Genetic Transformation of Plasmid DNA into Escherichia coli Using High Frequency Electromagnetic Energy.

We present a novel technique of genetic transformation of bacterial cells mediated by high frequency electromagnetic energy (HF EME). Plasmid DNA, pGLO (5.4 kb), was successfully transformed into Escherichia coli JM109 cells after exposure to 18 GHz irradiation at a power density between 5.6 and 30 kW m-2 for 180 s at temperatures ranging from 30 to 40 °C. Transformed bacteria were identified by the expression of green fluorescent protein (GFP) using confocal scanning microscopy (CLSM) and flow cytometry (FC). Approximately 90.7% of HF EME treated viable E. coli cells exhibited uptake of the pGLO plasmid. The interaction of plasmid DNA with bacteria leading to transformation was confirmed by using cryogenic transmission electron microscopy (cryo-TEM). HF EME-induced plasmid DNA transformation was shown to be unique, highly efficient, and cost-effective. HF EME-induced genetic transformation is performed under physiologically friendly conditions in contrast to existing techniques that generate higher temperatures, leading to altered cellular integrity. This technique allows safe delivery of genetic material into bacterial cells, thus providing excellent prospects for applications in microbiome therapeutics and synthetic biology.

Affinity fine-tuning anti-CAIX CAR-T cells mitigate on-target off-tumor side effects.

One of the major hurdles that has hindered the success of chimeric antigen receptor (CAR) T cell therapies against solid tumors is on-target off-tumor (OTOT) toxicity due to sharing of the same epitopes on normal tissues. To elevate the safety profile of CAR-T cells, an affinity/avidity fine-tuned CAR was designed enabling CAR-T cell activation only in the presence of a highly expressed tumor associated antigen (TAA) but not when recognizing the same antigen at a physiological level on healthy cells. Using direct stochastic optical reconstruction microscopy (dSTORM) which provides single-molecule resolution, and flow cytometry, we identified high carbonic anhydrase IX (CAIX) density on clear cell renal cell carcinoma (ccRCC) patient samples and low-density expression on healthy bile duct tissues. A Tet-On doxycycline-inducible CAIX expressing cell line was established to mimic various CAIX densities, providing coverage from CAIX-high skrc-59 tumor cells to CAIX-low MMNK-1 cholangiocytes. Assessing the killing of CAR-T cells, we demonstrated that low-affinity/high-avidity fine-tuned G9 CAR-T has a wider therapeutic window compared to high-affinity/high-avidity G250 that was used in the first anti-CAIX CAR-T clinical trial but displayed serious OTOT effects. To assess the therapeutic effect of G9 on patient samples, we generated ccRCC patient derived organotypic tumor spheroid (PDOTS) ex vivo cultures and demonstrated that G9 CAR-T cells exhibited superior efficacy, migration and cytokine release in these miniature tumors. Moreover, in an RCC orthotopic mouse model, G9 CAR-T cells showed enhanced tumor control compared to G250. In summary, G9 has successfully mitigated OTOT side effects and in doing so has made CAIX a druggable immunotherapeutic target.

Electrophoretically Co-Deposited Collagen-Lactoferrin Membranes with Enhanced Pro-Regenerative Properties for Oral Soft Tissue Regeneration.

The quality of soft tissue defect regeneration after dental surgeries largely determines their final success. Collagen membranes have been proposed for the healing of such defects, but in some cases, they do not guarantee a sufficient volume of the regenerated tissue and vascularization. For this purpose, lactoferrin, a protein with natural pro-regenerative, anti-inflammatory, and pro-angiogenic activity, can be added to collagen. In this article, we used a semipermeable barrier-assisted electrophoretic deposition (SBA-EPD) method for the production of collagen-lactoferrin membranes. The membrane structure was studied by SEM, and its mechanical properties were shown. The lactoferrin release kinetics were shown by ELISA within 75 h. When tested in vitro, we demonstrated that the collagen-lactoferrin membranes significantly increased the proliferation of keratinocytes (HaCaT) and fibroblasts (977hTERT) compared to blank collagen membranes. In vivo, on the vestibuloplasty and free gingival graft harvesting models, we showed that collagen-lactoferrin membranes decreased the wound inflammation and increased the healing rates and regeneration quality. In some parameters, collagen-lactoferrin membranes outperformed not only blank collagen membranes, but also the commercial membrane Mucograft®. Thus, we proved that collagen-lactoferrin membranes produced by the SBA-EPD method may be a valuable alternative to commercially used membranes for soft tissue regeneration in the oral cavity.

