 after IDR. (<b>C</b>) The occurrence rate of hard tissue deposition in the dental pulp differed significantly between the CG and EG after IR (<span class="html-italic">p</span> &lt; 0.002). (<b>D</b>) A slight positive trend in the EG was noted at this stage following IDR.</p>
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<p>The percentage of Ki-67-positive cells in the crown (<b>A</b>,<b>C</b>,<b>E</b>,<b>G</b>) and root pulp (<b>B</b>,<b>D</b>,<b>F</b>,<b>H</b>) at weeks 1 (<b>A</b>,<b>B</b>,<b>E</b>,<b>F</b>) and 2 (<b>C</b>,<b>D</b>,<b>G</b>,<b>H</b>) after IR (<b>A</b>–<b>D</b>) and IDR (<b>E</b>–<b>H</b>). (<b>A</b>,<b>B</b>) There are no significant differences in the number of proliferating cells at week 1 in the crown and root pulp between CG and EG after IR. (<b>C</b>,<b>D</b>) Similarly, no significant differences are noted between groups at week 2 after IR. (<b>E</b>,<b>F</b>) Cell proliferation appears slightly higher in the CG in the coronal and root pulp at week 1 after IDR. (<b>G</b>,<b>H</b>) The proliferative cells further decrease at week 2, whereas a significant difference between the CG and EG is observed in the coronal pulp at this stage (<span class="html-italic">p</span> &lt; 0.047).</p>
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11 pages, 1566 KiB  
Article
The Applicability and Limitations of the Spectrofluorometric Method for Determination of ALDH1 Activity in Serum and Plasma
by Sylwia Michorowska, Agnieszka Wiśniewska, Renata Wolinowska, Piotr Wroczyński and Joanna Giebułtowicz
Diagnostics 2024, 14(23), 2721; https://doi.org/10.3390/diagnostics14232721 - 3 Dec 2024
Viewed by 530
Abstract
Background: Aldehyde dehydrogenase class 1 (ALDH1) is an enzyme that is ubiquitously distributed in adult tissues and may serve as a prognostic marker in various cancer types. In blood, 99% of ALDH1 is found in erythrocytes; although, it was also demonstrated that leukocytes [...] Read more.
Background: Aldehyde dehydrogenase class 1 (ALDH1) is an enzyme that is ubiquitously distributed in adult tissues and may serve as a prognostic marker in various cancer types. In blood, 99% of ALDH1 is found in erythrocytes; although, it was also demonstrated that leukocytes and platelets exhibit ALDH activity. No ALDH activity was detected in plasma, even when employing the highly sensitive fluorometric method with 7-methoxy-1-naphthaldehyde as a substrate. However, some reports have been released describing stable and measurable ALDH1 activity in the serum of healthy subjects using 6-methoxy-2-naphthaldehyde as a substrate and a Shimadzu RF—5301 spectrofluorometer. Methods: Our study aimed to verify whether ALDH1 activity can be measured in plasma or serum (n = 80) using 6-methoxy-2-naphthaldehyde as a substrate and a highly sensitive Hitachi F7000 spectrofluorometer, which offers a higher signal-to-noise ratio compared to the Shimadzu RF-5301. Additionally, HPLC with fluorometric detection was used to validate the results (n = 25) and analyze the influence of hemolysis (n = 5) and liver cell damage (n = 15) on ALDH1 activity in serum. Results: Measurable ALDH activity in serum/plasma was very rarely detected using a spectrofluorometer (2 cases out of 80). However, background drift in assays without coenzyme addition was observed, and it may be easily mistaken for ALDH or oxidase activity. Therefore, the spectrofluorometer drift observed in blank assays and modified by a matrix, e.g., enhanced in protein-rich samples, should be considered in ALDH1 activity assays. Conclusions: The spectrofluorometric method has limited applicability for determining ALDH activity in plasma and serum. HPLC can measure ALDH1 activity in plasma or serum; however, factors like hemolysis and elevated liver enzymes significantly affect activity and must be considered in diagnostic interpretations. To enhance research quality on ALDH1 as a biomarker for diseases, including cancers, we recommend using control samples, reference materials, and purifying commercially available aldehyde substrates to improve method sensitivity. Full article
