Isolation and characterization of plant-derived exosome-like nanoparticles from Carica papaya L. fruit and their potential as anti-inflammatory agent.

Inflammation is an immune system response that identifies and eliminates foreign material. However, excessive and persistent inflammation could disrupt the healing process. Plant-derived exosome-like nanoparticles (PDENs) are a promising candidate for therapeutic application because they are safe, biodegradable and biocompatible. In this study, papaya PDENs were isolated by a PEG6000-based method and characterized by dynamic light scattering (DLS), transmission Electron Microscopy (TEM), bicinchoninic acid (BCA) assay method, GC-MS analysis, total phenolic content (TPC) analysis, and 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay. For the in vitro test, we conducted internalization analysis, toxicity assessment, determination of nitrite concentration, and assessed the expression of inflammatory cytokine genes using qRT-PCR in RAW 264.7 cells. For the in vivo test, inflammation was induced by caudal fin amputation followed by analysis of macrophage and neutrophil migration in zebrafish (Danio rerio) larvae. The result showed that papaya PDENs can be well isolated using the optimized differential centrifugation method with the addition of 30 ppm pectolyase, 15% PEG, and 0.2 M NaCl, which exhibited cup-shaped and spherical morphological structure with an average diameter of 168.8±9.62 nm. The papaya PDENs storage is stable in aquabidest and 25 mM trehalose solution at -20˚C until the fourth week. TPC estimation of all papaya PDENs ages did not show a significant change, while the DPPH test exhibited a significant change in the second week. The major compounds contained in Papaya PDENs is 2,3-dihydro-3,5-dihydroxy-6-methyl-4H-pyran-4-one (DDMP). Papaya PDENs can be internalized and is non-cytotoxic to RAW 264.7 cells. Moreover, LPS-induced RAW 264.7 cells treated with papaya PDENs showed a decrease in NO production and downregulation mRNA expression of pro-inflammatory cytokine genes (IL-1B and IL-6) and an upregulation in mRNA expression of anti-inflammatory cytokine gene (IL-10). In addition, in vivo tests conducted on zebrafish treated with PDENs papaya showed inhibition of macrophage and neutrophil cell migration. These findings suggest that PDENs papaya possesses anti-inflammatory properties.

Anti-inflammatory potential of goldenberry-derived exosome-like nanoparticles in macrophage polarization.

Objective: Overpopulated M1 macrophages can trigger chronic inflammation. Plant-derived exosome-like nanoparticles have been reported to show beneficial bioactivities. Aim: To isolate PDEN from goldenberry fruits and evaluate its anti-inflammatory potential in macrophage polarization. Methods: GDEN were isolated by centrifugation and precipitation methods. LPS-induced RAW 264.7 cells were treated with GDEN before being evaluated with nitric oxide production assay and flow cytometry of CD80 and CD209. Results: GDEN averaged 227.7 nm in size and spherical-shaped. GDEN 40 µg/ml decreased NO production in LPS-induced cells. Flow cytometry showed that CD209 (M2 marker) positive cells were up-regulated after being treated with 20 µg/ml GDEN. Conclusion: GDEN showed anti-inflammatory potential through the ability to reduce M1 macrophages product and promote M2 polarization.

During inflammation, there are immune cells called macrophages which can act in two different ways. They are M1 cells which drive inflammation and M2 cells which resolve inflammation. The imbalance of M1 and M2 cells can be harmful to the body. For example, having too many M1 macrophages will cause chronic inflammation. Nano-sized particles found in plants are known to have some benefits as future drugs. They have good biocompatibility, can be produced in large quantities and show various bioactivities. Goldenberry has been used as an herb and is known to have good effects on human health, like reducing inflammation. In this study, nanoparticles from goldenberry fruits, also known as goldenberry-derived exosome-like nanoparticles (GDEN), were successfully extracted using a chemical, polyethylene glycol (PEG) 6000. GDEN showed no harmful effect on mice macrophage cells (RAW 264.7) and had been successfully taken inside by the cells. GDEN also showed reducing inflammation potential. This was supported by the decrease of M1 cell product and the increase of M2 cell marker. Hence, GDEN are worthy to be studied further for its usage to treat inflammation and other related health problems.

