Expression Of Specific Proteins Research Articles

Unveiling the native architecture of adult cardiac tissue using the 3D-NaissI method

Accurately imaging adult cardiac tissue in its native state is essential for regenerative medicine and understanding heart disease. Current fluorescence methods encounter challenges with tissue fixation. Here, we introduce the 3D-NaissI (3D-Native Tissue Imaging) method, which enables rapid, cost-effective imaging of fresh cardiac tissue samples in their closest native state, and has been extended to other tissues. We validated the efficacy of 3D-NaissI in preserving cardiac tissue integrity using small biopsies under hypothermic conditions in phosphate-buffered saline, offering unparalleled resolution in confocal microscopy for imaging fluorescent small molecules and antibodies. Compared to conventional histology, 3D-NaissI preserves cardiac tissue architecture and native protein epitopes, facilitating the use of a wide range of commercial antibodies without unmasking strategies. We successfully identified specific cardiac protein expression patterns in cardiomyocytes (CMs) from rodents and humans, including for the first time ACE2 localization in the lateral membrane/T-Tubules and SGTL2 in the sarcoplasmic reticulum. These findings shed light on COVID-19-related cardiac complications and suggest novel explanations for therapeutic benefits of iSGLT2 in HFpEF patients. Additionally, we challenge the notion of "connexin-43 lateralization” in heart pathology, suggesting it may be an artifact of cardiac fixation, as 3D-NaissI clearly revealed native connexin-43 expression at the lateral membrane of healthy CMs. We also discovered previously undocumented periodic ring-like 3D structures formed by pericytes that cover the lateral surfaces of CMs. These structures, positive for laminin-2, delineate a specific spatial architecture of laminin-2 receptors on the CM surface, underscoring the pivotal role of pericytes in CM function. Lastly, 3D-NaissI facilitates the mapping of native human protein expression in fresh cardiac autopsies, offering insights into both pathological and non-pathological contexts. Therefore, 3D-NaissI provides unparalleled insights into native cardiac tissue biology and holds the promise of advancing our understanding of physiology and pathophysiology, surpassing standard histology in both resolution and accuracy.

Regional Gene Therapy for Bone Tissue Engineering: A Current Concepts Review

The management of segmental bone defects presents a complex reconstruction challenge for orthopedic surgeons. Current treatment options are limited by efficacy across the spectrum of injury, morbidity, and cost. Regional gene therapy is a promising tissue engineering strategy for bone repair, as it allows for local implantation of nucleic acids or genetically modified cells to direct specific protein expression. In cell-based gene therapy approaches, a variety of different cell types have been described including mesenchymal stem cells (MSCs) derived from multiple sources—bone marrow, adipose, skeletal muscle, and umbilical cord tissue, among others. MSCs, in particular, have been well studied, as they serve as a source of osteoprogenitor cells in addition to providing a vehicle for transgene delivery. Furthermore, MSCs possess immunomodulatory properties, which may support the development of an allogeneic “off-the-shelf” gene therapy product. Identifying an optimal cell type is paramount to the successful clinical translation of cell-based gene therapy approaches. Here, we review current strategies for the management of segmental bone loss in orthopedic surgery, including bone grafting, bone graft substitutes, and operative techniques. We also highlight regional gene therapy as a tissue engineering strategy for bone repair, with a focus on cell types and cell sources suitable for this application.

Robust peptide/RNA complexes prepared with microfluidic mixing for pulmonary delivery by nebulisation.