(This article belongs to the Section Clinical Laboratory Medicine)
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<p>Kinetics of fluorescence increase in plasma (<b>a</b>,<b>b</b>) and serum (<b>c</b>,<b>d</b>) samples from two exemplar patients, with measurements taken both with (green circles) and without (red triangles) NAD<sup>+</sup> addition. One of the two cases in which measurable ALDH activity was found in both plasma (<b>a</b>) and serum (<b>c</b>) is illustrated. The mean slopes and their standard deviations are as follows: (<b>a</b>) (19.2 ± 0.1) × 10<sup>3</sup> FU/s with NAD<sup>+</sup> and (1.7 ± 0.6) × 10<sup>3</sup> FU/s without NAD<sup>+</sup>; (<b>b</b>) (0.47 ± 0.07) × 10<sup>3</sup> FU/s with NAD<sup>+</sup> and (0.49 ± 0.06) × 10<sup>3</sup> FU/s without NAD<sup>+</sup>; (<b>c</b>) (13.7 ± 0.7) x10<sup>3</sup> FU/s with NAD<sup>+</sup> and (1.8 ± 0.1) × 10<sup>3</sup> FU/s without NAD<sup>+</sup>; (<b>d</b>) (0.62 ± 0.09) × 10<sup>3</sup> FU/s with NAD<sup>+</sup> and (0.677 ± 0.003) × 10<sup>3</sup> FU/s without NAD<sup>+</sup>.</p>
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<p>An exemplar chromatogram is presented for the serum sample, which demonstrated the highest measured ALDH activity, yet was still classified as having immeasurable activity by a spectrofluorometer. The peak shown corresponds to MONCO. The black line represents the sample at t = 0, the grey line represents the sample after 30 min of reaction without NAD<sup>+</sup>, and the dashed line represents the sample with NAD<sup>+</sup> after 30 min of reaction. It is noteworthy that a prominent peak is observable even at time zero, likely attributed to the low oxidative stability of the aldehyde during storage. The marginal increase in peak area observed in the absence of NAD<sup>+</sup> can be attributed to the low activity of other enzymes that do not require NAD<sup>+</sup> as a cofactor, such as aldehyde oxidase.</p>
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<p>Expression and purification of the recombinant ALDH1A1 protein analyzed by SDS-PAGE (polyacrylamide gel electrophoresis) with Coomassie staining. Recombinant protein, with a molecular weight of 55 kD, was expressed as a fusion protein. Lane ‘marker’: pre-stained molecular weight markers (PageRuler, Fermentas, Vilnius, Lithuania) with the following masses: 17, 26, 34, 43, 55, 72, 95, 130, and 170 kDa; lane ‘no induction’: extracts from bacteria containing pET-28a-ALDH1A1 before IPTG induction; lane ‘induction’: extracts from bacteria containing pET-28a-ALDH1A1 after IPTG induction; lane ‘k1’: pellet from centrifugation of bacterial cell lysate after IPTG induction, before purification with His-binding columns; lane ‘k2’: flowthrough from the bacteria extract during purification with the His-binding columns; lanes E1 and E2: purified recombinant protein following purification with His-binding columns.</p>
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<p>The course of fluorescence increase obtained for the recombinant protein, with an activity of 50 mU/L. The mean slopes (and their standard deviations) are as follows: (0.60 ± 0.10) × 10<sup>3</sup> FU/s without NAD<sup>+</sup>, (0.47 ± 0.03) × 10<sup>3</sup> FU/s with 50 nM NAD<sup>+</sup>, and (1.1 ± 0.2) × 10<sup>3</sup> FU/s with 100 μM NAD<sup>+</sup>.</p>
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11 pages, 2911 KiB  
Article
Arthrocentesis of Temporomandibular Joints—A Clinical Comparative Study
by Marta Siewert, Rafał Pokrowiecki, Paweł J. Zawadzki and Zygmunt Stopa
Life 2024, 14(12), 1594; https://doi.org/10.3390/life14121594 - 3 Dec 2024
Viewed by 425
Abstract
The objective of this study was to compare single-needle arthrocentesis with the conventional two-needle arthrocentesis, as well as the additional intracapsular injection of hyaluronic acid or platelet-rich fibrin. A total of 96 patients with established osteoarthritis (OA) (n = 48) or with [...] Read more.