Characterizing Nanoparticle Isolated by Yam Bean (Pachyrhizus erosus) as a Potential Agent for Nanocosmetics: An in vitro and in vivo Approaches.

BACKGROUND: This study investigated the potential of Plant-Derived Exosome-Like Nanoparticles (PDENs) as cosmeceutical nanocarriers for treating skin problems, such as scar removal, face rejuvenation, anti-aging, and anti-pigmentation. OBJECTIVE: Researchers isolated PDENs from Yam Bean (Pachyrhizus erosus) using PEG-based precipitation, gradual filtration, and various centrifugations at low temperatures. Followed by in vitro and in vivo studies using HDF cells and Zebrafish. METHODS: The morphology of the YB-PDENs was determined using TEM analysis, they had a spherical shape with diameters of 236,83 ± 9,27nm according to PSA. The study found that YB-PDENs were stable in aquabidest at 4°C for one month of storage and had ~-26,5 mV of Zeta Potential. The concentration of YB-PDENs was measured using the BCA Assay, and internalization of YB-PDENs to HDF cells was observed using a Confocal Laser Scanning Microscope labelled with PKH67. RESULT: As for cytotoxicity, after 24 and 72 hours of incubation with YB-PDENs, the viability of HDF cells remained more than 80%. The study also examined cell migration using the Scratch Assay and found that at 2,5 µg/mL, YB-PDENs had better migration results than other concentrations. Immunocytochemistry showed that collagen expression was higher after 14 days of incubation with YB-PDENs, and melanocytes in zebrafish decreased at each concentration compared with controls. CONCLUSION: In conclusion, this study is the first to extract and describe PDENs from Yam Bean (Pachyrhizus erosus), with YB-PDENs having a promising anti-melanogenic effect in skin treatment. This study highlights the potential of YB-PDENs as a promising alternative to depigmentation and skin whitening treatments.

Isolation of plant-derived exosome-like nanoparticles (PDENs) from Solanum nigrum L. berries and Their Effect on interleukin-6 expression as a potential anti-inflammatory agent.

Inflammation is a temporary response of the immune system that can be treated using common anti-inflammatory drugs. However, prolonged use of these drugs increases the risk of adverse side effects. Accordingly, there is an increasing need for alternative treatments for inflammation with fewer side effects. Exosomes are extracellular vesicles secreted by most eukaryotic cells and have been studied as a candidate for cell-free therapy for inflammatory diseases due to their immunomodulatory and anti-inflammatory properties. In recent years, the focus of exosome research has shifted from animal cell-derived exosomes to plant-derived exosome-like nanoparticles (PDENs). Plant-derived exosome-like nanoparticles (PDENs) are easier to obtain, have minimal safety concerns, and can be produced in higher quantities and lower cost than exosomes derived from animal cells. In this study, the isolation and analysis of the anti-inflammatory potential of PDENs from black nightshade berries (Solanum nigrum L.) were carried out. The results of isolation and characterization showed that PDENs had a spherical morphology, measuring around 107 nm with zeta potential of -0.6 mV, and had a protein concentration of 275.38 µg/mL. PDENs were also shown to be internalized by RAW264.7 macrophage cell line after 2 hours of incubation and had no cytotoxicity effect up to the concentration of 2.5 µg/mL. Furthermore, exposure to several doses of PDENs to the LPS-stimulated RAW264.7 cell significantly decreased the expression of pro-inflammatory cytokine gene IL-6, as well as the expression of IL-6 protein up to 97,28%. GC-MS analysis showed the presence of neral, a monoterpene compound with known anti-inflammatory properties, which may contribute to the anti-inflammatory activity of PDENs isolated from Solanum nigrum L. berries. Taken together, the present study was the first to isolate and characterize PDENs from Solanum nigrum L. berries. The results of this study also demonstrated the anti-inflammatory activity of PDEN by suppressing the production of IL-6 in LPS-stimulated RAW264.7 cells.

Potential of Plant-derived Exosome-like Nanoparticles from Physalis peruviana Fruit for Human Dermal Fibroblast Regeneration and Remodeling.