The first record of Chironomusnuditarsis Keyl, 1961 from Sevan Lake (Armenia) confirmed by morphology, karyotype and COI gene sequence.

Chironomusnuditarsis Keyl, 1961 is recorded from Sevan Lake for the first time. This species is widespread in Europe, the Caucasus, and Siberia. For species identification, we used a comprehensive approach that included morphological, cytogenetic and molecular genetic analyses. Morphological analysis showed a high similarity with the description. Nine chromosome banding sequences ndtA1, ndtA2, ndtB2, ndtC1, ndtD1, ndtE1, ndtF1, ndtG1, and ndtG2 were found. The banding sequences ndtA1, ndtA2, ndtG1, and ndtG2 are species-specific for C.nuditarsis and allow us to accurately distinguish it from the sibling species Ch.curabilis Belyanina, Sigareva et Loginova, 1990. Molecular-genetic analysis of the COI gene sequences has shown low genetic distances of 0.38-0.95% in the sibling species Ch.nuditarsis and Ch.curabilis complex and the insufficiency of using a single COI as a molecular marker for their separation.

Engineered Sustainable Omniphobic Coatings to Control Liquid Spreading on Food-Contact Materials.

The adhesion of sticky liquid foods to a contacting surface can cause many technical challenges. The food manufacturing sector is confronted with many critical issues that can be overcome with long-lasting and highly nonwettable coatings. Nanoengineered biomimetic surfaces with distinct wettability and tunable interfaces have elicited increasing interest for their potential use in addressing a broad variety of scientific and technological applications, such as antifogging, anti-icing, antifouling, antiadhesion, and anticorrosion. Although a large number of nature-inspired surfaces have emerged, food-safe nonwetted surfaces are still in their infancy, and numerous structural design aspects remain unexplored. This Review summarizes the latest scientific research regarding the key principles, fabrication methods, and applications of three important categories of nonwettable surfaces: superhydrophobic, liquid-infused slippery, and re-entrant structured surfaces. The Review is particularly focused on new insights into the antiwetting mechanisms of these nanopatterned structures and discovering efficient platform methodologies to guide their rational design when in contact with food materials. A detailed description of the current opportunities, challenges, and future scale-up possibilities of these nanoengineered surfaces in the food industry is also provided.

Green synthesis of non-isocyanate hydroxyurethane and its hybridization with carboxymethyl cellulose to produce films.

Non-isocyanate polyurethanes (NIPUs) have attracted increasing attention as a sustainable alternative to conventional isocyanate-based polyurethanes. This study synthesized non-isocyanate hydroxyurethanes (NIHUs) through an addition reaction between propylene carbonate (PC) and 1,2-ethylenediamine (EDA). The resulting NIHU was then hybridized with carboxymethyl cellulose (CMC) to investigate its hybridization potential. Structural analysis through FTIR, NMR, and XRD confirmed the crystalline nature of NIHU, featuring urethane bonds and abundant hydroxyl groups. It was found that NIHU and CMC interacted by forming hydrogen bonds between hydroxyl groups of NIHU and carboxyl groups of CMC, resulting in a dense CMC/NIHU hybrid structure. NMR and XRD analyses revealed changes in the hybrids' chain mobility, the Young's modulus of the hybrid with 30 % NIHU content decreased from 1627 MPa to 502 MPa relative to CMC, and the elongation at break increased from 4.44 % to 17.2 %. Increasing the concentration of NIHU in CMC reduced the hydrophobicity, in terms of water contact angle, from 70° to 41.7°. The simplicity of the synthesis method for NIHU, coupled with the desirable structure, strength, and balanced flexibility of CMC/NIHU hybrids, is expected to facilitate the production of NIHU-rich hybrids and increase their application in packaging.