Small interfering RNA (siRNA) and messengerRNA (mRNA) have drawn considerable attention in recent years due to their ability to modulate the expression of specific disease-related proteins. However, it is difficult to find safe, robust, and effective RNA delivery systems suitable for pulmonary delivery to treat lung diseases. In this study, two cationic peptides, namely LAH4-L1 and PEG12KL4, were employed as non-viral vectors for siRNA and mRNA delivery. Four formulations (i.e. LAH4-L1/siRNA; PEG12KL4/siRNA; LAH4-L1/mRNA and PEG12KL4/mRNA) were investigated. Microfluidic mixing method was utilised to fabricate RNA complexes in a controllable and reproducible manner. Upon optimisation of the microfluidic mixing protocol, a vibrating mesh nebuliser was employed to aerosolise the RNA complexes, and their transfection efficiency was evaluated on A549 and BEAS-2B cells. Following nebulisation, inhalable mist was generated for all RNA formulations with mass median aerodynamic diameter below 5μm. Although the hydrodynamic particle sizes of the RNA complexes were significantly reduced to around 100nm after nebulisation regardless of the original size of the complexes prior to nebulisation, the RNA binding efficiency and the in vitro RNA transfection ability of all the peptide formulations were successfully preserved with no significant differences compared to the same system before nebulisation. The current result indicates that both LAH4-L1 and PEG12KL4 hold significant potential for future clinical application for pulmonary siRNA and mRNA delivery through nebulisation.

The influence of strain on human keratocyte redox homeostasis and behavior

Aims/Purpose: To study the influence of strain on the expression of specific proteins in human keratocytes and its consequential effects on keratocyte intracellular redox homeostasis and behavior.Methods: Strain was applied to human keratocytes using the Flexcell® Tension Systems. Proteomics and western blot were used to identify proteins regulated in response to strain. Immunofluorescence (IF) staining and live‐cell imaging were employed to monitor reactive oxygen species (ROS) production and mitochondrial membrane potential (ΔΨM). Expression of oxidative stress‐related protein, and its influence on nuclear factor kappa‐light‐chain‐enhancer of activated B cells (NF‐κB) nuclear translocation and keratocyte proliferation and migration, were assessed by western blot, IF staining and bromodeoxyuridine (BrdU) assay. Mouse injury models and keratocytes from keratoconus patients were used to assess how strain (intraocular pressure, IOP) influences downstream protein in vivo.Results: The expression of protein Cytochrome P450 1B1 (CYP1B1) and Aldehyde Dehydrogenase 3 Family Member A1 (ALDH3A1) was significantly upregulated in strained keratocytes. Increased CYP1B1 effectively suppressed H2O2‐induced ROS production and mitigated the dissipation of ΔΨM. ALDH3A1 expression was negatively related to ROS accumulation. When the expression of ALDH3A1 was knocked down, NF‐κB nuclear translocation was promoted, ultimately resulting in increased keratocyte proliferation and migration. Mice and keratoconus patients with increased corneal strain also showed elevated ALDH3A1.Conclusions: Corneal strain significantly upregulates the expression of CYP1B1 and ALDH3A1. Increased CYP1B1 helps maintain keratocyte intracellular redox homeostasis and subsequently regulates keratocyte proliferation and migration via ALDH3A1.

Functional assays for the diagnosis of chronic granulomatous disease.

Chronic granulomatous disease (CGD) is a rare immunodeficiency characterized by recurrent bacterial and fungal infections that are attributed to reduced production of reactive oxygen species (ROS) by a multi-component enzyme complex known as the phagocyte NADPH oxidase or NOX2. Presented in this review are descriptions of several assays that assess the production of ROS as well as assays that characterize the expression of specific proteins of NOX2.

What are the potential mechanisms of fatigue-induced skeletal muscle hypertrophy with low-load resistance exercise training?

High-load resistance exercise (>60% of 1-repetition maximum) is a well-known stimulus to enhance skeletal muscle hypertrophy with chronic training. However, studies have intriguingly shown that low-load resistance exercise training (RET) (≤60% of 1-repetition maximum) can lead to similar increases in skeletal muscle hypertrophy as compared to high-load RET. This has raised questions about the underlying mechanisms for eliciting the hypertrophic response with low-load RET. A key characteristic of low-load RET is performing resistance exercise to, or close to, task failure, thereby inducing muscle fatigue. The primary aim of this evidence-based narrative review is to explore whether muscle fatigue may act as an indirect or direct mechanism contributing to skeletal muscle hypertrophy during low-load RET. It has been proposed that muscle fatigue could indirectly stimulate muscle hypertrophy through increased muscle fibre recruitment, mechanical tension, ultrastructural muscle damage, the secretion of anabolic hormones, and/or alterations in the expression of specific proteins involved in muscle mass regulation (e.g., myostatin). Alternatively, it has been proposed that fatigue could directly stimulate muscle hypertrophy through the accumulation of metabolic by-products (e.g., lactate), and/or inflammation and oxidative stress. This review summarizes the existing literature eluding to the role of muscle fatigue as a stimulus for low-load RET-induced muscle hypertrophy and provides suggested avenues for future research to elucidate how muscle fatigue could mediate skeletal muscle hypertrophy.

