It is not just about prescription. A cohort study of the impact of enteral nutrition on mortality of hospitalized patients with CoVID-19 / No es solo una prescripción. Estudio de cohorte del impacto de la nutrición enteral en la mortalidad de pacientes hospitalizados con COVID-19

Introduction: during COVID-19 pandemic, international societies released guidelines and recommendations for patients requiring nutritionalsupport according to previous similar respiratory diseases.Objectives: the aim of the study was to evaluate the nutritional support provided by enteral nutrition (EN) in patients with COVID-19 infection,identify if the recommendations from international societies were met and their impact on mortality rate.Methods: a cohort study was conducted on adult patients with COVID-19 admitted to a tertiary hospital. Demographic, clinical, biochemical, andnutritional variables were obtained. A random-effect parametric survival-time model was performed to quantify the risk of death for each variable,and the Hausman test was used to confirm the model.Results: two hundred and twenty-nine patients were enrolled. The delivered energy was > 80 % of adequacy in the first two days, as suggestedby international guidelines (11.7 ± 4.9 kcal/kg); however, an adequacy rate less than 60 % was achieved on day 14 (25.4 ± 7.4 kcal/kg). Theprotein adequacy was > 75 % on the first days of infusion (1.3 ± 0.3 g/kg); however, the infusion was < 50 % (1.5 ± 0.4 g/kg) after being extu-bated. Age, sex, and nutritional risk were related to higher mortality in patients with EN, whereas the infused energy and protein, the percentageof protein adequacy, arginine, and n-3 PUFA were associated with lower mortality.Conclusion: achieving at least 80 % of the energy and protein requirements, as well as n-3 PUFA and arginine supplementation could be asso-ciated with lower mortality in COVID-19 patients. More studies are needed to confirm the role of these nutrients on the mortality rate.(AU)

Introducción: durante la pandemia de COVID-19, las sociedades internacionales publicaron guías y recomendaciones para pacientes querequieren apoyo nutricional basándose en lo previamente recomendado en enfermedades respiratorias similares.Objetivos: evaluar el soporte nutricional con nutrición enteral (NE) en pacientes con COVID-19 e identificar el cumplimiento de las recomenda-ciones hechas por las sociedades internacionales y su impacto en la tasa de mortalidad.Métodos: estudio de cohorte en adultos con COVID-19 ingresados en un hospital de tercer nivel. Se registraron variables demográficas, clínicas,bioquímicas y nutricionales. Se realizó un modelo de supervivencia de efectos aleatorios para cuantificar el riesgo de muerte para cada variabley la prueba de Hausman para confirmar el modelo.Resultados: se incluyeron 229 pacientes. La energía administrada fue > 80 % de adecuación en los dos primeros días (11,7 ± 4,9 kcal/kg);sin embargo, fue < 60 % el día 14 (25,4 ± 7,4 kcal/kg). La adecuación de proteínas fue > 75 % en los primeros días de infusión (1,3 ± 0,3g/kg), pero < 50 % (1,5 ± 0,4 g/kg) después de ser extubado. La edad, el sexo y el riesgo nutricional se relacionaron con mayor mortalidad,mientras que la energía y proteína infundidas, el porcentaje de adecuación proteica, la arginina y el contenido de ácidos grasos poliinsaturados(AGPI) n-3 se asociaron con menor mortalidad.Conclusión: aunque se necesitan más estudios para confirmarlo, alcanzar al menos el 80 % de los requerimientos energéticos y proteicos, asícomo la suplementación de fórmulas con AGPI n-3 y arginina, podría asociarse con menor mortalidad en pacientes con COVID-19.(AU)

Rosacea treatment with mussel adhesive protein delivered via microneedling: In vivo and clinical studies.

