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Biosensors May 2024Intracellular delivery, the process of transporting substances into cells, is crucial for various applications, such as drug delivery, gene therapy, cell imaging, and... (Review)
Review
Intracellular delivery, the process of transporting substances into cells, is crucial for various applications, such as drug delivery, gene therapy, cell imaging, and regenerative medicine. Among the different approaches of intracellular delivery, mechanoporation stands out by utilizing mechanical forces to create temporary pores on cell membranes, enabling the entry of substances into cells. This method is promising due to its minimal contamination and is especially vital for stem cells intended for clinical therapy. In this review, we explore various mechanoporation technologies, including microinjection, micro-nano needle arrays, cell squeezing through physical confinement, and cell squeezing using hydrodynamic forces. Additionally, we highlight recent research efforts utilizing mechanoporation for stem cell studies. Furthermore, we discuss the integration of mechanoporation techniques into microfluidic platforms for high-throughput intracellular delivery with enhanced transfection efficiency. This advancement holds potential in addressing the challenge of low transfection efficiency, benefiting both basic research and clinical applications of stem cells. Ultimately, the combination of microfluidics and mechanoporation presents new opportunities for creating comprehensive systems for stem cell processing.
Topics: Stem Cells; Humans; Microfluidics; Animals; Drug Delivery Systems
PubMed: 38785730
DOI: 10.3390/bios14050256 -
Archives of Insect Biochemistry and... May 2024Invasive insect pests, currently, pose a serious economic threat to several staple crops all over the world, one such being the fall armyworm, Spodoptera frugiperda. It...
CRISPR/Cas9 mediated validation of spermatogenesis-related gene, tssk2 as a component of genetic pest management of fall armyworm, Spodoptera frugiperda (J. E. Smith) (Lepidoptera: Noctuidae).
Invasive insect pests, currently, pose a serious economic threat to several staple crops all over the world, one such being the fall armyworm, Spodoptera frugiperda. It was first observed in Africa since 2016, outside of its natural habitat in the Americas. Subsequently, it invaded several countries in South and South East Asia and also very recently in Australia. In all the newly invaded regions, maize is the principal crop attacked causing a serious economic concern to the poor farmers, particularly in the developing countries. Owing to the innate genetic ability, it defies many of the management options that include insecticides, Bt transgenics, and so forth. This is due to its high mobility, polyphagy and ability for quick development of resistance to several classes of insecticides. At this critical juncture, CRISPR/Cas9 mediated genome editing has shown a lot of promise in developing a novel area-wide pest management strategy called precision-guided sterile insect technique (pgSIT). pgSIT was initially demonstrated in Drosophila melanogaster which holds a greater promise for the environmentally friendly management of several globally significant agricultural pests such as S. frugiperda. Therefore, before developing both sgRNA and Cas9 transgenic lines, we have validated the target gene such as tssk2 through a non-transgenic approach by microinjecting ribo nucleo protein complex (Cas9 protein and tssk2 sgRNA) into G eggs of S. frugiperda. In the current investigation, we have obtained five edited males with distinct mutations which were further used for crossing studies to ascertain the effect of tssk2 editing affecting egg hatchability.
Topics: Animals; Spodoptera; CRISPR-Cas Systems; Male; Pest Control, Biological; Gene Editing; Spermatogenesis; Insect Proteins; Female; Insect Control
PubMed: 38783691
DOI: 10.1002/arch.22121 -
Insect Science May 2024Vitellogenin receptor (VgR) plays a crucial role in oogenesis by mediating endocytosis of vitellogenin and a portion of the yolk proteins in many insect species....
Vitellogenin receptor (VgR) plays a crucial role in oogenesis by mediating endocytosis of vitellogenin and a portion of the yolk proteins in many insect species. However, the function of VgR in minute parasitoid wasps is largely unknown. Here, we applied Trichogramma dendrolimi, a minute egg parasitoid, as a study model to investigate the function of VgR in parasitoids. We developed RNA interference (RNAi) methods based on microinjection of prepupae in T. dendrolimi. RNAi employs nanomaterial branched amphipathic peptide capsules (BAPC) as a carrier for double-stranded RNA (dsRNA), significantly enhancing delivery efficiency. Also, artificial hosts without medium were used to culture the injected prepupae in vitro. Utilizing these methods, we found that ovarian growth was disrupted after knockdown of TdVgR, as manifested by the suppressed development of the ovariole and the inhibition of nurse cell internalization by oocytes. Also, the initial mature egg load in the ovary was significantly reduced. Notably, the parasitic capacity of the female adult with ovarian dysplasia was significantly decreased, possibly resulting from the low availability of mature eggs. Moreover, ovarian dysplasia in T. dendrolimi caused by VgR deficiency are conserved despite feeding on different hosts. The results confirmed a critical role of TdVgR in the reproductive ability of T. dendrolimi and provided a reference for gene functional studies in minute insects.