USE OF GENE THERAPY IN THE TREATMENT OF ALZHEIMER’S DISEASE – LITERATURE REVIEW

Neurodegenerative diseases are the most common cause of dementia, affecting around 55 million elderly worldwide, Alzheimer’s Disease (AD) corresponds to 60% of all cases. These elderlies reveal memory loss, impairment of language and thought formulation. AD has no cure, only its symptoms are treated, which does not prevent the disease from progressing. Also, throughout the course of brain degeneration progress, drugs no longer show significant effects. For this reason, the set of changes in the DNA of individuals who suffer from the disease can provide an important biomarker that has been the target of studies involving gene therapy. This technique occurs through the correction of modified genes or site-specific modifications, which can be through DNA or RNA, intending to treat or prevent diseases and uses a viral or non-viral vector which helps deliver genetic material to the cell. In the following we searched for clinical studies that portrayed some advances in the treatment of AD using gene therapy techniques. Due to the ability to alter and induce the expression of specific proteins, gene therapy presents a promising future in the restoration and correction of the pathogenic mechanism.

Completely defined cell culture medium for advanced alveolar models

Abstract In vitro alveolar models are an important tool study respiratory physiology, investigate lung diseases, develop and test new therapies, and simulate inhalation exposure to environmental pollutants or therapeutic agents. A defined cell culture medium is essential for maintaining consistent and reliable in vitro models. In this study a defined cell culture medium for advanced alveolar models based on epithelial A549 cell line and endothelial EaHy926 cell line is developed. Cellular survival was proven by quantification of LDH release. It was shown that co-culture of the cells enhances cellular survival compared to the endothelial monoculture. The formation of a homogeneous monolayer on laminin coated membranes was confirmed by actin staining. Further, the expression of cell type specific proteins (endothelial: CD31, VE-cadherin; epithelial: ZO1 and E-cadherin) was shown. Overall, we demonstrated a completely defined cell culture medium that can be used for the setup of an advanced in vitro alveolar model for biomedical application like drug development or breath gas analysis.

Resveratrol-Loaded Pluronic Micelles Ameliorate Scopolamine-Induced Cognitive Dysfunction Targeting Acetylcholinesterase Activity and Programmed Cell Death.

Numerous experimental studies suggest the potential for resveratrol (RVT) to be useful in the Alzheimer's disease treatment, but its low bioavailability limits its application. This study aimed to assess the potential of resveratrol-loaded micelles as a neuronal delivery platform to protect rats from scopolamine-induced memory impairment. Resveratrol was incorporated into Pluronic micelles, and the effects of micellar (mRVT) and pure resveratrol (RVT) were compared in the model of scopolamine-induced dementia in male Wistar rats. Memory performance was assessed by a T maze test. The effect of the treatment on specific neurotransmitter levels and protein expression in the cortex and the hippocampus were evaluated biochemically. Our results revealed that the polymeric micelles were in nanoscale (approximately 33 nm) and reached 79% encapsulation efficiency. The treatment with mRVT demonstrated better spatial memory protective effect. The biochemical assays showed that mRVT in a dose of 10 mg/kg enhanced the effects of the pure drug in regard to noradrenalin neurotransmission and acetylcholinesterase inhibitory activity in the hippocampus. Furthermore, micellar resveratrol increased the cAMP-response element-binding protein expression in the cortex and hippocampus of rats as well as the Bcl2/BAX ratio, which indicated an anti-apoptotic effect in the experimental dementia model. In conclusion, our results indicated the potential of a micellar system loaded with resveratrol for neurodegenerative diseases treatment.