AIM: This research aimed to study the potential of PDEN from P. peruviana fruits (PENC) for regenerating and remodeling HDF. BACKGROUND: Large wounds are dangerous and require prompt and effective healing. Various efforts have been undertaken, but have been somewhat ineffective. Plant-derived exosome-like nanoparticles (PDEN) are easily sampled, relatively cost-effective, exhibit high yields, and are nonimmunogenic. OBJECTIVE: The objective of the study was to isolate and characterize PDEN from Physalis peruviana (PENC), and determine PENC's internalization and toxicity on HDF cells, PENC's ability to regenerate HDF (proliferation and migration), and PENC ability's to remodel HDF (collagen I and MMP- 1 production). METHOD: PENC was isolated using gradual filtration and centrifugation, followed by sedimentation using PEG6000. Characterization was done using a particle size analyzer, zeta potential analyzer, TEM, and BCA assay. Internalization was done using PKH67 staining. Toxicity and proliferation assays were conducted using MTT assay; meanwhile, migration assay was carried out by employing the scratch assay. Collagen I production was performed using immunocytochemistry and MMP-1 production was conducted using ELISA. RESULT: MTT assay showed a PENC concentration of 2.5 until 500 µg/mL and being non-toxic to cells. PENC has been found to induce cell proliferation in 1, 3, 5, and 7 days. PENC at a concentration of 2.5, 5, and 7.5 µg/mL, also accelerated HDF migration using the scratch assay in two days. In remodeling, PENC upregulated collagen-1 expression from day 7 to 14 compared to control. MMP-1 declined from day 2 to 7 in every PENC concentration and increased on day 14. Meanwhile, ROS declined from day 2 to 7 and 14, with PENC inducing a rise in the ROS level compared to control. Overall, PENC at concentrations of 2.5, 5, and 7.5 µg/mL induced HDF proliferation and migration, upregulated collagen I production, and decreased MMP-1 levels. CONCLUSION: Isolated PENC was 190-220 nm in size, circular, covered with membrane, and its zeta potential was -6.7 mV; it could also be stored at 4°C for up to 2 weeks in aqua bidest. Protein concentration ranged between 170-1,395 µg/mL. Using PKH67, PENC could enter HDF within 6 hours. PENC was non-toxic up to a concentration of 500 µg/mL. Using MTT and scratch assay, PENC was found to elevate HDF proliferation and migration, and reorganize actin. Using immunocytochemistry, collagen I was upregulated by PENC, whereas MMP-1 concentration was reduced.

Plant-Derived Exosome-like Nanoparticles for Biomedical Applications and Regenerative Therapy.

Plant-derived exosome-like nanoparticles (PDENs) comprise various bioactive biomolecules. As an alternative cell-free therapeutic approach, they have the potential to deliver nano-bioactive compounds to the human body, and thus lead to various anti-inflammatory, antioxidant, and anti-tumor benefits. Moreover, it is known that Indonesia is one of the herbal centers of the world, with an abundance of unexplored sources of PDENs. This encouraged further research in biomedical science to develop natural richness in plants as a source for human welfare. This study aims to verify the potential of PDENs for biomedical purposes, especially for regenerative therapy applications, by collecting and analyzing data from the latest relevant research and developments.

Development and Characterization of a Gel Formulation Containing Golden Cherry Exosomes (Physalis minima) as a Potential Anti-Photoaging.

AIM: The present study aims to produce a novel therapeutic approach for the treatment of photoaging. BACKGROUND: Plant-derived exosome-like nanoparticles (PDENs) are nano-sized vesicles containing biomolecules released by multivesicular bodies. Recently, studies have shown the efficacy of exosomes in treating photoaging through increasing collagen synthesis and decreasing collagen degradation. In addition, some PDENs were also proven to contain bioactive metabolites, which also have potential antioxidant activity to mitigate the risk of photoaging. OBJECTIVE: Formulating and developing a gel and incorporating it with exosomes derived from golden cherry (Physalis minima). METHOD: The formulation was developed by first preparing various base formulations with different compositions and selecting the best through evaluation tests. The results showed that only polymer base natrosol with a concentration of 0.25% was suitable for incorporating exosomes. The selected base was then incorporated with various concentrations of golden cherry exosomes and was evaluated regarding its physical and stability profile. RESULT: The result demonstrates that the incorporated gel displayed pleasant organoleptic properties and a pH compatible with the skin, with pseudoplastic flow and a suitable viscosity for topical application. The stability study also only revealed minor changes in viscosity and pH without affecting the general stability of the formulation. Formulation incorporating 0.25% golden cherry exosomes had shown the best stability profile compared to other concentrations. On characterization, although the incorporated exosomes showed heterogeneous particle size distribution (PI > 0.3), they still maintained their structural integrity. In addition, the incorporated exosomes showed antioxidant activity with IC 50 of 372.435 µg/mL, which can help mitigate the risk of photoaging. CONCLUSION: Golden cherry exosomes have been successfully incorporated into gel and, thus, can be potentially utilized as a novel therapeutic approach for the treatment of photoaging.