The objective of this study was to compare single-needle arthrocentesis with the conventional two-needle arthrocentesis, as well as the additional intracapsular injection of hyaluronic acid or platelet-rich fibrin. A total of 96 patients with established osteoarthritis (OA) (n = 48) or with internal de-arrangement (DD) (n = 48) were assigned single-needle arthrocentesis with distension of the joint or conventional two-needle arthrocentesis with or without intracapsular injection of the medication (hyaluronic acid (HA) or platelet-rich plasma (PRP)) performed every month over a period of 6 months. The maximum mouth opening and pain, as measured by the visual analog scale (VAS), were compared. Each group exhibited significant improvement, i.e., a decrease in pain and an increase in mouth opening. The single-puncture technique provided similar pain reduction as the two-needle approach but provided significantly better results in terms of maximum mouth opening. The reduction in pain was similar when comparing the OA and DD cohorts; however, patients with disc displacement achieved significantly better mouth opening than OA. Intracapsular application of medication contributed to a significant decrease in pain in both HA and PRP groups, with platelet-rich fibrin being significantly superior to HA in terms of mouth opening improvement. Full article
(This article belongs to the Section Medical Research)
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<p>Intraoperative photographs showing two arthrocentesis techniques: (<b>A</b>) a two-needle approach where fluid is introduced into the TMJ through a posterior entry point and evacuated simultaneously through an anterior exit point, which provides passive joint lavage; (<b>B</b>,<b>C</b>) a single puncture technique where fluid is pumped into the TMJ through a posterior point and then evacuated through the same needle after removal of the syringe, which provides a pressured lavage through joint capsule hydro-dissection and stretching; (<b>D</b>) intracapsular application of medication after arthrocentesis (HA or PRF).</p>
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<p>Change in mean VAS and MMO scores (N = 96)<b>.</b> A one-way analysis of variance (ANOVA) with repeated measures showed a significant difference between the variables: F = 205.36, <span class="html-italic">p</span>-value ≤ 0.001 for the VAS decrease and F = 91.54, <span class="html-italic">p</span>-value ≤ 0.001 for increase in MMO before and after the treatment.</p>
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<p>Graph showing gradual decrease in pain between each treatment in the OA and DD cohorts.</p>
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<p>Graph showing gradual increase in maximal mouth opening between each treatment in the OA and DD cohorts.</p>
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<p>Graph showing gradual decrease in pain between each treatment in the HA and PRP cohorts.</p>
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<p>Graph showing gradual increase in maximal mouth opening between each treatment in the HA and PRP cohorts.</p>
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17 pages, 7904 KiB  
Article
An Oxygenated Layer Coating to Enhance the Healing Process of Non-Healing Wounds—A Case Report
by Tomasz Miłek, Anna Grzeczkowicz, Agata Lipko, Maria Mesjasz, Leszek Oklesiński, Angelika Kwiatkowska, Marcin Strawski, Monika Drabik and Ludomira H. Granicka
Appl. Sci. 2024, 14(23), 11200; https://doi.org/10.3390/app142311200 - 1 Dec 2024
Viewed by 506
Abstract
The ever-increasing problem of chronic, non-healing wound treatment is a challenge in contemporary medicine. This study aims to verify the effect of a designed dressing containing a coating involving oxygenating factors on the promotion of wound healing in all healing phases in a [...] Read more.
The ever-increasing problem of chronic, non-healing wound treatment is a challenge in contemporary medicine. This study aims to verify the effect of a designed dressing containing a coating involving oxygenating factors on the promotion of wound healing in all healing phases in a single clinical case. An alginate-based coating, which incorporates the oxygenating factor, was produced. The obtained surface physicochemical features were examined using FTIR and AFM. The bandage support was modified with the developed coating and applied to patients who suffered from chronic wounds (treated with non-modified commercially available bandages) and had required treatment at least 3 months before starting the study. During bandage application, the chosen characteristics of the wound-healing process were examined. A histological evaluation of granulation tissue confirmed the presence of fibroblasts, hemorrhage, and lymphocytes during the second week of treatment. Moreover, epithelization tissue in the third week of treatment exhibited the presence of keratinocytes. A 100% wound reduction was observed between 3 and 4 weeks of treatment with the modified dressing. It can be recommended that the applied coating, which provides a supportive effect through oxygenating elements, inducing the remodeling phase in up to 4 weeks of treatment, be developed for use in hard-to-heal wound treatment. Full article
(This article belongs to the Section Biomedical Engineering)
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<p>The coating (ALG—PFC)<sub>net</sub>—patient platelet-rich plasma interface model in configuration with treated tissue. Key to the symbol: (ALG—PFC)<sub>net</sub>—alginate–perfluorooctyl cross-linked.</p>