The Effects of Dexamethasone on Human Lens Epithelial Cells and the Analysis of Related Pathways with Transcriptome Sequencing.

The goal of this study was to investigate the effects of dexamethasone on human lens epithelial cells (HLECs) and the potential mechanisms. HLECs (HLE-B3) were cultured in vitro to assess the effects of dexamethasone on cell size at different concentrations. Immunofluorescence staining was used to detect specific protein expression in HLE-B3 cells. The cell size was observed using phase-contrast microscopy, and the length and area were quantitatively measured with ImageJ software for statistical analysis. Flow cytometry was used to verify these outcomes. The means of three groups were statistically analyzed using one-way analysis of variance, whereas the means of two groups were statistically analyzed with the parametric Student's t-test. Additionally, high-throughput transcriptome sequencing was performed to compare messenger RNA (mRNA) expression levels between different concentrations of dexamethasone treatment groups and the control group, to identify potential signaling pathways. Subsequently, we performed quantitative Polymerase Chain Reaction (qPCR), immunofluorescence staining, and molecular docking experiments on the key differentially expressed genes. Dexamethasone affected the size of HLE-B3 cells. Both 0.25 and 0.5 μmol/L dexamethasone increased cell length and area, exhibiting no significant difference between the two treatment groups. Flow cytometry showed that dexamethasone increased cell size and granularity, with 0.25 μmol/L dexamethasone leading to larger cell areas and higher intracellular granularity. High-throughput transcriptome sequencing revealed significant upregulation of lysophosphatidic acid receptor 1 (LPAR1) and the pathways related to the glucocorticoid (GC) receptor. Certain concentrations of dexamethasone impact the morphology and biological functions of HLECs. As a subtype of G protein-coupled receptors, LPAR1 on the cell membrane may interact with dexamethasone, affecting cell size and inhibiting autophagy via the phosphoinositide 3-kinase (PI3K)/protein kinase B (AKT)/mammalian target of rapamycin (mTOR) pathway. These discoveries offer crucial biological insights into how dexamethasone influences the morphology and function of HLECs and the pathogenesis of GC-induced cataracts, offering potential molecular targets for future therapeutic strategies.

Succinylation modification-mediated upregulation of Sp1 promotes hepatocellular carcinoma cell proliferation

Hepatocellular carcinoma (HCC) remains one of the most prevalent malignant tumors globally, characterized by high incidence and mortality rates. Despite ongoing research, the underlying molecular mechanisms of HCC development are not yet fully understood. Utilizing bioinformatic analysis, real-time quantitative reverse transcription polymerase chain reaction (RT-qPCR), and Western blot assays, we identified that the expression of specificity protein 1 (Sp1) was significantly elevated in HCC cells compared to normal cells. Knockdown of the Sp1 gene led to a marked reduction in the viability and clonogenic potential of HCC cells. Further investigation revealed that the succinylation level of Sp1 was also increased in HCC cells. The upregulation of Sp1 expression was attributed to its succinylation, mediated by KAT2A, with lysine (K)562 identified as the succinylation site. Additionally, KAT2A and Sp1 were found to influence the upregulation of mTOR phosphorylation. Collectively, these findings suggest that KAT2A-promoted succinylation of Sp1 enhances the proliferative capacity of HCC cells by activating the mTOR pathway, providing a theoretical foundation for potential therapeutic strategies against HCC.

Kerstinginone, a new flavanone derivative from Commiphora kerstingii Engl. (Burseraceae) with potent apoptosis-inducing activity and inhibition of AKT/mTOR signaling pathway in non-sensitive prostate cancer cells