Low-Affinity NMDA Receptor Antagonist Hemantane in a Topical Formulation Attenuates Arthritis Induced by Freund's Complete Adjuvant in Rats.

Purpose: N-methyl-D-aspartate (NMDA) receptors that are expressed by T-cells modulate T-cell proliferation, cytotoxicity and cell migration toward chemokines. Several studies have shown an anti-inflammatory effect of NMDA receptor antagonists. This study compares the effect of the noncompetitive low-affinity NMDA receptor antagonist N-(2-adamantyl)-hexamethyleneimine hydrochloride (hemantane) in a topical formulation (gel) with the cyclooxygenase (COX) inhibitor diclofenac in a topical formulation (gel) in rats with arthritis induced by Freund's Complete Adjuvant (FCA). Methods: On day 14 after an FCA injection into the left hind paw, rats with contralateral hind paw edema were selected for further investigation (29/65). They were treated with 5% hemantane gel or 1% diclofenac gel applied locally to hind paws daily for 2 weeks starting 14 days after the FCA injection. Rats with arthritis were examined hind paw edema, hyperalgesia, and motor deficits; their body weight and hematological parameters were recorded. The rats were euthanized on day 28, followed by histological examination of the ankle joint (HE stain). Results: Rats with arthritis exhibited hind paw inflammation and hyperalgesia, motor deficits, changes of hematological parameters, reduced weight gain and spleen hypertrophy. Histological examination of the ankle joint revealed degenerative-dystrophic lesions of the cartilaginous tissue, proliferative inflammation of the synovium, edema and lymphocytic/macrophage infiltration of periarticular tissues. Hemantane gel reduced hind paw edema, pain, motor deficits and histological signs of inflammation; its effect was comparable to diclofenac gel. Conclusion: Hemantane gel alleviates FCA-induced arthritis in rats, and its effect is comparable to diclofenac gel.

Impact of multiscale surface topography characteristics on Candida albicans biofilm formation: From cell repellence to fungicidal activity.

While there has been significant research conducted on bacterial colonization on implant materials, with a focus on developing surface modifications to prevent the formation of bacterial biofilms, the study of Candida albicans biofilms on implantable materials is still in its infancy, despite its growing relevance in implant-associated infections. C. albicans fungal infections represent a significant clinical concern due to their severity and associated high fatality rate. Pathogenic yeasts account for an increasing proportion of implant-associated infections, since Candida spp. readily form biofilms on medical and dental device surfaces. In addition, these biofilms are highly antifungal-resistant, making it crucial to explore alternative solutions for the prevention of Candida implant-associated infections. One promising approach is to modify the surface properties of the implant, such as the wettability and topography of these substrata, to prevent the initial Candida attachment to the surface. This review summarizes recent research on the effects of surface wettability, roughness, and architecture on Candida spp. attachment to implantable materials. The nanofabrication of material surfaces are highlighted as a potential method for the prevention of Candida spp. attachment and biofilm formation on medical implant materials. Understanding the mechanisms by which Candida spp. attach to surfaces will allow such surfaces to be designed such that the incidence and severity of Candida infections in patients can be significantly reduced. Most importantly, this approach could also substantially reduce the need to use antifungals for the prevention and treatment of these infections, thereby playing a crucial role in minimizing the possibility contributing to instances of antimicrobial resistance. STATEMENT OF SIGNIFICANCE: In this review we provide a systematic analysis of the role that surface characteristics, such as wettability, roughness, topography and architecture, play on the extent of C. albicans cells attachment that will occur on biomaterial surfaces. We show that exploiting bioinspired surfaces could significantly contribute to the prevention of antimicrobial resistance to antifungal and chemical-based preventive measures. By reducing the attachment and growth of C. albicans cells using surface structure approaches, we can decrease the need for antifungals, which are conventionally used to treat such infections.