PubMed: 38783625
DOI: 10.1111/1744-7917.13385 -
Addiction Biology May 2024Addictive properties of propofol have been demonstrated in both humans and animals. The nucleus accumbens (NAc) shell (NAsh) in the brain, along with the interactions...
Addictive properties of propofol have been demonstrated in both humans and animals. The nucleus accumbens (NAc) shell (NAsh) in the brain, along with the interactions between N-methyl-D-aspartate receptor (NMDAR) and the dopamine D1 receptor (D1R), as well as their downstream ERK/CREB signalling pathway in the NAc, are integral in regulating reward-seeking behaviour. Nevertheless, it remains unclear whether NMDARs and the NMDAR-D1R/ERK/CREB signalling pathway in the NAsh are involved in mediating propofol addiction. To investigate it, we conducted experiments with adult male Sprague-Dawley rats to establish a model of propofol self-administration behaviour. Subsequently, we microinjected D-AP5 (a competitive antagonist of NMDARs, 1.0-4.0 μg/0.3 μL/site) or vehicle into bilateral NAsh in rats that had previously self-administered propofol to examine the impact of NMDARs within the NAsh on propofol self-administration behaviour. Additionally, we examined the protein expressions of NR2A and NR2B subunits, and the D1R/ERK/CREB signalling pathways within the NAc. The results revealed that propofol administration behaviour was enhanced by D-AP5 pretreatment in NAsh, accompanied by elevated expressions of phosphorylation of NR2A (Tyr1246) and NR2B (Tyr1472) subunits. There were statistically significant increases in the expressions of D1Rs, as well as in the phosphorylated ERK1/2 (p-ERK1/2) and CREB (p-CREB). This evidence substantiates a pivotal role of NMDARs in the NAsh, with a particular emphasis on the NR2A and NR2B subunits, in mediating propofol self-administration behaviour. Furthermore, it suggests that this central reward processing mechanism may operate through the NMDAR-D1R/ERK/CREB signal transduction pathway.
Topics: Animals; Nucleus Accumbens; Propofol; Receptors, N-Methyl-D-Aspartate; Male; Rats, Sprague-Dawley; Receptors, Dopamine D1; Self Administration; Rats; Signal Transduction; Cyclic AMP Response Element-Binding Protein; MAP Kinase Signaling System
PubMed: 38782631
DOI: 10.1111/adb.13401 -
American Journal of Hypertension May 2024In the hypothalamic paraventricular nucleus (PVN) of spontaneously hypertensive rats (SHRs), the expression of Testis specific protein, Y-encoded-like 2 (TSPYL2) and the...
BACKGROUND
In the hypothalamic paraventricular nucleus (PVN) of spontaneously hypertensive rats (SHRs), the expression of Testis specific protein, Y-encoded-like 2 (TSPYL2) and the phosphorylation level of Janus kinase 2 (JAK2)/signal transducer and activator of transcription 3 (STAT3) are higher comparing with the normotensive Wistar-Kyoto rats (WKY). But how they are involved in hypertension remains unclear. TSPYL2 may interact with JAK2/STAT3 in PVN to sustain the high blood pressure during hypertension.
METHODS
Knockdown of TSPYL2 via adeno-associated virus (AAV) carrying shRNA was conducted through bilateral micro-injection into the PVN of SHR and WKY rats. JAK2/STAT3 inhibition was achieved by intraperitoneally or PVN injection of AG490 into the SHRs. Blood pressure (BP), plasma norepinephrine (NE), PVN inflammatory response, and PVN oxidative stress were measured.
RESULTS
TSPYL2 knock-down in the PVN of SHRs but not WKYs led to reduced BP and plasma NE, and deactivation of JAK2/STAT3, decreased expression of pro-inflammatory cytokine IL-1β, and increased expression of anti-inflammatory cytokine IL-10 in the PVN. Meanwhile, AG490 administrated in both ways reduced the blood pressure in the SHRs and deactivated JAK2/STAT3 but failed to change the expression of TSPYL2 in PVN. AG490 also downregulated expression of IL-1β and upregulated expression of IL-10. Both knockdown of TSPYL2 and inhibition of JAK2/STAT3 can reduce the oxidative stress in the PVN of SHRs.