Commiphora kerstingii Engl is a tree which is 20-30m in height and commonly called "ararrabi" in Hausa. It is found in the Sahelian region (Cameroon, Chad, and Nigeria) where it is utilized for the treatment of several ailments including cancer. This study was aimed at investigating the chemical constituents and cytotoxic effect of extracts and isolates from the stem barks and leaves of C. kerstingii. Using classical chromatography technique coupled with spectroscopic analysis and literature information, ten (10) compounds were isolated from C. kerstingii stem barks and leaves, out of which two [kerstingilactone (3) and kerstinginone (10)] were new. To evaluate their potential cytotoxic effect, the impact on cell viability, growth, and proliferation was assessed using MTT and CCK-8 assays. Cell death mechanisms were analyzed via flow cytometry, and Western blotting was utilized to examine the expression of specific regulatory proteins. Furthermore, anti-metastatic properties were investigated through assays on cell migration, adhesion, and chemotaxis. Among the tested compounds, 2 (Masticadienonic Acid) and 10 (kerstinginone) exhibited significant dose-dependent inhibition of PC3 and LNCaP cell growth. Compound 2 displayed optimal inhibitory effects within a concentration range of 10-40μg/mL, while compound 10 demonstrated potent growth inhibition at concentrations of 2.5-10μg/mL. Both compounds suppressed cell proliferation and the formation of clones. Specifically, compound 2 induced apoptosis solely in the androgen-sensitive LNCaP prostate cancer cells, whereas compound 10 induced a stronger and concentration-dependent apoptotic response in both PC3 and LNCaP cells, resulting in approximately 50-70% apoptotic cells. It also induced potent cell migration/invasion arrest at concentrations ranging from 2.5 to 5μg/mL and increased cell adhesion to the extracellular matrix. Kerstinginone exhibits potent cytotoxicity and apoptosis-inducing activity, making it a promising lead for discovering a new anticancer drug.

Leveraging deep learning for identification and segmentation of “CAF-1/p60-positive” nuclei in oral squamous cell carcinoma tissue samples

In the current study, we introduced a unique method for identifying and segmenting oral squamous cell carcinoma (OSCC) nuclei, concentrating on those predicted to have significant CAF-1/p60 protein expression. Our suggested model uses the StarDist architecture, a deep-learning framework designed for biomedical image segmentation tasks. The training dataset comprises painstakingly annotated masks created from tissue sections previously stained with hematoxylin and eosin (H&E) and then restained with immunohistochemistry (IHC) for p60 protein. Our algorithm uses subtle morphological and colorimetric H&E cellular characteristics to predict CAF-1/p60 IHC expression in OSCC nuclei. The StarDist-based architecture performs exceptionally well in localizing and segmenting H&E nuclei, previously identified by IHC-based ground truth. In summary, our innovative approach harnesses deep learning and multimodal information to advance the automated analysis of OSCC nuclei exhibiting specific protein expression patterns. This methodology holds promise for expediting accurate pathological assessment and gaining deeper insights into the role of CAF-1/p60 protein within the context of oral cancer progression.

Time dependent changes in protein expression induced by intermittent theta burst stimulation in a cell line.

Intermittent Theta Burst Stimulation (iTBS), a non-invasive brain stimulation technique, is recognized for its ability to modulate cortical neuronal activity. However, its effects over time and the dynamics following stimulation are less well understood. Understanding the temporal dynamics of iTBS effects is essential for optimizing the timing and frequency of stimulation in therapeutic applications. This study investigated the temporal changes in protein expression induced by iTBS in Neuro-2a cells. We analyzed protein expression in retinoic acid-differentiated Neuro-2a cells at multiple time points - 0.5, 3, 6, 12, and 24 hours post-iTBS - using Western blot and immunocytochemistry techniques. Our findings reveal a significant early increase in neurotransmitter receptor subunits, neurotrophic factors, and cytoskeletal proteins within the first 0.5 hour following iTBS. Notably, proteins such as mGLuR1, NMDAR1, GABBR2, and β-tubulin III showed substantial increase in expression. However, the effects of iTBS on protein expression was not sustained at later timepoints. Our results suggest that iTBS can transiently alter the expression of specific proteins in Neuro-2a cells. Future research should investigate the potential benefits of repeated stimulations within the early time window to refine iTBS interventions, potentially expanding their research and clinical applications.