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<p>The package modified with (ALG–PFC)<sub>net</sub> coating dressings. Key to the symbol: (ALG—PFC)<sub>net</sub>—alginate–perfluorooctyl cross-linked.</p>
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<p>AFM visualization of (ALG) and (ALG-PFC) layers on the gold-covered mica substrate. (<b>A</b>) ALG; (<b>B</b>) ALG-PFC. Key to the symbols: ALG—alginate, ALG-PFC—alginate with oxygenating factor.</p>
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<p>AFM visualization of the profile of the ALG-PFC layer deposited on the gold mica substrate cover. Key to the symbol: ALG-PFC—alginate with oxygenating factor.</p>
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<p>FT-IR spectra of ALG, ALG-PFC. Key to the symbols: ALG—alginate; ALG-PFC—alginate–perfluorooctyl.</p>
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<p>SEM visualization of the surface of (ALG-PFC)<sub>net</sub> coating. (<b>A</b>) magnification 100; (<b>B</b>) magnification 300. Key to the symbol: (ALG-PFC)<sub>net</sub>—alginate–perfluorooctyl cross-linked.</p>
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<p>Visualization of the surface of the (<b>A</b>) ALGnet coating; (<b>B</b>) (ALG-PFC)<sub>net</sub> coating. Key to the symbols: ALG<sub>net</sub>—alginate cross-linked; (ALG-PFC)<sub>net</sub>—alginate–perfluorooctyl cross-linked.</p>
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<p>Fluorescence microscopic picture of (ALG-PFC_µB-FITC)<sub>net</sub> layer coating in different magnifications (<b>A</b>,<b>B</b>). Key to the symbol: (ALG-PFC_µB-FITC)<sub>net</sub>—alginate–perfluorooctyl labeled with fluorescent microbeads, cross-linked.</p>
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<p>Three-dimensional (3D) view in the coordinate system of the oxygenating layer (ALG-PFC_µB-FITC)<sub>net</sub> in confocal microscope in different planes of observation (<b>A</b>,<b>B</b>). Key to the symbol: (ALG-PFC_µB-FITC)<sub>net</sub>—alginate–perfluorooctyl labeled with fluorescent microbeads, cross-linked.</p>
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<p>The water contact angle. Key to the symbols: ALG—alginate, ALG-PFC—alginate–perfluorooctyl.</p>
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<p>The representing images of the droplets of water exhibiting the contact angles for assessed layer coatings: (<b>A</b>) support; (<b>B</b>) alginate; (<b>C</b>) alginate–perfluorooctyl.</p>
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<p>Wound treatment with the designed system (patient 002M).</p>
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<p>Wound treatment with the designed system (patient 003M).</p>
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<p>Wound treatment with the designed system (patient 004M).</p>
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<p>Wound treatment with the designed system (patient 005F).</p>
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<p>Compilation of the wound treatment effects in time for three different patients (001M, 002M, 003M): (1<b>A</b>–<b>3A</b>) before the treatment, (1<b>B</b>–<b>3B</b>) after 1 week of the treatment, and (1<b>C</b>–<b>3C</b>) after 2 weeks of the treatment.</p>
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<p>Illustration of wound healing with visible granulation and epithelization. Granulation and epithelization models based on Alven et al., 2022 [<a href="#B50-applsci-14-11200" class="html-bibr">50</a>]. ROS—reactive oxygen species.</p>
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<p>Theoretical (for normally healing wounds) and experimental duration of phases of proliferation and maturation. The histological pictures visualize the time range of granulation, indicating the proliferation phase, and epithelialization, indicating the maturation stage during the healing of treated non-healing wounds.</p>
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14 pages, 3787 KiB  
Article
Distinct Hemostasis and Blood Composition in Spiny Mouse Acomys cahirinus
by Nikita S. Filatov, Rafael R. Khismatullin, Airat I. Bilyalov, Alina I. Khabirova, Shakhnoza M. Salyakhutdinova, Roman V. Ursan, Roza N. Kasimova, Alina D. Peshkova, Insaf I. Gazizov, Elena I. Shagimardanova, Mary V. Woroncow, Andrey P. Kiyasov, Rustem I. Litvinov and Oleg A. Gusev
Int. J. Mol. Sci. 2024, 25(23), 12867; https://doi.org/10.3390/ijms252312867 - 29 Nov 2024
Viewed by 445
Abstract
The spiny mouse (Acomys species) is capable of scarless wound regeneration through largely yet unknown mechanisms. To investigate whether this capacity is related to peculiarities of the hemostatic system, we studied the blood of Acomys cahirinus in comparison to Mus musculus (Balb/c) [...] Read more.