Design and Preparation of Lifetime-Based Dual Fluorescent/Phosphorescent Sensor of pH and Oxygen and its Exploration in Model Physiological Solutions and Cells.

In the present report, a novel dual pH-O2 sensor based on covalent conjugate of rhodamine 6G and cyclometalated iridium complex with poly(vinylpyrrolidone-block-vinyltetrazole) copolymer is reported. In model physiological solutions the sensor chromophores display independent phosphorescent and fluorescent lifetime responses onto variations in oxygen concentration and pH, respectively. Colocalization studies on Chinese hamster ovary cells demonstrate the preferential localization in endosomes and lysosomes. The fluorescent lifetime imaging microscopy-phosphorescent lifetime imaging microscopy (FLIM-PLIM) experiments show that the phosphorescent O2 sensor provides unambiguous information onto hypoxia versus normoxia cell status as well as semi-quantitative data on the oxygen concentration in cells in between these two states. However, the results of FLIM measurements indicate that dynamic lifetime interval of the sensor (≈0.5 ns between pH values 5.0 and 8.0) is insufficient even for qualitative estimation of pH in living cells because half-width of lifetime distribution in the studied samples is higher than the sensor dynamic interval. Nevertheless, the variations in rhodamine emission intensity are much higher and allow rough discrimination of acidic and neutral cell conditions. Thus, the results of this study indicate that the suggested approach to the design of dual pH-O2 sensors makes possible to prepare the biocompatible and water-soluble conjugate with fast cellular uptake.

Effects of lipid extraction on stable isotope ratios of carbon and nitrogen in muscles of freshwater fish.

The extraction of lipids by the Folch method from the muscles of all the fish studied led to statistically significant differences in the values of δ15N. At the same time, lipid extraction led to a statistically significant increase in δ13C in pike and roach, and to a statistically insignificant decrease in δ13C in perch and bream. Thus, lipid extraction cannot serve as a universal method of sample preparation for the analysis of the isotopic composition of carbon (13C/12C) and nitrogen (15N/14N) in fish muscles. The differences between the δ13C values in the samples before and after lipid extraction were statistically investigated by different models. It is shown that mathematical correction method models can be used, but the results are depending on the fish types.

Fluorescent Nanocarbons: From Synthesis and Structure to Cancer Imaging and Therapy.

Nanocarbons are emerging at the forefront of nanoscience, with diverse carbon nanoforms emerging over the past two decades. Early cancer diagnosis and therapy, driven by advanced chemistry techniques, play a pivotal role in mitigating mortality rates associated with cancer. Nanocarbons, with an attractive combination of well-defined architectures, biocompatibility, and nanoscale dimension, offer an incredibly versatile platform for cancer imaging and therapy. This paper aims to review the underlying principles regarding the controllable synthesis, fluorescence origins, cellular toxicity, and surface functionalization routes of several classes of nanocarbons: carbon nanodots, nanodiamonds, carbon nanoonions, and carbon nanohorns. This review also highlights recent breakthroughs regarding the green synthesis of different nanocarbons from renewable sources. It also presents a comprehensive and unified overview of the latest cancer-related applications of nanocarbons and how they can be designed to interface with biological systems and work as cancer diagnostics and therapeutic tools. The commercial status for large-scale manufacturing of nanocarbons is also presented. Finally, it proposes future research opportunities aimed at engendering modifiable and high-performance nanocarbons for emerging applications across medical industries. This work is envisioned as a cornerstone to guide interdisciplinary teams in crafting fluorescent nanocarbons with tailored attributes that can revolutionize cancer diagnostics and therapy.

Piercing of the Human Parainfluenza Virus by Nanostructured Surfaces.