BACKGROUND: Rosacea is a prevalent chronic dermatological condition marked by facial inflammation and erythema, significantly compromising the quality of life for affected individuals. Current treatment methods for rosacea are not considered ideal because of the complex etiology of the disease. Mussel adhesive protein (MAP) is a glycoprotein derived from the foot gland of mussels. The protein exhibits anti-inflammatory properties, relieves skin itching, and promotes wound healing. AIMS: We aimed to explore the feasibility of using MAP administered via microneedle delivery for treating rosacea and the potential molecular mechanism involved. MATERIALS AND METHODS: The therapeutic effect and mechanism of MAP microneedle delivery in an LL-37-induced rosacea-like mouse model were observed using morphological and histological methods. Twenty-seven patients with erythematotelangiectatic rosacea (ETR) underwent treatment once every 1 month, with three treatments constituting one treatment course. The therapeutic effect was evaluated by comparing the clinical images taken at baseline, after the first treatment course, and after the second treatment course. The red value, CEA, and GFSS score were also calculated. RESULTS: In response to the microneedle delivery of MAP, innate immunity, inflammatory infiltration, and abnormal neurovascular regulation improved significantly in rosacea-like mice. In the clinical experiments, the microneedle delivery of MAP significantly improved the symptoms of erythema, flushing, and telangiectasia in patients with ETR, and no obvious adverse reactions were observed. CONCLUSIONS: MAP delivered by microneedling is effective and safe for treating ETR.

The strawberry-derived permeation enhancer pelargonidin enables oral protein delivery.

Although patients generally prefer oral drug delivery to injections, low permeability of the gastrointestinal tract makes this method impossible for most biomacromolecules. One potential solution is codelivery of macromolecules, including therapeutic proteins or nucleic acids, with intestinal permeation enhancers; however, enhancer use has been limited clinically by modest efficacy and toxicity concerns surrounding long-term administration. Here, we hypothesized that plant-based foods, which are well tolerated by the gastrointestinal tract, may contain compounds that enable oral macromolecular absorption without causing adverse effects. Upon testing more than 100 fruits, vegetables, and herbs, we identified strawberry and its red pigment, pelargonidin, as potent, well-tolerated enhancers of intestinal permeability. In mice, an oral capsule formulation comprising pelargonidin and a 1 U/kg dose of insulin reduced blood glucose levels for over 4 h, with bioactivity exceeding 100% relative to subcutaneous injection. Effects were reversible within 2 h and associated with actin and tight junction rearrangement. Furthermore, daily dosing of mice with pelargonidin for 1 mo resulted in no detectable side effects, including weight loss, tissue damage, or inflammatory responses. These data suggest that pelargonidin is an exceptionally effective enhancer of oral protein uptake that may be safe for routine pharmaceutical use.

Effectiveness Comparison of Polysaccharides, Proteins, and Lipids as Composite Edible Coatings on the Quality of Food Products / Comparación de la Efectividad de Polisacáridos, Proteínas y Lípidos como Recubrimientos Compuestos Comestibles en la Calidad de los Productos Alimenticios

Background: This research was motivated by the complaints of tomato farmers about their crops that quickly rotted before being sold, as well as the many research results (raw materials and methods) that edible coating films could not be applied optimally. Objectives: The research was a practical recommendation by comparing the effectiveness of raw materials (polysaccharides, proteins, and lipids) with the dipping and spray methods. Materials and methods used in the comparison process were the application of Structural Equation Modeling (SEM) with the Partial Least Square (PLS) approach. Results: Dipping has a strong effect (f2 ≥ 0.35; p<0.05), while spray had a moderate effect (f2: 0.15-0.35; p<0.05). Thus, the role of dipping as a mediator was more dominant than spray. Compared to proteins and lipids, polysaccharides had the best effectiveness (ß:0.460-0.584; f2: 0.15-0.35; p<0.05). Conclusion: the three ingredients improved the quality of tomatoes, and the dipping method was easier to apply by farmers than the spray method, which had many obstacles in its application