The spiny mouse (Acomys species) is capable of scarless wound regeneration through largely yet unknown mechanisms. To investigate whether this capacity is related to peculiarities of the hemostatic system, we studied the blood of Acomys cahirinus in comparison to Mus musculus (Balb/c) to reveal differences in blood composition and clotting in both males and females. In response to surgical manipulations, blood clots formed in wounds of Acomys comprised a stronger hemostatic seal with reduced surgical bleeding in comparison with Balb/c. Acomys demonstrated notably shorter tail bleeding times and elevated clottable fibrinogen levels. Histological analysis revealed that clots from Acomys blood had densely packed fibrin-rich clots with pronounced fibrin segregation from erythrocytes. Acomys exhibited superior plasma clot stiffness as revealed with thromboelastography. The latter two characteristics are likely due to hyperfibrinogenemia. Light transmission platelet aggregometry demonstrated that ADP-induced platelet aggregates in Acomys males are stable, unlike the aggregates formed in the plasma of Balb/c undergoing progressive disaggregation over time. There were no apparent distinctions in platelet contractility and baseline expression of phosphatidylserine. Hematological profiling revealed a reduced erythrocytes count but increased mean corpuscular volume and hemoglobin content in Acomys. These results demonstrate the distinctive hemostatic potential of Acomys cahirinus, which may contribute to their remarkable regenerative capacity. Full article
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<p>Results of the tail bleeding assay (<b>A</b>) and levels of clottable fibrinogen in blood plasma (<b>B</b>) of male and female <span class="html-italic">Acomys cahirinus</span> mice versus Balb/c. The results are presented as mean ± SD (n = 5 for each species and gender group). * <span class="html-italic">p</span> &lt; 0.05; ** <span class="html-italic">p</span> &lt; 0.01.</p>
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<p>Representative histological images of blood clots from <span class="html-italic">Acomys cahirinus</span> (<b>A</b>,<b>C</b>) and Balb/c (<b>B</b>,<b>D</b>) (n = 3 for each). Arrows indicate <span class="html-italic">red</span> fibrin in the clot. Erythrocytes are <span class="html-italic">yellow</span>. (<b>E</b>) Characteristic dense <span class="html-italic">red</span> fibrin accumulations within the <span class="html-italic">Acomys cahirinus</span> blood clot. Multiple giant densely packed clusters and fibrous structures of fibrin are visualized (within dashed ovals), mostly separated from <span class="html-italic">yellow</span> erythrocytes. (<b>F</b>) Typical loose distribution of fibrin within the Balb/c blood clot. A significant predominance of loosely arranged fibrin (<span class="html-italic">red</span>), colocalized with erythrocytes (stained <span class="html-italic">pink</span>, not <span class="html-italic">yellow</span> as in the <span class="html-italic">Acomys</span> clot). Single islets of fibrin aggregates (marked by dashed ovals) were observed. Picro–Mallory stain. Magnification 25× (<b>A</b>–<b>D</b>) and 200× (<b>E</b>,<b>F</b>).</p>
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<p>TEG parameters of whole blood (<b>A</b>–<b>C</b>) and PFP (<b>D</b>–<b>F</b>) in male and female <span class="html-italic">Acomys cahirinus</span> and Balb/c: <span class="html-italic">R</span> (reaction time), α-angle (fibrin polymerization rate), and <span class="html-italic">G</span> (shear elastic modulus). n = 6 for paired whole blood samples and n = 9 for PFP samples from males; n = 5 for paired blood and PFP samples in females. The results are presented as mean ± SD. * <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, **** <span class="html-italic">p</span> &lt; 0.0001. ns—not significant.</p>
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<p>Parameters of clot contraction in whole blood (<b>A</b>,<b>B</b>) and in PRP (<b>C</b>,<b>D</b>) from <span class="html-italic">Acomys cahirinus</span> and Balb/c males and females. n = 6 for paired whole blood samples and n = 5 for PRP samples from males; n = 5 for paired blood and PRP samples from females. The results are presented as mean ± SD. * <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. ns—not significant.</p>
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<p>Averaged light transmission aggregograms recorded after ADP-induced platelet aggregation in PRP from male (<b>A</b>) and female (<b>B</b>) <span class="html-italic">Acomys cahirinus</span> and Balb/c (n = 5 for each species and gender group). Two-way ANOVA test in (<b>A</b>): <span class="html-italic">p</span> &lt; 0.05 at 12 min, <span class="html-italic">p</span> &lt; 0.01 at 15 min, <span class="html-italic">p</span> &lt; 0.001 at 20–30 min.</p>
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<p>A characteristic thromboelastogram and the key parameters of clot formation: <span class="html-italic">R</span>—reaction time, α—alpha angle, <span class="html-italic">MA</span>—maximum amplitude.</p>
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20 pages, 3616 KiB  
Article
Green Tea Epigallocatechin 3-Gallate Reduced Platelet Aggregation and Improved Anticoagulant Proteins in Patients with Transfusion-Dependent β-Thalassemia: A Randomized Placebo-Controlled Clinical Trial
by Touchwin Petiwathayakorn, Sasinee Hantrakool, Kornvipa Settakorn, Nuntouchaporn Hutachok, Adisak Tantiworawit, Nopphadol Chalortham, Pimpisid Koonyosying and Somdet Srichairatanakool
Foods 2024, 13(23), 3864; https://doi.org/10.3390/foods13233864 - 29 Nov 2024
Viewed by 813
Abstract
Patients with transfusion-dependent β-thalassemia (TDT) with iron overload have been linked to hypercoagulability and increased platelet (PLT) activation that causes thrombosis. Green tea extract (GTE) rich in epigallocatechin-3-gallate (EGCG) exerts iron-chelating and antithrombotic properties. The study aimed to assess the effects of GTE [...] Read more.