This paper presents a comprehensive experimental and theoretical investigation into the antiviral properties of nanostructured surfaces and explains the underlying virucidal mechanism. We used reactive ion etching to fabricate silicon (Si) surfaces featuring an array of sharp nanospikes with an approximate tip diameter of 2 nm and a height of 290 nm. The nanospike surfaces exhibited a 1.5 log reduction in infectivity of human parainfluenza virus type 3 (hPIV-3) after 6 h, a substantially enhanced efficiency, compared to that of smooth Si. Theoretical modeling of the virus-nanospike interactions determined the virucidal action of the nanostructured substrata to be associated with the ability of the sharp nanofeatures to effectively penetrate the viral envelope, resulting in the loss of viral infectivity. Our research highlights the significance of the potential application of nanostructured surfaces in combating the spread of viruses and bacteria. Notably, our study provides valuable insights into the design and optimization of antiviral surfaces with a particular emphasis on the crucial role played by sharp nanofeatures in maximizing their effectiveness.

Prevalence and Therapeutic Targeting of High-Level ERBB2 Amplification in NSCLC.

INTRODUCTION: ERBB2 amplification in lung cancer remains poorly characterized. HER2 (encoded by ERBB2) is a transmembrane tyrosine kinase capable of ligand-independent dimerization and signaling when overexpressed, and a common cause of HER2 overexpression is ERBB2 amplification. Here, we evaluated the clinicopathologic and genomic characteristics of ERBB2-amplified NSCLC and explored a HER2 antibody-drug conjugate (ADC) therapeutic strategy. METHODS: Our institutional next-generation DNA sequencing data (OncoPanel) from 5769 NSCLC samples (5075 patients) were queried for cases having high-level ERBB2 amplification (≥6 copies). Clinical and demographic characteristics were extracted from the electronic medical records. Efficacy of the pan-ERBB inhibitor afatinib or HER2 ADCs (trastuzumab deruxtecan and trastuzumab emtansine) was evaluated in NSCLC preclinical models and patients with ERBB2 amplification. RESULTS: High-level ERBB2 amplification was identified in 0.9% of lung adenocarcinomas and reliably predicted overexpression of HER2. ERBB2 amplification events are detected in two distinct clinicopathologic and genomic subsets of NSCLC: as the sole mitogenic driver in tumors arising in patients with a smoking history or as a concomitant alteration with other mitogenic drivers in patients with a light or never smoking history. We further reveal that trastuzumab deruxtecan is effective therapy in in vitro and in vivo preclinical models of NSCLC harboring ERBB2 amplification and report two cases of clinical activity of an anti-HER2 ADC in patients who acquired ERBB2 amplification after previous targeted therapy. CONCLUSIONS: High-level ERBB2 amplification reliably predicts HER2 overexpression in patients with NSCLC, and HER2 ADC is effective therapy in this population.

Biocatalytic Method for Producing an Affinity Resin for the Isolation of Immunoglobulins.

Affinity chromatography is a widely used technique for antibody isolation. This article presents the successful synthesis of a novel affinity resin with a mutant form of protein A (BsrtA) immobilized on it as a ligand. The key aspect of the described process is the biocatalytic immobilization of the ligand onto the matrix using the sortase A enzyme. Moreover, we used a matrix with primary amino groups without modification, which greatly simplifies the synthesis process. The resulting resin shows a high dynamic binding capacity (up to 50 mg IgG per 1 mL of sorbent). It also demonstrates high tolerance to 0.1 M NaOH treatment and maintains its effectiveness even after 100 binding, elution, and sanitization cycles.

Electrophysical Characteristics of Acrylonitrile Butadiene Styrene Composites Filled with Magnetite and Carbon Fiber Fillers.

