Search Results (928)

Abstract: Cell surface signaling (CSS) is a means of rapidly adjusting transcription in response to extracellular stimuli in Gram-negative bacteria. The pseudobactin BN7/8 uptake (Pup) system not only imports iron but also upregulates its own transcription through CSS in Pseudomonas capeferrum. In the absence of ferric pseudobactin BN7/8, the signaling components are maintained in a resting state via the formation of a periplasmic complex between the N-terminal signaling domain (NTSD) of the outer membrane iron-transporter, PupB, and the C-terminal CSS domain (CCSSD) of the sigma regulator, PupR. The previously determined 1.6 Å crystal structure of this periplasmic complex has allowed us to probe the structural and thermodynamic consequences of mutating key interfacial residues. In this report, we describe the solution structure of the PupB NTSD and use Nuclear Magnetic Resonance spectroscopy, Isothermal Titration Calorimetry, and Circular Dichroism spectroscopy together with thermal denaturation to investigate whether three PupB point mutations, Q69K, H72D, and L74A, influence the interaction merely due to the chemical nature of the amino acid substitution or also cause changes in overall protein structure. Our results demonstrate that binding to the PupR CCSSD does not alter the structure of PupB NTSD and that the individual mutations have only minor effects on structure. The mutations generally lower thermodynamic stability of the NTSD and weaken binding to the CCSSD. These findings validate the X-ray crystal structure interface, emphasizing the importance of amino acid chemical nature at the interface. Full article

The control of metabolic networks is incompletely understood, even for glycolysis in highly studied model organisms. Direct real-time observations of metabolic pathways can be achieved in cellular systems with ¹³C NMR using dissolution Dynamic Nuclear Polarization (dDNP NMR). The method relies on a short-lived boost of NMR sensitivity using a redistribution of nuclear spin states to increase the alignment of the magnetic moments by more than four orders of magnitude. This temporary boost in sensitivity allows detection of metabolism with sub-second time resolution. Here, we hypothesized that dDNP NMR would be able to investigate molecular phenotypes that are not easily accessible with more conventional methods. The use of dDNP NMR allows real-time insight into carbohydrate metabolism in a Gram-positive bacterium (Lactoccocus lactis), and comparison to other bacterial, yeast and mammalian cells shows differences in the kinetic barriers of glycolysis across the kingdoms of life. Nevertheless, the accumulation of non-toxic precursors for biomass at kinetic barriers is found to be shared across the kingdoms of life. We further find that the visualization of glycolysis using dDNP NMR reveals kinetic characteristics in transgenic strains that are not evident when monitoring the overall glycolytic rate only. Finally, dDNP NMR reveals that resting Lactococcus lactis cells use the influx of carbohydrate substrate to produce acetoin rather than lactate during the start of glycolysis. This metabolic regime can be emulated using suitably designed substrate mixtures to enhance the formation of the C4 product acetoin more than 400-fold. Overall, we find that dDNP NMR provides analytical capabilities that may help to clarify the intertwined mechanistic determinants of metabolism and the optimal usage of biotechnologically important bacteria. Full article

Bacterial superantigens are T-cell-stimulatory protein molecules which produce massive cytokines and cause human diseases. Due to their ability to activate up to 20% of resting T-cells, they have effectively killed T-cell-dependent tumours in vivo. However, the intrinsic toxicity of whole SAg molecules highlights the urgent need to develop more effective and safer SAg-based immunotherapy. With its unique approach, our study is a significant step towards developing safer tumour-targeted superantigen peptides (TTSP). We identified the T-cell activation function regions on the SEA superantigen and produced variants with minimal lethality, ensuring a safer approach to cancer treatment. This involved the creation of twenty 50-amino-acid-long overlapping peptides covering the full-length SEA superantigen (P1-P20). We then screened these peptides for T-cell activation, successfully isolating two peptides (P5 and P15) with significant T-cell activation. These selected peptides were used to design and synthesise tumour-targeted superantigen peptides, which were linked to a cancer-specific third loop (L3) of transforming growth factor-α (TGF-α), TGFαL3 from either a C’ or N’ terminal with an eight-amino-acid flexible linker in between. We also produced several P15 variants by changing single amino acids or by amino acid deletions. The novel molecules were then investigated for cytokine production and tumour-targeted killing. The findings from our previous study and the current work open up new avenues for peptide-based immunotherapy, particularly when combined with other immunotherapy techniques, thereby ensuring effective and safer cancer treatment. Full article