CONCLUSION
JAK2/STAT3 is regulated by TSPYL2 in the PVN of SHRs, and PVN TSPYL2/JAK2/STAT3 is essential for maintaining high blood pressure in the hypertensive rats, making it a potential therapeutic target for hypertension.
PubMed: 38782571
DOI: 10.1093/ajh/hpae067 -
International Journal of Pharmaceutics Jun 2024There is a growing and urgent need for developing novel biomaterials and therapeutic approaches for efficient wound healing. Microneedles (MNs), which can penetrate... (Review)
Review
There is a growing and urgent need for developing novel biomaterials and therapeutic approaches for efficient wound healing. Microneedles (MNs), which can penetrate necrotic tissues and biofilm barriers at the wound and deliver active ingredients to the deeper layers in a minimally invasive and painless manner, have stimulated the interests of many researchers in the wound-healing filed. Among various materials, polymeric MNs have received widespread attention due to their abundant material sources, simple and inexpensive manufacturing methods, excellent biocompatibility and adjustable mechanical strength. Meanwhile, due to the unique properties of nanomaterials, the incorporation of nanomaterials can further extend the application range of polymeric MNs to facilitate on-demand drug release and activate specific therapeutic effects in combination with other therapies. In this review, we firstly introduce the current status and challenges of wound healing, and then outline the advantages and classification of MNs. Next, we focus on the manufacturing methods of polymeric MNs and the different raw materials used for their production. Furthermore, we give a summary of polymeric MNs incorporated with several common nanomaterials for chronic wounds healing. Finally, we discuss the several challenges and future prospects of transdermal drug delivery systems using nanomaterials-based polymeric MNs in wound treatment application.
Topics: Wound Healing; Needles; Humans; Polymers; Animals; Nanostructures; Drug Delivery Systems; Administration, Cutaneous; Microinjections
PubMed: 38782153
DOI: 10.1016/j.ijpharm.2024.124247 -
Journal of Controlled Release :... Jul 2024Microneedle patches have been developed as favorable platforms for delivery systems, such as the locoregional application of therapeutic drugs, and implantation systems,...
Microneedle patches have been developed as favorable platforms for delivery systems, such as the locoregional application of therapeutic drugs, and implantation systems, such as electronic devices on visceral tissue surfaces. However, the challenge lies in finding materials that can achieve both biocompatibility and stable fixation on the target tissue. To address this issue, utilizing a biocompatible adhesive biomaterial allows the flat part of the patch to adhere as well, enabling double-sided adhesion for greater versatility. In this work, we propose an adhesive microneedle patch based on mussel adhesive protein (MAP) with enhanced mechanical strength via ultraviolet-induced polyacrylate crosslinking and Coomassie brilliant blue molecules. The strong wet tissue adhesive and biocompatible nature of engineered acrylated-MAP resulted in the development of a versatile wet adhesive microneedle patch system for in vivo usage. In a mouse tumor model, this microneedle patch effectively delivered anticancer drugs while simultaneously sealing the skin wound. Additionally, in an application of rat subcutaneous implantation, an electronic circuit was stably anchored using a double-sided wet adhesive microneedle patch, and its signal location underneath the skin did not change over time. Thus, the proposed acrylated-MAP-based wet adhesive microneedle patch system holds great promise for biomedical applications, paving the way for advancements in drug delivery therapeutics, tissue engineering, and implantable electronic medical devices.
Topics: Animals; Needles; Drug Delivery Systems; Proteins; Microinjections; Rats, Sprague-Dawley; Transdermal Patch; Tissue Adhesives; Mice; Humans; Antineoplastic Agents; Male; Cell Line, Tumor; Rats; Female; Mice, Inbred BALB C; Skin; Adhesives; Acrylates
PubMed: 38782066
DOI: 10.1016/j.jconrel.2024.05.038 -
The Journal of Clinical and Aesthetic... May 2024Nose reshaping with hyaluronic acid (HA) fillers, also known as medical rhinoplasty, is an increasingly popular, minimally invasive aesthetic procedure. As the demand...
INTRODUCTION
Nose reshaping with hyaluronic acid (HA) fillers, also known as medical rhinoplasty, is an increasingly popular, minimally invasive aesthetic procedure. As the demand for nasal reshaping continues to rise, it is essential to develop safe and efficient injection techniques and assess satisfaction to ensure optimal outcomes and patient-centered care.
OBJECTIVE
This study aims to evaluate patient and physician satisfaction with hyaluronic acid filler applications using microinjection technique for nasal reshaping.