The combinatory effect of scaffold topography and culture condition: an approach to nucleus pulposus tissue engineering.

Scaffold topography and culture medium conditions for human wharton's jelly mesenchymal stem cells (hWJ-MSC) are critical components of the approach to nucleus pulposus (NP) tissue engineering. Aim: To evaluate the silk fibroin (SF) scaffold topography analysis (optimal thickness and pore diameter) and to determine culture medium conditions for the growth and differentiation of hWJ-MSC. Method: hWJ-MSCs were seeded into different thicknesses and pore size diameters and grown in different concentrations of glucose, platelet rich plasma (PRP) and oxygen. The cell-seeded scaffold was evaluated for cell attachment, growth and differentiation potency. Results & discussion: The results indicated that SF scaffold with a minimum thickness 3.5 mm and pore diameter of 500 µm with cells cultured under low glucose, 10% PRP and normoxia conditions induced the growth and differentiation of hWJ-MSCs, indicated by the accumulation of glycosaminoglycans content and the presence of type II collagen, as markers of NP-like cells.

Infectious Disease Modeling with Socio-Viral Behavioral Aspects-Lessons Learned from the Spread of SARS-CoV-2 in a University.

When it comes to understanding the spread of COVID-19, recent studies have shown that pathogens can be transmitted in two ways: direct contact and airborne pathogens. While the former is strongly related to the distancing behavior of people in society, the latter are associated with the length of the period in which the airborne pathogens remain active. Considering those facts, we constructed a compartmental model with a time-dependent transmission rate that incorporates the two sources of infection. This paper provides an analytical and numerical study of the model that validates trivial insights related to disease spread in a responsive society. As a case study, we applied the model to the COVID-19 spread data from a university environment, namely, the Institut Teknologi Bandung, Indonesia, during its early reopening stage, with a constant number of students. The results show a significant fit between the rendered model and the recorded cases of infections. The extrapolated trajectories indicate the resurgence of cases as students' interaction distance approaches its natural level. The assessment of several strategies is undertaken in this study in order to assist with the school reopening process.

Potential Cell-Based and Cell-Free Therapy for Patients with COVID-19.

Since it was first reported, the novel coronavirus disease 2019 (COVID-19) remains an unresolved puzzle for biomedical researchers in different fields. Various treatments, drugs, and interventions were explored as treatments for COVID. Nevertheless, there are no standard and effective therapeutic measures. Meanwhile, mesenchymal stem cell (MSC) therapy offers a new approach with minimal side effects. MSCs and MSC-based products possess several biological properties that potentially alleviate COVID-19 symptoms. Generally, there are three classifications of stem cell therapy: cell-based therapy, tissue engineering, and cell-free therapy. This review discusses the MSC-based and cell-free therapies for patients with COVID-19, their potential mechanisms of action, and clinical trials related to these therapies. Cell-based therapies involve the direct use and injection of MSCs into the target tissue or organ. On the other hand, cell-free therapy uses secreted products from cells as the primary material. Cell-free therapy materials can comprise cell secretomes and extracellular vesicles. Each therapeutic approach possesses different benefits and various risks. A better understanding of MSC-based and cell-free therapies is essential for supporting the development of safe and effective COVID-19 therapy.

Spidroin striped micropattern promotes chondrogenic differentiation of human Wharton's jelly mesenchymal stem cells.