The blooms of Alexandrium catenella, the main producer of paralytic shellfish toxins worldwide, have become the main threat to coastal activities in Southern Chile, such as artisanal fisheries, aquaculture and public health. Here, we explore retrospective data from an intense Paralytic Shellfish Poisoning outbreak in Southern Chile in Summer–Autumn 2016, identifying environmental drivers, spatiotemporal dynamics, and detoxification rates of the main filter-feeder shellfish resources during an intense A. catenella bloom, which led to the greatest socio-economic impacts in that area. Exponential detoxification models evidenced large differences in detoxification dynamics between the three filter-feeder species surf clam (Ensis macha), giant barnacle (Austromegabalanus psittacus), and red sea squirt (Pyura chilensis). Surf clam showed an initial toxicity (9054 µg STX-eq·100 g⁻¹) around 10-fold higher than the other two species. It exhibited a relatively fast detoxification rate and approached the human safety limit of 80 µg STX-eq·100 g⁻¹ towards the end of the 150 days. Ecological implications and future trends are also discussed. Based on the cell density evolution, data previously gathered on the area, and the biology of this species, we propose that the bloom originated in the coastal area, spreading offshore thanks to the resting cysts formed and transported in the water column. Full article

The improper disposal of plastic waste has become a significant problem, with only a small amount recycled and the rest ending up in landfills or being burned, leading to environmental pollution. In addition, the cost of electric energy has risen by over 100% in the last 20 years, making it unaffordable for remote areas to access this service due to high installation costs, leaving people living far from major cities without electricity. This study proposes an innovative solution to these issues using microbial fuel cell (MFC) technology to simultaneously reduce plastic waste and generate electric energy by utilizing the fungus Aspergillus sp. As a substrate for 45 days. The MFCs reached maximum values of 0.572 ± 0.024 V and 3.608 ± 0.249 mA of voltage and electric current on the thirty-first day, with the substrate operating at a pH of 6.57 ± 0.27 and an electrical conductivity of 257.12 ± 20.9 mS/cm. Furthermore, it was possible to reduce the chemical oxygen demand by 73.77% over the 45 days of MFC operation, while the recorded internal resistance was 27.417 ± 9.810 Ω, indicating a power density of 0.124 ± 0.006 mW/cm². The initial and final transmittance spectra, obtained using FTIR (Fourier Transform Infrared), showed the characteristic peaks of polyethylene (plastic), with a noticeable reduction in the final spectrum, particularly in the vibration of the C-H compound. After 45 days of fungus operation, the plastic surface used as a sample exhibited perforations and cracks, resulting in a thickness reduction of 313.56 µm. This research represents an initial step in using fungi for plastic reduction and electric energy generation in an alternative and sustainable manner. Full article

Background and Objectives: Urinary incontinence (UI) is the involuntary loss of urine caused by a weakness in the pelvic floor muscles (PFMs) that affects urethral closure. Myostatin, which prevents the growth of muscles, is a protein expressed by human skeletal muscle cells. Indeed, it has been observed that myostatin concentration rises during skeletal muscle inactivity and that suppressing serum myostatin promotes muscle growth and strength. Furthermore, therapeutic interventions that reduce myostatin signalling may lessen the effects of aging on skeletal muscle mass and function. For this reason, the aim of the study was to assess if flat magnetic stimulation technology affects serum myostatin levels, as myostatin can block cell proliferation at the urethral sphincter level. Materials and Methods: A total of 19 women, 75% presenting stress urinary incontinence (SUI) and 25% urgency urinary incontinence (UUI), were enrolled. A non-invasive electromagnetic therapeutic system designed for deep pelvic floor area stimulation was used for eight sessions. Results: The ELISA (enzyme linked immunosorbent assay) test indicated that the myostatin levels in blood sera had significantly decreased. Patients’ ultrasound measurements showed a significant genital hiatus length reduction at rest and in a stress condition. The Pelvic Floor Bother Questionnaire consistently revealed a decrease in mean scores when comparing the pre- and post-treatment data. Conclusions: Effective flat magnetic stimulation reduces myostatin concentration and genital hiatus length, minimizing the severity of urinary incontinence. The results of the study show that without causing any discomfort or unfavourable side effects, the treatment plan significantly improved the PFM tone and strength in patients with UI. Full article