With the rapid development of wireless communication technologies and the miniaturization trend in the electronics industry, the reduction of electromagnetic interference has become an important issue. To solve this problem, a lot of attention has been focused on polymer composites with combined functional fillers. In this paper, we report a method for creating an acrylonitrile butadiene styrene (ABS) plastic composite with a low amount of conductive carbon and magnetic fillers preparation. Also, we investigate the mechanical, thermophysical, and electrodynamic characteristics of the resulting composites. Increasing the combined filler amount in the ABS composite from 1 to 5 wt % leads to a composite conductivity growth of almost 50 times. It is necessary to underline the temperature decrease of 5 wt % mass loss and, accordingly, the composite heat resistance reduction with an increase in the combined filler from 1 to 5 wt %, while the thermal conductivity remains almost constant. It was established that electrodynamic and physical-mechanical characteristics depend on the agglomeration of fillers. This work is expected to reveal the potential of combining commercially available fillers to construct effective materials with good electromagnetic interference (EMI) protection using mass production methods (extrusion and injection molding).

Targeting TREX1 induces innate immune response in drug-resistant Small Cell Lung Cancer.

Small cell lung cancer (SCLC) is the most lethal type of lung cancer. Paradoxically, this tumor displays an initial exquisite response to chemotherapy; however, at relapse, the tumor is highly resistant to subsequent available therapies. Here, we report that the expression of three prime repair exonuclease 1 (TREX1) is strongly induced in chemoresistant SCLCs. ATAC-seq and ChIP-seq revealed a significant increase in chromatin accessibility and transcriptional activity of TREX1 gene locus in chemoresistant SCLC. Analyses of human SCLC tumors and patient-derived xenografts (PDXs) also showed increase in TREX1 expression in post-chemotherapy samples. TREX1 depletion caused the activation of cyclic GMP-AMP synthase (cGAS)-stimulator of interferon genes (STING) pathway due to cytoplasmic accumulation of damage-associated double-stranded DNA, inducing immunogenicity and enhancing the sensitivity of drug resistant cells to chemotherapy. These findings suggest TREX1 upregulation may partially contribute to the survival of resistant cells, and its inhibition may represent a promising therapeutic strategy to enhance antitumor immunity and potentiate the efficacy of chemotherapy and/or immunotherapy in chemoresistant SCLCs.

ForCenS-LGM: a dataset of planktonic foraminifera species assemblage composition for the Last Glacial Maximum.

Species assemblage composition of marine microfossils offers the possibility to investigate ecological and climatological change on time scales inaccessible using conventional observations. Planktonic foraminifera - calcareous zooplankton - have an excellent fossil record and are used extensively in palaeoecology and palaeoceanography. During the Last Glacial Maximum (LGM; 19,000 - 23,000 years ago), the climate was in a radically different state. This period is therefore a key target to investigate climate and biodiversity under different conditions than today. Studying LGM climate and ecosystems indeed has a long history, yet the most recent global synthesis of planktonic foraminifera assemblage composition is now nearly two decades old. Here we present the ForCenS-LGM dataset with 2,365 species assemblage samples collected using standardised methods and with harmonised taxonomy. The data originate from marine sediments from 664 sites and present a more than 50% increase in coverage compared to previous work. The taxonomy is compatible with the most recent global core top dataset, enabling direct investigation of temporal changes in foraminifera biogeography and facilitating seawater temperature reconstructions.

Experience in the Application of Augmented Reality Technology in the Surgical Treatment of Patients Suffering Primary and Recurrent Pelvic Tumors.

Surgical treatment of locally spread tumors in pelvic organs remains an urgent and complicated oncological problem. The recurrence rate after radical treatment ranges from 15.1% to 45.2%. The key to successful and safe surgical intervention lies in meticulous planning and intraoperative navigation, including the utilization of augmented reality (AR) technology. This paper presents the experience of clinically testing an AR technology application algorithm in the surgical treatment of 11 patients. The main stages of the algorithm are described. Radical operations incorporating intraoperative AR technology with favorable outcomes were performed on eight patients. One patient underwent a palliative intervention, while two patients did not undergo surgery. The testing of the algorithm for the application of AR technology in the surgical treatment of primary and recurrent pelvic tumors demonstrated both a technical possibility and reproducibility of this algorithm and the AR technology itself in clinical practice.