Antecedentes: esta investigación está motivada por las quejas de los productores de tomate sobre sus cultivos que se pudren rápidamente antes de ser vendidos, así como por los muchos resultados de la investigación (materias primas y métodos) de que las películas de recubrimiento comestibles no se pudieron aplicar de manera óptima. Objetivos: La investigación consiste en recomendaciones prácticas mediante la comparación de la eficacia de las materias primas (polisacáridos, proteínas y lípidos) con los métodos de inmersión y aspersión. Métodos: El método utilizado en el proceso de comparación es la aplicación del modelo de ecuaciones estructurales (SEM) con el enfoque de mínimos cuadrados parciales (PLS). Resultados: La inmersión tiene un efecto fuerte (f2 ≥ 0,35; p<0,05), mientras que la pulverización tiene un efecto moderado (f2: 0,15-0,35; p<0,05). Por lo tanto, el papel de la inmersión como mediador es más dominante que el del rociado. Los polisacáridos tienen la mejor eficacia (ß:0,460-0,584; f2: 0,15-0,35; p<0,05) en comparación con las proteínas y los lípidos. Conclusión: es que los tres ingredientes pueden mejorar la calidad de los tomates, y el método de inmersión es más fácil de aplicar por los agricultores que el método de aspersión, que tiene muchos obstáculos en su aplicación

Intralesional Infiltrations of Arteriosclerotic Tissue Cells-Free Filtrate Reproduce Vascular Pathology in Healthy Recipient Rats.

Lower-extremity arterial disease is a major health problem with increasing prevalence, often leading to non-traumatic amputation, disability and mortality. The molecular mechanisms underpinning abnormal vascular wall remodeling are not fully understood. We hypothesized on the existence of a vascular tissue memory that may be transmitted through soluble signaling messengers, transferred from humans to healthy recipient animals, and consequently drive the recapitulation of arterial wall thickening and other vascular pathologies. We examined the effects of the intralesional infiltration for 6 days of arteriosclerotic popliteal artery-derived homogenates (100 µg of protein) into rats' full-thickness wounds granulation tissue. Animals infiltrated with normal saline solution or healthy brachial arterial tissue homogenate obtained from traumatic amputation served as controls. The significant thickening of arteriolar walls was the constant outcome in two independent experiments for animals receiving arteriosclerotic tissue homogenates. This material induced other vascular morphological changes including an endothelial cell phenotypic reprogramming that mirrored the donor's vascular histopathology. The immunohistochemical expression pattern of relevant vascular markers appeared to match between the human tissue and the corresponding recipient rats. These changes occurred within days of administration, and with no cross-species limitation. The identification of these "vascular disease drivers" may pave novel research avenues for atherosclerosis pathobiology.

Oral delivery of therapeutic peptides and proteins: Technology landscape of lipid-based nanocarriers.

The oral administration of therapeutic peptides and proteins is favoured from a patient and commercial point of view. In order to reach the systemic circulation after oral administration, these drugs have to overcome numerous barriers including the enzymatic, sulfhydryl, mucus and epithelial barrier. The development of oral formulations for therapeutic peptides and proteins is therefore necessary. Among the most promising formulation approaches are lipid-based nanocarriers such as oil-in-water nanoemulsions, self-emulsifying drug delivery systems (SEDDS), solid lipid nanoparticles (SLN), nanostructured lipid carriers (NLC), liposomes and micelles. As the lipophilic character of therapeutic peptides and proteins can be tremendously increased such as by the formation of hydrophobic ion pairs (HIP) with hydrophobic counter ions, they can be incorporated in the lipophilic phase of these carriers. Since gastrointestinal (GI) peptidases as well as sulfhydryl compounds such as glutathione and dietary proteins are too hydrophilic to enter the lipophilic phase of these carriers, the incorporated therapeutic peptide or protein is protected towards enzymatic degradation as well as unintended thiol/disulfide exchange reactions. Stability of lipid-based nanocarriers towards lipases can be provided by the use to excipients that are not or just poorly degraded by these enzymes. Nanocarriers with a size <200 nm and a mucoinert surface such as PEG or zwitterionic surfaces exhibit high mucus permeating properties. Having reached the underlying absorption membrane, lipid-based nanocarriers enable paracellular and lymphatic drug uptake, induce endocytosis and transcytosis or simply fuse with the cell membrane releasing their payload into the systemic circulation. Numerous in vivo studies provide evidence for the potential of these delivery systems. Within this review we provide an overview about the different barriers for oral peptide and protein delivery, highlight the progress made on lipid-based nanocarriers in order to overcome them and discuss strengths and weaknesses of these delivery systems in comparison to other technologies.