Resveratrol has been found to stimulate the growth of human dermal papilla cells (hDPCs) and protect them from oxidative damage. The topical application of resveratrol effectively enhanced hair growth and stimulated the transition of the hair cycle from the resting phase (telogen) into the active growth phase (anagen phase). The aim of this study was to examine the effects of resveratrol-rich peanut callus and sprout extracts on promoting hair growth and anti-hair loss effects in hDPCs and then select the most effective extract (callus) for further assessment in clinical applications. The peanut sprout and callus were produced from plant seeds and extracted with ethanol. Resveratrol was identified as the major constituent in the extracts obtained using the HPLC approach, although no resveratrol was found in the original seed extract. Hair cell proliferation, growth factor expression, and 5α-reductase (5-AR) inhibition assays were used to examine the extracts’ promotion of hair growth and inhibition of anti-hair loss in an in vitro hDPCs model, and standard resveratrol served as a positive reference. Initially, the cytotoxicity of the samples was assessed, revealing that 2.5% of all extracts and 0.02% of resveratrol exhibited a non-cytotoxic effect to hDPCs. The 50% cytotoxic concentration of peanut callus extract (15.50 ± 0.05%) was significantly less cytotoxic to hDPCs than sprout (12.30 ± 0.14%) and seed (11.69 ± 0.26%) extracts (p < 0.05). In vitro experiments indicated that peanut callus extract had the greatest proliferated hDPCs (20.44 ± 0.78%), increased the expression of growth factors (IGF-1, KGF, and HGF) and inhibited 5AR (27.83 ± 2.48%). Then, the callus extract and resveratrol as the positive standard were separately incorporated into a hair serum base and assessed for anti-hair loss effect in a randomized, double-blind, placebo-controlled clinical trial on 60 subjects, and the daily hair shedding count, one-minute combing, and forcible hair pluck tests were monitored for 12 weeks. The clinical study showed that the hair serum containing peanut callus extract and resveratrol were effective in reducing hair shedding (66.98% and 61.47%), decreased hair fall from combing by (83.92% and 68.19%), and increased the ratio of the anagen/telogen by (3.3 and 2.8, respectively). The results suggest that peanut callus extract has a significant amount of resveratrol and the potential to stimulate hair growth and prevent hair loss, making it an alternative bioactive option for cosmetic applications. Full article

We developed a technique that can measure corneal transepithelial electrical resistance (TER) in mice, which was used for evaluating corneal toxicity induced by ophthalmic drugs. We used a tissue culture well and its insert to mount the mouse globe and separated the cornea from the rest of the globe to enable corneal TER measurements to be taken. The explanted mouse eyes were divided into groups, and the corneal epithelia were exposed to different concentrations of BAC. Half of these eyes were fixed for transmission electron microscopy (TEM) examination and the other for ZO-1 immunohistochemical (IHC) evaluation. After exposure to control, 0.1%, 0.2%, and 0.5% BAC, the TER was 100 ± 0%, 91 ± 14%, 83 ± 13%, and 34 ± 12% of the pre-exposure TER at 1 min, respectively, with a statistically significant decrease in the 0.5% group. After 3 min, the TER showed a statistically significant decrease in the 0.2% and 0.5% groups. The TEM examinations showed a loss of epithelial tight junctions between superficial cells in the 0.2% and 0.5% groups. The IHC examination showed decreased ZO-1 staining of the corneal epithelium of the same groups as compared to the control. To the best of our knowledge, we succeeded in developing an innovative technique for corneal TER measurement in mice. Full article

Postural Orthostatic Tachycardia Syndrome (POTS) affects up to 1% of the US population, predominantly women, and is characterized by a complex, elusive etiology and heterogeneous phenotypes. This review delves into the intricate physiology and etiology of POTS, decoding the roles of the sinoatrial node, the autonomic nervous system, fluid dynamics, and the interplay between the immune and endocrine systems. It further examines key contributing factors such as dysautonomia, thoracic hypovolemia, autonomic neuropathies, sympathetic denervation, autoimmune responses, and associations with conditions such as small-fiber neuropathy and mast cell activation syndrome. Given the numerous mysteries surrounding POTS, we also cautiously bring attention to sinoatrial node and myocardial function, particularly in how the heart responds to stress despite exhibiting a normal cardiac phenotype at rest. The potential of genomic research in elucidating the underlying mechanisms of POTS is emphasized, suggesting this as a valuable approach that is likely to improve our understanding of the genetic underpinnings of POTS. The review introduces a tentative classification system for the etiological factors in POTS, which seeks to capture the condition’s diverse aspects by categorizing various etiological factors and acknowledging co-occurring conditions. This classification, while aiming to enhance understanding and optimize treatment targets, is presented as a preliminary model needing further study and refinement. This review underscores the ongoing need for research to unravel the complexities of POTS and to develop targeted therapies that can improve patient outcomes. Full article