Patients with transfusion-dependent β-thalassemia (TDT) with iron overload have been linked to hypercoagulability and increased platelet (PLT) activation that causes thrombosis. Green tea extract (GTE) rich in epigallocatechin-3-gallate (EGCG) exerts iron-chelating and antithrombotic properties. The study aimed to assess the effects of GTE treatment on plasma coagulation state and PLT function in vitro and in patients with TDT. The subjects consumed a placebo or GTE tablets (50 mg and 2 × 50 mg EGCG equivalent) every day for two months. Blood was then collected from the treated patients for analyses of PLT numbers, agonist-induced PLT aggregation, and anti-coagulation proteins. In our findings indicate that the in vitro treatment of GTE (at least 1 mg EGCG equivalent) inhibited PLT aggregation in patients who were healthy and with thalassemia platelet-rich plasma (PRP), which was significant in the healthy PRP. Consistently, GTE treatment inhibited the PLT aggregation that had been ex vivo generated by collagen or ADP. In addition, consumption of GTE tablets greatly inhibited PLT aggregation and increased the plasma levels of proteins C and S, as well as the free protein S concentrations depending upon the time course, but not the GTE dosage. Moreover, plasma ferritin levels decreased in both green tea tablet groups in a time-dependent manner (p < 0.05 in the second month). In conclusion, EGCG-rich GTE diminished PLT aggregation in patients who were healthy and patients with thalassemia plasma. It also improved PLT aggregation and hypercoagulability in patients with TDT by increasing the antithrombotic activity of protein C and protein S. This would suggest an adjuvant of GTE could reduce the risk of thrombosis associated with iron overload. Full article
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Figure 1
<p>CONSORT flow diagram of the clinical study.</p>
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<p>Mean ± SEM values of PLT aggregation in N-PRP samples treated with PBS, GTE (1.25–5 mg EGCG eq) or EGCG (1.25–5 mg), which were induced by (<b>A</b>) 5 μM of ADP or (<b>B</b>) 2 μg/mL of Col agonist. Accordingly, * <span class="html-italic">p</span> &lt; 0.05, ** <span class="html-italic">p</span> &lt; 0.01 when compared without treatment; <sup>#</sup> <span class="html-italic">p</span> &lt; 0.05 when compared between genders.</p>
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<p>Mean ± SEM values of PLT aggregation in T-PRP samples treated with PBS, GTE (1.25–5 mg EGCG eq), and standard EGCG (1.25–5 mg) that had been induced by (<b>A</b>) 5 μM of ADP or (<b>B</b>) 2 μg/mL of Col agonist.</p>
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<p>PLT aggregation in N-PRP, T-PRP, and T-PRP^ samples treated with GTE (1.25–5.0 mg EGCG eq) that were induced by (<b>A</b>) 5 μM of ADP and (<b>B</b>) 2 μg/mL of Col. Data are expressed as mean ± SEM values, which ** <span class="html-italic">p</span> &lt; 0.01 when compared without treatment.</p>
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<p>PLT aggregation in T-PRP samples obtained from patients with BE, BM, and H thalassemia treated with PBS, GTE (1.25–5.0 mg EGCG eq), and standard EGCG (1.25–5.0 mg) that were induced by (<b>A</b>) 5 μM of ADP and (<b>B</b>) 2 μg/mL of Col. Data are expressed as individual and mean ± SEM values.</p>
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<p>PT (<b>A</b>), aPTT (<b>B</b>), PC (<b>C</b>), PS (<b>D</b>), and free PS activity (<b>E</b>) values in plasma of patients with TDT who had consumed a PB tablet (n = 11), a single GTE (50 mg EGCG eq) tablet (n = 9), and double-GTE (50 mg EGCG equivalent) tablets (n = 10) tablet once daily for 60 days. Data are expressed as individual and mean ± SD values. Accordingly, * <span class="html-italic">p</span> &lt; 0.05 when compared with day 0.</p>
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<p>PT (<b>A</b>), aPTT (<b>B</b>), PC (<b>C</b>), PS (<b>D</b>), and free PS activity (<b>E</b>) values in plasma of patients with TDT who had consumed a PB tablet (n = 11), a single GTE (50 mg EGCG eq) tablet (n = 9), and double-GTE (50 mg EGCG equivalent) tablets (n = 10) tablet once daily for 60 days. Data are expressed as individual and mean ± SD values. Accordingly, * <span class="html-italic">p</span> &lt; 0.05 when compared with day 0.</p>
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<p>Percentages for PLT aggregation in PRP samples of patients with TDT who had consumed PB tablet (n = 11), a single GTE (50 mg EGCG eq) tablet (n = 9), and double-GTE tablets (50 mg EGCG eq) tablet (n = 10) once daily for 60 days after induction by the addition of an agonist, either 5 μM of ADP (<b>A</b>) or 2 mg/mL of collagen (<b>B</b>) in vitro. Data are expressed as individual and mean ± SD values. Accordingly, * <span class="html-italic">p</span> &lt; 0.05 and ** <span class="html-italic">p</span> &lt; 0.01 when compared with T0.</p>
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<p>Levels of PI (<b>A</b>), TIBC (<b>B</b>), TS (<b>C</b>), and Ft (<b>D</b>) in the plasma of patients with TDT who had consumed a PB tablet (n = 11), a single GTE tablet (50 mg EGCG equivalent) (n = 9), and double-GTE tablets (100 mg EGCG eq) (n = 10) once daily for 60 days. Data are expressed as mean ± SD values.</p>
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12 pages, 1213 KiB  
Review
Platelet-Rich Plasma in Diabetic Foot Ulcer Healing: Contemplating the Facts
by Jacob Smith and Vikrant Rai
Int. J. Mol. Sci. 2024, 25(23), 12864; https://doi.org/10.3390/ijms252312864 - 29 Nov 2024
Viewed by 684
Abstract
Diabetic foot ulcers (DFUs), debilitating complication of diabetes, often lead to amputation even in the presence of current advanced treatment for DFUs. Platelet-rich plasma (PRP) containing growth factors and other proteins has been suggested as a potent therapeutic in promoting DFU healing. PRP [...] Read more.