Cartilage tissue engineering, particularly micropattern, can influence the biophysical properties of mesenchymal stem cells (MSCs) leading to chondrogenesis. In this research, human Wharton's jelly MSCs (hWJ-MSCs) were grown on a striped micropattern containing spider silk protein (spidroin) from Argiope appensa. This research aims to direct hWJ-MSCs chondrogenesis using micropattern made of spidroin bioink as opposed to fibronectin that often used as the gold standard. Cells were cultured on striped micropattern of 500 µm and 1000 µm width sizes without chondrogenic differentiation medium for 21 days. The immunocytochemistry result showed that spidroin contains RGD sequences and facilitates cell adhesion via integrin ß1. Chondrogenesis was observed through the expression of glycosaminoglycan, type II collagen, and SOX9. The result on glycosaminoglycan content proved that 1000 µm was the optimal width to support chondrogenesis. Spidroin micropattern induced significantly higher expression of SOX9 mRNA on day-21 and SOX9 protein was located inside the nucleus starting from day-7. COL2A1 mRNA of spidroin micropattern groups was downregulated on day-21 and collagen type II protein was detected starting from day-14. These results showed that spidroin micropattern enhances chondrogenic markers while maintains long-term upregulation of SOX9, and therefore has the potential as a new method for cartilage tissue engineering.

Extracellular vesicles: a promising cell-free therapy for cartilage repair.

Few effective therapies for cartilage repair have been found as cartilage has a low regenerative capacity. Extracellular vesicles (EVs), including exosomes, are produced by cells and contain bioactive components such as nucleic acids, proteins, lipids and other metabolites that have potential for treating cartilage injuries. Challenges like the difficulty in standardizing targeted therapy have prevented EVs from being used frequently as a treatment option. In this review we present current studies, mechanisms and delivery strategies of EVs. Additionally, we describe the challenges and future directions of EVs as therapeutic agents for cartilage repair.

Nanotopography in directing osteogenic differentiation of mesenchymal stem cells: potency and future perspective.

Severe bone injuries can result in disabilities and thus affect a person's quality of life. Mesenchymal stem cells (MSCs) can be an alternative for bone healing by growing them on nanopatterned substrates that provide mechanical signals for differentiation. This review aims to highlight the role of nanopatterns in directing or inducing MSC osteogenic differentiation, especially in bone tissue engineering. Nanopatterns can upregulate the expression of osteogenic markers, which indicates a faster differentiation process. Combined with growth factors, nanopatterns can further upregulate osteogenic markers, but with fewer growth factors needed, thereby reducing the risks and costs involved. Nanopatterns can be applied in scaffolds for tissue engineering for their lasting effects, even in vivo, thus having great potential for future bone treatment.

Conductive Polymeric-Based Electroactive Scaffolds for Tissue Engineering Applications: Current Progress and Challenges from Biomaterials and Manufacturing Perspectives.

The practice of combining external stimulation therapy alongside stimuli-responsive bio-scaffolds has shown massive potential for tissue engineering applications. One promising example is the combination of electrical stimulation (ES) and electroactive scaffolds because ES could enhance cell adhesion and proliferation as well as modulating cellular specialization. Even though electroactive scaffolds have the potential to revolutionize the field of tissue engineering due to their ability to distribute ES directly to the target tissues, the development of effective electroactive scaffolds with specific properties remains a major issue in their practical uses. Conductive polymers (CPs) offer ease of modification that allows for tailoring the scaffold's various properties, making them an attractive option for conductive component in electroactive scaffolds. This review provides an up-to-date narrative of the progress of CPs-based electroactive scaffolds and the challenge of their use in various tissue engineering applications from biomaterials perspectives. The general issues with CP-based scaffolds relevant to its application as electroactive scaffolds were discussed, followed by a more specific discussion in their applications for specific tissues, including bone, nerve, skin, skeletal muscle and cardiac muscle scaffolds. Furthermore, this review also highlighted the importance of the manufacturing process relative to the scaffold's performance, with particular emphasis on additive manufacturing, and various strategies to overcome the CPs' limitations in the development of electroactive scaffolds.

Challenges and strategy in treatment with exosomes for cell-free-based tissue engineering in dentistry.