Protein and peptide delivery to lungs by using advanced targeted drug delivery.

The challenges and difficulties associated with conventional drug delivery systems have led to the emergence of novel, advanced targeted drug delivery systems. Therapeutic drug delivery of proteins and peptides to the lungs is complicated owing to the large size and polar characteristics of the latter. Nevertheless, the pulmonary route has attracted great interest today among formulation scientists, as it has evolved into one of the important targeted drug delivery platforms for the delivery of peptides, and related compounds effectively to the lungs, primarily for the management and treatment of chronic lung diseases. In this review, we have discussed and summarized the current scenario and recent developments in targeted delivery of proteins and peptide-based drugs to the lungs. Moreover, we have also highlighted the advantages of pulmonary drug delivery over conventional drug delivery approaches for peptide-based drugs, in terms of efficacy, retention time and other important pharmacokinetic parameters. The review also highlights the future perspectives and the impact of targeted drug delivery on peptide-based drugs in the coming decade.

Advances in the development of antimicrobial peptides and proteins for inhaled therapy.

Antimicrobial peptides and proteins (APPs) are becoming increasingly important in targeting multidrug-resistant (MDR) bacteria. APPs is a rapidly emerging area with novel molecules being produced and further optimised to enhance antimicrobial efficacy, while overcoming issues associated with biologics such as potential toxicity and low bioavailability resulting from short half-life. Inhalation delivery of these agents can be an effective treatment of respiratory infections owing to the high local drug concentration in the lungs with lower exposure to systemic circulation hence reducing systemic toxicity. This review describes the recent studies on inhaled APPs, including in vitro and in vivo antimicrobial activities, toxicity assessments, and formulation strategies whenever available. The review also includes studies on combination of APPs with other antimicrobial agents to achieve enhanced synergistic antimicrobial effect. Since different APPs have different biological and chemical stabilities, a targeted formulation strategy should be considered for developing stable and inhalable antimicrobial peptides and proteins. These strategies include the use of sodium chloride to reduce electrostatic interaction between APP and extracellular DNA in sputum, the use of D-enantiomers or dendrimers to minimise protease-mediated degradation and or the use of prodrugs to reduce toxicity. Although great effort has been put towards optimising the biological functions of APPs, studies assessing biological stability in inhalable aerosols are scarce, particularly for novel molecules. As such, formulation and manufacture of inhalable liquid and powder formulations of APPs are underexplored, yet they are crucial areas of research for clinical translation.

Ion-dependent slow protein release from in vivo disintegrating micro-granules.

Through the controlled addition of divalent cations, polyhistidine-tagged proteins can be clustered in form of chemically pure and mechanically stable micron-scale particles. Under physiological conditions, these materials act as self-disintegrating protein depots for the progressive release of the forming polypeptide, with potential applications in protein drug delivery, diagnosis, or theragnosis. Here we have explored the in vivo disintegration pattern of a set of such depots, upon subcutaneous administration in mice. These microparticles were fabricated with cationic forms of either Zn, Ca, Mg, or Mn, which abound in the mammalian body. By using a CXCR4-targeted fluorescent protein as a reporter building block we categorized those cations regarding their ability to persist in the administration site and to sustain a slow release of functional protein. Ca2+ and specially Zn2+ have been observed as particularly good promoters of time-prolonged protein leakage. The released polypeptides result is available for selective molecular interactions, such as specific fluorescent labeling of tumor tissues, in which the protein reaches nearly steady levels.