Innate immune response in COVID-19: single-cell multi-omics profile of NK lymphocytes in a clinical case series

BackgroundCOVID-19 pandemic caused by the Severe Acute Respiratory Syndrome-Coronavirus-2 (SARS-CoV-2) represents the biggest global health emergency in recent decades. The host immune response to SARS-CoV-2 seems to play a key role in disease pathogenesis and clinical manifestations, with Natural Killer (NK) lymphocytes being among the targets of virus-induced regulation.MethodsThis study performed a single-cell multi-omics analysis of transcripts and proteins of NK lymphocytes in COVID-19 patients, for the characterization of the innate immunological response to infection. NK cells were isolated from peripheral blood samples collected from adult subjects divided into 3 study groups: (1) non-infected subjects (Naïve group, n = 3), (2) post COVID-19 convalescent subjects (Healed group, n = 3) and (3) patients that were vaccinated against SARS-CoV-2 (Vaccine group, n = 3). Cells were then analysed by the BD Rhapsody System for the single-cell multi-omics investigation of transcriptome and membrane proteins.ResultsThe bioinformatic analysis identified 5 cell clusters which differentially expressed gene/protein markers, defining NK cell subsets as “Active NK cells” and “Mature NK cells”. Calculating the relative proportion of each cluster within patient groups, more than 40% of the Naïve group cell population was found to belong to Mature NKs, whereas more than 75% of the Vaccine group cell population belonged to the cluster of Active NKs. Regarding the Healed group, it seemed to show intermediate phenotype between Active and Mature NK cells. Differential expression of specific genes, proteins and signaling pathways was detected comparing the profile of the 3 experimental groups, revealing a more activated NK cell phenotype in vaccinated patients versus recovered individuals.ConclusionsThe present study detected differential expression of NK cell markers in relation to SARS-CoV-2 infection and vaccine administration, suggesting the possibility to identify key molecular targets for clinical-diagnostic use of the individual response to viral infection and/or re-infection.

Differential effect of plakoglobin in restoring the tumor suppressor activities of p53-R273H vs. p53-R175H mutants.

The six most common missense mutations in the DNA binding domain of p53 are known as "hot spots" and include two of the most frequently occurring p53 mutations (p53-R175H and p53-R273H). p53 stability and function are regulated by various post-translational modifications such as phosphorylation, acetylation, sumoylation, methylation, and interactions with other proteins including plakoglobin. Previously, using various carcinoma cell lines we showed that plakoglobin interacted with wild-type and several endogenous p53 mutants (e.g., R280K, R273H, S241F, S215R, R175H) and restored their tumor suppressor activities in vitro. Since mutant p53 function is both mutant-specific and cell context-dependent, we sought herein, to determine if plakoglobin tumor suppressive effects on exogenously expressed p53-R273H and p53-R175H mutants are similarly maintained under the same genetic background using the p53-null and plakoglobin-deficient H1299 cell line. Functional assays were performed to assess colony formation, migration, and invasion while immunoblotting and qPCR were used to examine the subcellular distribution and expression of specific proteins and genes that are typically regulated by or regulate p53 function and are altered in mutant p53-expressing cell lines and tumors. We show that though, plakoglobin interacted with both p53-R273H and p53-R175H mutants, it had a differential effect on the transcription and subcellular distribution of their gene targets and their overall oncogenic properties in vitro. Notably, we found that plakoglobin's tumor suppressive effects were significantly stronger in p53-R175H expressing cells compared to p53-R273H cells. Together, our results indicate that exploring plakoglobin interactions with p53-R175H may be useful for the development of cancer therapeutics focused on the restoration of p53 function.

BMSCs-derived exosomes carrying miR-668-3p promote progression of osteoblasts in osteonecrosis of the femoral head: Expression of proteins CD63 and CD9