Diabetic foot ulcers (DFUs), debilitating complication of diabetes, often lead to amputation even in the presence of current advanced treatment for DFUs. Platelet-rich plasma (PRP) containing growth factors and other proteins has been suggested as a potent therapeutic in promoting DFU healing. PRP is safe and effective in improving the DFU healing rate, decreasing healing time, and making chronic wounds viable for treatment. Though PRP is safe and effective in promoting DFU healing, there are inconsistencies in clinical outcomes. These varying results may be due to various concentrations of PRP being used. Most studies report dosage and timing, but none have reported the concentration of various factors. This is important, as the concentration of factors in PRP can vary significantly with each preparation and may directly impact the healing outcome. This critical review discusses the limiting factors and issues related to PRP therapy and future directives. A systematic search of PubMed and Google Scholar was performed with keywords including diabetic foot ulcer, ulcer healing, platelet-rich plasma, DFU treatment, and PRP limitations and efficacy, alone or in combination, to search the related articles. The articles describing DFU and the use of PRP in DFU healing were included. The existing literature suggests that PRP is effective and safe for promoting DFU healing, but larger clinical trials are needed to improve clinical outcomes. There is a need to consider multiple factors including the role of epigenetics, lifestyle modification, and the percentage composition of each constituent in PRP. Full article
(This article belongs to the Section Bioactives and Nutraceuticals)
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<p>Preparation of platelet-rich plasma. The first centrifugation separates the whole blood into three layers: 1. red blood cell layer (RBC), 2. buffy coat (BC) containing white blood cells and most of the platelets, and 3. platelet-poor plasma (PPP) layer. The second centrifugation of BC and PPP separates all platelets in platelet-rich plasma (PRP) layer and PPP.</p>
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<p>Molecular and cellular role supplemented by platelet-rich plasma (PRP) when administered to wound. The growth factors stored in platelets are secreted and stimulate the proliferation and migration of fibroblasts, keratinocytes, vascular smooth muscle cells (VSMCs), and endothelial cells toward the wound area. These cells and their interaction in-between and with immune cells help in promoting wound healing by suppressing inflammation and promoting tissue regeneration. PDGF—platelet-derived growth factor, EGF—epidermal growth factor, FGF—fibroblast growth factor, VEGF—vascular endothelial growth factor, IGF—insulin-like growth factor, TGF-β—transforming growth factor-beta, CD34—cluster of differentiation 34, HSP47—heat shock protein 47, SFA—fibroblast-specific surface antigen, SMA—smooth muscle actin, ECM—extracellular matrix, and IL-interleukin.</p>
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19 pages, 6477 KiB  
Article
First- vs. Second-Generation Autologous Platelet Concentrates and Their Implications for Wound Healing: Differences in Proteome and Secretome
by Hanna L. Stiller, Natarajan Perumal, Caroline Manicam, Emily R. Trzeciak, Julia Todt, Kerstin Jurk, Andrea Tuettenberg, Sven Schumann, Eik Schiegnitz and Sebastian Blatt
Bioengineering 2024, 11(11), 1171; https://doi.org/10.3390/bioengineering11111171 - 20 Nov 2024
Viewed by 712
Abstract
Differences in cell count and growth factor expression between first- and second-generation autologous platelet concentrates (APCs) have been well described. The debate over which formula best supports wound healing in various surgical procedures is still ongoing. This study aims to assess the whole [...] Read more.