METHODS
The study included healthy adult patients who underwent medical rhinoplasty with the same HA filler using the microinjection technique. Patient satisfaction levels were evaluated at one and six months after the last injection using the Global Patient Satisfaction Scale (GPSS). Additionally, an independent dermatologist conducted a clinical evaluation for each patient by comparing before and after clinical pictures, using the Aesthetic Improvement Scale (AIS). Any side effects were recorded during each session and follow-up period for six months.
RESULTS
A total of 40 patients (37 women and 3 men) participated in the study. The most frequently targeted anatomical areas for filler injections were the nasal tip (100%), columella (100%), nasal prominence (100%), nasal dorsum (85%), and nasal root (82.5%). Injections distal to the nasolabial fold (NFL) were performed in 2.5 percent of patients. Patients expressed high satisfaction with the results at both one and six months after the procedure (mean GPSS, respectively; 4.65 and 4.47). Similarly, clinicians reported satisfaction with outcomes at the same time points (mean AIS, respectively; 1.7 and 1.4). Apart from mild pain during the procedure and transient erythema afterward, no side effects were recorded.
CONCLUSION
Medical rhinoplasty with HA fillers using the microinjection technique is an effective and reliable procedure. This technique provides safe and aesthetically pleasing results from both patient and dermatologist perspectives, making it a favorable option for nasal reshaping with HA dermal fillers.
PubMed: 38779374
DOI: No ID Found -
Heliyon May 2024To construct models of high-risk human papillomavirus (HPV) infection with precancerous lesions or cervical cancer and explore the immune function.
OBJECTIVE
To construct models of high-risk human papillomavirus (HPV) infection with precancerous lesions or cervical cancer and explore the immune function.
METHODS
Using CRISPR/Cas9, the expression vector -Rosa26 was microinjected into fertilized eggs of C57BL/6 N mice using homologous recombination, and the F0 generation was obtained for reproduction. Then, the formation of precancerous lesions was promoted via intramuscular injection of estradiol. Presence of precancerous cervical-vaginal intraepithelial lesions, Ki67 and p16 expression levels, and CD8 T cell proportions in the spleen were evaluated.
RESULTS
Two F0 generation mice exhibited correct the homologous recombination. Seven positive mice were identified in the F1 generation. After breeding and mating, 25 homozygous and 11 heterozygous -engineered mice were obtained from the F2 generation. After estradiol benzoate treatment, the cervical-vaginal epithelium appeared as precancerous lesions with positive Ki67 and p16 expression. The percentage of CD8 T cells decreased.
CONCLUSION
HPV16-E6-E7-Rosa26 induced low immune function in mice, and provides a good model for the basic research of the mechanisms of action of HPV infection-associated precancerous lesions or cervical cancer.
PubMed: 38765051
DOI: 10.1016/j.heliyon.2024.e29881 -
Yi Chuan = Hereditas May 2024Lesch-Nyhan syndrome (LNS) is a congenital defect disease that results in defective purine metabolism. It is caused by pathogenic variants of the HPRT gene. Its clinical...
Lesch-Nyhan syndrome (LNS) is a congenital defect disease that results in defective purine metabolism. It is caused by pathogenic variants of the HPRT gene. Its clinical symptoms mainly include high uric acid levels, gout, and kidney stones and damage. The mechanism of LNS has not been fully elucidated, and no cure exists. Animal models have always played an important role in exploring causative mechanisms and new therapies. This study combined CRISPR/Cas9 and microinjection to knock out the HPRT gene to create an LNS rabbit model. A sgRNA targeting exon 3 of HPRT gene was designed. Subsequently, Cas9 mRNA and sgRNA were injected into rabbit zygotes, and injected embryos were transferred to the uterus. The genotype and phenotype of rabbits were analyzed after birth. Four infant rabbits (named R1, R2, R3 and R4), which showed varying levels of gene modification, were born. The gene-editing efficiency was 100%. No wild-type sequences at the target HPRT gene were detected in R4 rabbit. Next, 6-thioguanine drug testing confirmed that HPRT enzymatic activity was deficient in R4 infant rabbit. HE staining revealed kidney abnormalities in all infant rabbits. Overall, an sgRNA capable of knocking out the HPRT gene in rabbits was successfully designed, and HPRT gene-modified rabbits were successfully constructed by using CRISPR/Cas9 technology and microinjection. This study provides a new nonrodent animal model for studying LNS syndrome.
Topics: Animals; Rabbits; Lesch-Nyhan Syndrome; Disease Models, Animal; Hypoxanthine Phosphoribosyltransferase; CRISPR-Cas Systems; Female; Gene Editing; RNA, Guide, CRISPR-Cas Systems; Male; Phenotype
PubMed: 38763775
DOI: 10.16288/j.yczz.24-012