Recently, exosomes that are derived from bone marrow mesenchymal stem cells (BMSCs) have garnered considerable interest due to their significant roles in the processes of bone regeneration and repair. Among the various molecular components present within these exosomes, miR-668-3p has emerged as a pivotal microRNA that may be instrumental in modulating the function and proliferation of osteoblasts, the cells responsible for bone formation. The primary objective of this research was to examine the enhancing effects of BMSC-derived exosomes that are enriched with miR-668-3p on the advancement of osteoblasts in the context of osteonecrosis of the femoral head. Furthermore, the study aimed to analyze how the expression of specific exosomal proteins, namely CD63 and CD9, influences this biological process. To conduct the investigation, BMSCs were isolated from healthy rat models, followed by the extraction of their secreted exosomes. The subsequent phase of the study involved assessing the proliferation and differentiation of osteoblasts by introducing the exosomes enriched with miR-668-3p into an experimental setup representing osteonecrosis of the femoral head. The findings revealed that exosomes derived from BMSCs, which contained miR-668-3p, significantly enhanced the proliferation of osteoblasts as well as the expression of key osteogenic marker genes. Notably, the levels of CD63 and CD9 proteins were markedly increased in the treated groups, indicating that the mechanisms underlying this promotion might involve cell adhesion and the endocytic uptake of exosomes.

Region of interest determination algorithm of lensless calcium imaging datasets.

Advances in fluorescence imaging technology have been crucial to the progress of neuroscience. Whether it was specific expression of indicator proteins, detection of neurotransmitters, or miniaturization of fluorescence microscopes, fluorescence imaging has improved upon electrophysiology, the gold standard for monitoring brain activity, and enabled novel methods to sense activity in the brain. Hence, we developed a lightweight and compact implantable CMOS-based lensless Ca2+ imaging device for freely moving transgenic G-CaMP mouse experiments. However, without a lens system, determination of regions of interest (ROI) has proven challenging. Localization of fluorescence activity and separation of signal from noise are difficult. In this study, we report an ROI selection method using a series of adaptive binarizations with a gaussian method and morphological image processing. The parameters for each operation such as the kernel size, sigma and footprint size were optimized. We then validated the utility of the algorithm with simulated data and freely moving nociception experiments using the lensless devices. The device was implanted in the dorsal raphe nucleus to observe pain-related brain activity following a formalin test to stimulate pain. We observed significant increases in fluorescence activity after formalin injection compared to the control group when using the ROI determination algorithm.

Research on the biophysical properties of circulating tumor cells

This article reviews the biophysical properties of circulating tumor cells (CTCs) and their significance in tumor metastasis and liquid biopsy. CTCs are tumor cells that have shed from solid tumors or metastatic foci and entered the bloodstream, possessing various biological markers. They may be cleared by the immune system, enter a dormant state, or form clusters to facilitate metastasis. The physical properties of CTCs, including cell size, density, rigidity, and dielectric properties, differ from normal blood cells, providing a theoretical basis for separation and detection. The biological characteristics of CTCs include epithelial cell properties, expression of specific surface marker proteins, and the characteristics of epithelial-mesenchymal transition, which are crucial for the aggressiveness and metastatic potential of tumors. The development of CTC detection technologies, such as the Cell Search system, has been clinically applied, but the heterogeneity of tumor cells remains a major challenge in research. The development of composite separation technologies that combine various properties of CTCs is expected to improve the sensitivity, specificity, and efficiency of CTC detection, thereby playing an important role in the personalized treatment and prognostic assessment of tumors.

The order of green and red LEDs irradiation affects the neural differentiation of human umbilical cord matrix-derived mesenchymal cells

Different wavelengths emitted from light-emitting diodes (LEDs) are known as an influential factor in proliferation and differentiation of various cell types. Since human umbilical cord matrix-derived mesenchymal cells (hUCMs) are ideal tools for human regenerative medicine clinical trials and stem cell researches, in the present study we investigated the neurogenesis effects of single and intermittent green and red LED irradiation on hUCM cells. Exposure of hUCMs to single and intermittent green (530 nm, 1.59 J/cm2) and red (630 nm, 0.318 J/cm2) lights significantly increased the expression of specific genes including nestin, β-tubulin III and Olig2. Additionally, immunocytochemical analysis confirmed the expression of specific neural-related proteins including nestin, β-tubulin III, Olig2 and GFAP. Also, alternating exposure of hUCM cells to green and red lights increased the expression of some neural markers more than either light alone. Further research are required to develop the application of LED irradiation as a useful tool for therapeutic purposes including neural repair and regeneration.