One of the environmental problems that affects negatively orange productivity is drought because it greatly minimizes the growth attributes, photosynthetic process, water uptake, percentage of fruit set, and productivity; meanwhile, it raises the rates of respiration and transpiration, as well as the premature and preharvest fruit drop percentages. In addition, drought creates osmotic stress, affects the relationship between plants and water, reduces the amount of water in shoots, and prevents plant cell development and expansion. It is very important to search for a solution to minimize the effect of drought stress; therefore, the present study has investigated the effect of the application of humic acid (HA) at 0, 1 and 2 kg per tree and spraying of seaweed extract (SWE) at 0.2, 0.3 and 4% in combination with moringa leaf extract (MLE) at 2, 4 and 6%, respectively, on the productivity, fruit quality and nutritional status of navel orange cv. Washington during the 2022 and 2023 seasons. The results proved that the application of the biostimulants individually or in combination significantly positively changed the vegetative growth, productivity, fruit quality parameters and leaf mineral content of macro- and micronutrients of the treated trees compared to untreated trees. The superior treatments which gave the best results were 2 kg HA + 0.4% SWE + 6% MLE followed by 2 kg HA + 0.3% SWE + 4% MLE over the rest of the applied treatments. Full article

In the management of early-stage breast cancer (BC), lymph nodes (LNs) are typically characterised using the One-Step Nucleic Acid Amplification (OSNA) assay, a standard procedure for assessing subclinical metastasis in sentinel LNs (SLNs). The pivotal role of LNs in coordinating the immune response against BC is often overlooked. Our aim was to improve prognostic information provided by the OSNA assay and explore immune-related gene signatures in SLNs. The expression of an immune gene panel was analysed in SLNs from 32 patients with Luminal A early-stage BC (cT1-T2 N0). Using an unsupervised approach based on these expression values, this study identified two clusters, regardless of the SLN invasion: one evidencing an adaptive anti-tumoral immune response, characterised by an increase in naive B cells, follicular T helper cells, and activated NK cells; and another with a more undifferentiated response, with an increase in the activated-to-resting dendritic cells (DCs) ratio. Through a protein—protein interaction (PPI) network, we identified seven immunoregulatory hub genes: CD80, CD40, TNF, FCGR3A, CD163, FCGR3B, and CCR2. This study shows that, in Luminal A early-stage BC, SLNs gene expression studies enable the identification of distinct immune profiles that may influence prognosis stratification and highlight key genes that could serve as potential targets for immunotherapy. Full article

Combinatorial antiretroviral therapy (cART) has transformed HIV infection from a death sentence to a controllable chronic disease, but cannot eliminate the virus. Latent HIV-1 reservoirs are the major obstacles to cure HIV-1 infection. Previously, we engineered exosomal Tat (Exo-Tat) to reactivate latent HIV-1 from the reservoir of resting CD4+ T cells. Here, we present an HIV-1 eradication platform, which uses our previously described Exo-Tat to activate latent virus from resting CD4+ T cells guided by the specific binding domain of CD4 in interleukin 16 (IL16), attached to the N-terminus of exosome surface protein lysosome-associated membrane protein 2 variant B (Lamp2B). Cells with HIV-1 surface protein gp120 expressed on the cell membranes are then targeted for immune cytolysis by a BiTE molecule CD4-αCD3, which colocalizes the gp120 surface protein of HIV-1 and the CD3 of cytotoxic T lymphocytes. Using primary blood cells obtained from antiretroviral treated individuals, we find that this combined approach led to a significant reduction in replication-competent HIV-1 in infected CD4+ T cells in a clonal in vitro cell system. Furthermore, adeno-associated virus serotype DJ (AAV-DJ) was used to deliver Exo-Tat, IL16lamp2b and CD4-αCD3 genes by constructing them in one AAV-DJ vector (the plasmid was named pEliminator). The coculture of T cells from HIV-1 patients with Huh-7 cells infected with AAV-Eliminator viruses led to the clearance of HIV-1 reservoir cells in the in vitro experiment, which could have implications for reducing the viral reservoir in vivo, indicating that Eliminator AAV viruses have the potential to be developed into therapeutic biologics to cure HIV-1 infection. Full article