Differences in cell count and growth factor expression between first- and second-generation autologous platelet concentrates (APCs) have been well described. The debate over which formula best supports wound healing in various surgical procedures is still ongoing. This study aims to assess the whole proteome assembly, cell content, immunological potential and pro-angiogenic potential of second-generation APC, Platelet-Rich Fibrin (PRF) vs. first-generation APC, Platelet-Rich Plasma (PRP). The global proteome of the APCs was analyzed using nano-liquid chromatography mass spectrometry. Blood cell concentrations were determined by an automated cell counter. The effect of APCs on macrophage polarization was analyzed by flow cytometry. A yolk sac membrane (YSM) assay was used to monitor the neo-vessel formation and capillary branching in vivo. Cell count analysis revealed a higher number/concentration of leukocytes in PRF vs. PRP. Incubation of macrophages with PRP or platelet-free plasma (PFP) did not induce a significant pro-inflammatory state but led to a shift to the M0/M2 phenotype as seen in wound healing for all tested formulas. Label-free proteomics analysis identified a total of 387 proteins from three biological replicates of the respective designated groups. PRF induced increased formation of neo-vessels and branching points in vivo in comparison to PRP and PFP (each p < 0.001), indicating the enhanced pro-angiogenic potential of PRF. Overall, PRF seems superior to PRP, an important representative of first-generation formulas. Inclusion of leucocytes in PRF compared to PRP suggested rather an anti-inflammatory effect on macrophages. These results are important to support the versatile clinical applications in regenerative medicine for second-generation autologous platelet concentrates to optimize wound healing. Full article
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<p>Yolk sac membrane (YSM) assay. (<b>A</b>): In ovo analysis of PRF (2) and PFP (*) in safe distance from the embryo (1). (<b>B</b>): Microscopical analysis of the induced vessels (3) in direct contact with the PRF (2) and in safe distance from the embryo (1). (<b>C</b>): Vessel analysis after grayscale conversion and background extraction with FIJI software (Version 2.9.0). Arrow: vessel.</p>
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<p>(<b>a</b>) Cytological analysis of PRF, PRP, and PFP compared to EDTA-blood in platelets, erythrocytes, and leukocytes (ns: <span class="html-italic">p</span> &gt; 0.05; *: <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; ****: <span class="html-italic">p</span> &lt; 0.0001). (<b>b</b>) Cytological analysis of PRF, PRP, and PFP compared to EDTA-blood in neutrophilic granulocytes, lymphocytes, and monocytes (ns: <span class="html-italic">p</span> &gt; 0.05; *: <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).</p>
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<p>Platelet-rich plasma (PRP) and platelet-free plasma (PFP) polarize human monocyte derived macrophages to a “M0/M2-like” phenotype. Macrophages were polarized or treated with PRP or PFP. Following 2 days of incubation, surface marker expression was evaluated with flow cytometry. The bar diagram indicates the mean fluorescence intensity (MFI) of each marker, normalized to the matching donor “M0” control. The histograms show one representative result (<span class="html-italic">n</span> = 4 donors, means ± SD, * <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, and **** <span class="html-italic">p</span> &lt; 0.0001 analyzed by two-way ANOVA, corrected for multiple comparisons using Tukey tests).</p>
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<p>Representative protein profiles of the designated groups comprising the soluble proteins from sPRF, sPRP, and sPFP, and cell-derived proteins from pPRF, pPRP, and pPFP resolved in 1DE-PAGE after colloidal blue staining.</p>
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<p>Label-free quantitative proteomics analysis of the supernatant and pellet fragments of the APC. (<b>a</b>) Heat map depicts the hierarchical clustering of 387 total proteins based on the log<sub>2</sub> protein intensity corresponding to the designated groups. (<b>b</b>) Bar charts show the degree of mean percentage of top abundant proteins in the designated groups. (<b>c</b>,<b>d</b>) Venn diagrams illustrate the total number of proteins identified in all samples.</p>
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<p>Heat map depicts the hierarchical clustering of the total of 268 significantly differentially abundant proteins in the designated comparison groups. Significance threshold is at <span class="html-italic">p</span> &lt; 0.05. The up-regulated proteins are shown in red and the down-regulated proteins are in green.</p>
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<p>Volcano plots illustrate the significantly differentially abundant proteins identified based on the log<sub>2</sub> difference in the (<b>a</b>) pPRF vs. sPRF, (<b>b</b>) pPRP vs. sPRP (<b>c</b>) pPFP vs. sPFP, (<b>d</b>) sPRF vs. sPFP, (<b>e</b>) sPRP vs. sPFP, (<b>f</b>) sPRF vs. sPRP, (<b>g</b>) pPRF vs. pPFP, (<b>h</b>) pPRP vs. pPFP, and (<b>i</b>) pPRF vs. pPRP. The significance threshold is at <span class="html-italic">p</span> &lt; 0.05. The up-regulated proteins are shown in red and the down-regulated proteins are in green.</p>
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<p>Heat map depicts the top significant enriched biological functions and canonical pathways of the differentially abundant proteins in the designated comparison groups. The significance of enrichment (−log<sub>10</sub> (B-H <span class="html-italic">p</span>-value) is scaled by color intensity; <span class="html-italic">p</span> &lt; 0.05.</p>
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<p>Microscopic analysis of vessel formation in the yolk sac membrane (YSM) assay for PRF (<b>A</b>) and PFP (<b>B</b>). The red arrow in (<b>A</b>) highlights the enhanced vessel formation in the PRF group compared to PFP. Panel (<b>C</b>) quantifies vessel formation, and (<b>D</b>) depicts the number of branching points, demonstrating a statistically significant increase in pro-angiogenic activity in the PRF group (**** <span class="html-italic">p</span> &lt; 0.0001).</p>
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