In dentistry, problems of craniofacial, osteochondral, periodontal tissue, nerve, pulp or endodontics injuries, and osteoarthritis need regenerative therapy. The use of stem cells in dental tissue engineering pays a lot of increased attention, but there are challenges for its clinical applications. Therefore, cell-free-based tissue engineering using exosomes isolated from stem cells is regarded an alternative approach in regenerative dentistry. However, practical use of exosome is restricted by limited secretion capability of cells. For future regenerative treatment with exosomes, efficient strategies for large-scale clinical applications are being studied, including the use of ceramics-based scaffold to enhance exosome production and secretion which can resolve limited exosome secretory from the cells when compared with the existing methods available. Indeed, more research needs to be done on these strategies going forward.

DeepSperm: A robust and real-time bull sperm-cell detection in densely populated semen videos.

BACKGROUND AND OBJECTIVE: Object detection is a primary research interest in computer vision. Sperm-cell detection in a densely populated bull semen microscopic observation video presents challenges that are more difficult than those presented by other general object-detection cases. These challenges include partial occlusion, vast number of objects in a single video frame, tiny size of the object, artifacts, low contrast, low video resolution, and blurry objects because of the rapid movement of the sperm cells. This study proposes a deep neural network architecture, called DeepSperm, that solves the aforementioned problems and is more accurate and faster than state-of-the-art architectures. METHODS: In the proposed architecture, we use only one detection layer, which is specific for small object detection. For handling overfitting and increasing accuracy, we set a higher input network resolution, use a dropout layer, and perform data augmentation on saturation and exposure. Several hyper-parameters are tuned to achieve better performance. Mean average precision (mAP), confusion matrix, precision, recall, and F1-score are used to measure accuracy. Frame per second (fps) is used to measure speed. We compare our proposed method with you only look once (YOLO) v3 and YOLOv4. RESULTS: In our experiment, we achieve 94.11 mAP on the test dataset, F1-score of 0.93, and a processing speed of 51.9 fps. In comparison with YOLOv4, our proposed method is 2.18 x faster on testing, and 2.9 x faster on training with a small dataset, while achieving comparative detection accuracy. The weights file size was also reduced significantly, with one-twentieth that of YOLOv4. Moreover, it requires a 1.07 x less graphical processing unit (GPU) memory than YOLOv4. CONCLUSIONS: This study proposes DeepSperm, which is a simple, effective, and efficient deep neural network architecture with its hyper-parameters and configuration to detect bull sperm cells robustly in real time. In our experiments, we surpass the state-of-the-art in terms of accuracy, speed, and resource needs.

Propolis Can Improve Caudal Fin Regeneration in Zebrafish (Danio rerio) Induced by The Combined Administration of Alloxan and Glucose.

Hyperglycemia, a primary symptom in diabetes mellitus, is associated with difficulties in wound healing and regeneration. This condition is due to the length of the inflammatory phase and free radicals. Furthermore, there is evidence that molecular pathogenesis is involved in impaired wound healing in diabetics. As an animal model, zebrafish have many shared orthologous genes with human that are involved in protein regulation of wound healing and regeneration. Little is known about natural drugs that may be used to treat complications of wound healing in diabetes. Propolis, however, is known to consist of various organic compounds such as phenols and flavonoids with antioxidant and anti-inflammatory activities. This research aims to study propolis' effect on caudal fin regeneration and relative expression of several genes belonging to Hedgehog, bone morphogenetic protein (BMP), and Wnt signaling hyperglycemic (HG) zebrafish. GC-MS analysis and antioxidant activity testing were performed on ethanolic extract of propolis (EEP). Caudal fin regeneration was analyzed using ImageJ; blood glucose levels were measured; and relative gene expression analysis of shha, igf2a, bmp2b, and col1a2 was performed by the real-time polymerase chain reaction method with the ß-actin housekeeping gene. Impairment of caudal fin regeneration in zebrafish hyperglycemia was characterized by a low percentage of regeneration and decreased relative gene expression. EEP at 15 ppm could increase the percentage of caudal fin regeneration and the expression of shha, igf2a, bmp2b, and col1a2. Based on the results, it appears that phenols and flavonoids from the EEP can improve the caudal fin regeneration of HG zebrafish.

Cell penetration and chondrogenic differentiation of human adipose derived stem cells on 3D scaffold.