Background: The upper respiratory mucosa plays a crucial role in both the physical integrity and immunological function of the respiratory tract. However, in certain situations such as infections, trauma, or surgery, it might sustain damage. Tissue engineering, a field of regenerative medicine, has found applications in various medical fields including but not limited to plastic surgery, ophthalmology, and urology. However, its application to the respiratory system remains somewhat difficult due to the complex morphology and histology of the upper respiratory tract. To date, a culture protocol for producing a handleable, well-differentiated nasal mucosa has yet to be developed. The objective of this review is to describe the current state of research pertaining to cell culture techniques used for producing autologous healthy human upper respiratory cells and mucosal tissues, as well as describe its clinical applications. Methods: A search of the relevant literature was carried out with no time restriction across Embase, Cochrane, PubMed, and Medline Ovid databases. Keywords related to “respiratory mucosa” and “culture techniques of the human airway” were the focus of the search strategy for this review. The risk of bias in retained studies was assessed using the Joanna Briggs Institute’s (JBI) critical appraisal tools for qualitative research. A narrative synthesis of our results was then conducted. Results: A total of 33 studies were included in this review, and thirteen of these focused solely on developing a cell culture protocol without further use. The rest of the studies used their own developed protocol for various applications such as cystic fibrosis, pharmacological, and viral research. One study was able to develop a promising model for nasal mucosa that could be employed as a replacement in nasotracheal reconstructive surgery. Conclusions: This systematic review extensively explored the current state of research regarding cell culture techniques for producing tissue-engineered nasal mucosa. Bioengineering the nasal mucosa holds great potential for clinical use. However, further research on mechanical properties is essential, as the comparison of engineered tissues is currently focused on morphology rather than comprehensive mechanical assessments. Full article

This paper presents the concept of applying incremental capacity analysis (ICA) on the OCV curve in the SoC space. The OCV curve can be obtained from any sequence of discharge or charge current or power pulse with a necessary rest period to allow the cell to reach a pseudo-OCV after each pulse. With a high resolution (>100 pulses) in the full SoC window, an OCV-vs.-SoC curve can be obtained with sufficient accuracy to perform an ICA on the obtained OCV curve. ICA as a diagnostic technique has commonly been applied on Li-ion cells with constant charge and discharge at slow currents. However, a slow controlled constant current charge or discharge is normally not feasible and cannot be easily applied to a battery in an application. Here, we revisit pulse-based ICA to supplement the conventional constant-current-based technique. Based on actual ageing data, we show that ICA performed on a selection of high-resolution OCV curves is comparable or better than conventional ICA with constant current. The main advantage of OCV-ICA is that it can be applied to most cells and systems without a significant interruption of normal cell operation. OCV-ICA can provide valuable insights into ageing mechanisms as well as, e.g., detailed information on changes in internal resistance. Full article

Our previous work on an air-cooled stack of five pouch-format lithium-ion (Li-ion) cells showed that non-uniform temperature can cause accelerated degradation, especially of the middle cell. In this work, a stack of five similar cells was cycled at a higher C-rate and water-cooled to create a larger temperature gradient for comparison with the air-cooled stack. It was hypothesized that the larger temperature gradient in the water-cooled stack would exacerbate the degradation of the middle cell. However, the results showed that the middle cell degraded slightly slower than the side cells in the water-cooled stack. This trend is opposite to that in the air-cooled stack. This difference could be attributed to the combined effects of a smaller temperature rise and larger temperature gradient in the water-cooled stack than in the air-cooled stack. Post-mortem analysis of cycled cells and a fresh cell showed that the degradation mainly came from the anode. Increased lithium plating and decreased porosity in the side cells are possible mechanisms for the faster degradation compared with the middle cell. It was also found that all the cells in the water-cooled stack experienced a phenomenon of capacity drop and recovery after a low C-rate reference performance test and extended rest. This phenomenon can be attributed to lithium diffusion between the anode active area and the anode overhang area. Full article