The ability of cells to penetrate the scaffold and differentiate into chondrocyte is important in cartilage engineering. The aim of this research was to evaluate the use of silk fibroin 3D scaffold in facilitating the growth of stem cell and to study the role of L-ascorbic acid and platelet rich plasma (PRP) in proliferation and differentiation genes. Cell penetration and type II collagen content in the silk fibroin scaffold was analyzed by confocal microscopy. Relative expressions of CDH2, CCND1, CTNNB1 and COL2A1 were analyzed by reverse transcription-quantitative PCR (RT-qPCR). The silk fibroin 3D scaffold could facilitate cell penetration. L-ascorbic acid and PRP increased the expression of CDH2 and COL2A1 on the 21st day of treatment while PRP inhibited CTNNB1 and CCND1.

Plant-derived exosome-like nanoparticles: A concise review on its extraction methods, content, bioactivities, and potential as functional food ingredient.

Plant-derived exosome-like nanoparticles (PDENs) are small vesicles released by multivesicular bodies mainly to communicate between cells and regulate immunity against pathogen attack. Current studies have reported that PDENs could modulate gene expression in a cross-kingdom fashion. Therefore, PDENs could be a potential future functional food ingredient as their cross-kingdom communication abilities were reported to exert multiple health benefits. Macrophage and other cells have been reported to absorb PDENs in a manner regulated by the membrane lipid and protein profile and the intactness of the PDENs lipid bilayer. PDENs could be extracted from plant materials by various techniques such as ultracentrifugation, immunoaffinity, size-based isolation, and precipitation, though each method has its pros and cons. PDENs mainly contain lipid, protein, and genetic materials, mainly micro RNAs, which could exert multiple health benefits and functionalities when consumed in sufficient amounts. However, most studies on the health functionalities of PDENs were conducted through in-vitro and in-vivo studies, and its potency to be used as a functional ingredient remains a question as PDENs are sensitive to storage and processing condition and requires costly extraction method. This concise review features various exosome extraction methods, contents of PDENs and their roles, the health functionalities of PDENs, and its potency as a functional food ingredient.

Chondrogenic differentiation of Wharton's Jelly mesenchymal stem cells on silk spidroin-fibroin mix scaffold supplemented with L-ascorbic acid and platelet rich plasma.

In this research, hWJ-MSCs were grown on silk scaffolds and induced towards chondrogenesis by supplementation with L-ascorbic acid (LAA) or platelet rich plasma (PRP). Silk scaffolds were fabricated with salt leaching method by mixing silk fibroin (SF) with silk spidroin (SS). The silk fibroin was obtained from Bombyx mori cocoon that had been degummed, and the silk spidroin was obtained from wild-type spider Argiope appensa. The effect of scaffold composition and inducer on cell proliferation was observed through MTT assay. The most optimal treatment then continued to be used to induce hWJ-MSC towards chondrogenic differentiation for 7 and 21 days. Scaffolds characterization showed that the scaffolds produced had 3D structure with interconnected pores, and all were biocompatible with hWJ-MSCs. Scaffold with the addition of 10% SS + 90% SF showed higher compressive strength and better pore interconnectivity in comparison to 100% silk fibroin scaffold. After 48 h, cells seeded on scaffold with spidroin and fibroin mix had flattened morphology in comparison to silk fibroin scaffold which appeared to be more rounded on the scaffold surface. Scaffold with 10% (w/w) of silk spidroin (SS) + 90% (w/w) of silk fibroin (SF) was the most optimal composition for cell proliferation. Immunocytochemistry of integrin ß1 and RGD sequence, showed that scaffold with SS 10% provide better cell attachment with the presence of RGD sequence from the spidroin silk which could explain the higher cell proliferation than SF100% scaffold. Based on Alcian Blue staining and Collagen Type II immunocytochemistry (ICC), cells grown on 10% SS + 90% SF scaffold with 10% PRP supplementation were the most optimal to support chondrogenesis of hWJ-MSCs. These results showed that the addition of spidroin silk from A. appensa. had impact on scaffold compressive strength and chondrogenic differentiation of hWJ-MSC and had the potential for further development of bio-based material scaffold in cartilage tissue engineering.