Short-Term Pre-Operative Protein Caloric Restriction in Elective Vascular Surgery Patients: A Randomized Clinical Trial.

(1) Background: Vascular surgery operations are hampered by high failure rates and frequent occurrence of peri-operative cardiovascular complications. In pre-clinical studies, pre-operative restriction of proteins and/or calories (PCR) has been shown to limit ischemia-reperfusion damage, slow intimal hyperplasia, and improve metabolic fitness. However, whether these dietary regimens are feasible and safe in the vascular surgery patient population remains unknown. (2) Methods: We performed a randomized controlled trial in patients scheduled for any elective open vascular procedure. Participants were randomized in a 3:2 ratio to either four days of outpatient pre-operative PCR (30% calorie, 70% protein restriction) or their regular ad-libitum diet. Blood was drawn at baseline, pre-operative, and post-operative day 1 timepoints. A leukocyte subset flow cytometry panel was performed at these timepoints. Subcutaneous/perivascular adipose tissue was sampled and analyzed. Follow-up was one year post-op. (3) Results: 19 patients were enrolled, of whom 11 completed the study. No diet-related reasons for non-completion were reported, and there was no intervention group crossover. The PCR diet induced weight loss and BMI decrease without malnutrition. Insulin sensitivity was improved after four days of PCR (p = 0.05). Between diet groups, there were similar rates of re-intervention, wound infection, and cardiovascular complications. Leukocyte populations were maintained after four days of PCR. (4) Conclusions: Pre-operative PCR is safe and feasible in elective vascular surgery patients.

Delivering more for less: nanosized, minimal-carrier and pharmacoactive drug delivery systems.

Traditional nanoparticle carriers such as liposomes, micelles, and polymeric vehicles improve drug delivery by protecting, stabilizing, and increasing the circulatory half-life of the encapsulated drugs. However, traditional drug delivery systems frequently suffer from poor drug loading and require an excess of carrier materials. This carrier material excess poses an additional systemic burden through accumulation, if not degradable the need for metabolism, and potential toxicity. To address these shortcomings, minimal-carrier nanoparticle systems and pharmacoactive carrier materials have been developed. Both solutions provide drug delivery systems in which the majority of the nanoparticle is pharmacologically active. While minimal-carrier and pharmacoactive drug delivery systems can improve drug loading, they can also suffer from poor stability. Here, we review minimal-carrier and pharmacoactive delivery systems, discuss ongoing challenges and outline opportunities to translate minimal-carrier and pharmacoactive drug delivery systems into the clinic.

Cytosolic protein delivery using pH-responsive, charge-reversible lipid nanoparticles.

Although proteins have attractive features as biopharmaceuticals, the difficulty in delivering them into the cell interior limits their applicability. Lipid nanoparticles (LNPs) are a promising class of delivery vehicles. When designing a protein delivery system based on LNPs, the major challenges include: (i) formulation of LNPs with defined particle sizes and dispersity, (ii) efficient encapsulation of cargo proteins into LNPs, and (iii) effective cellular uptake and endosomal release into the cytosol. Dioleoylglycerophosphate-diethylenediamine (DOP-DEDA) is a pH-responsive, charge-reversible lipid. The aim of this study was to evaluate the applicability of DOP-DEDA-based LNPs for intracellular protein delivery. Considering the importance of electrostatic interactions in protein encapsulation into LNPs, a negatively charged green fluorescent protein (GFP) analog was successfully encapsulated into DOP-DEDA-based LNPs to yield diameters and polydispersity index of < 200 nm and < 0.2, respectively. Moreover, ~ 80% of the cargo proteins was encapsulated into the LNPs. Cytosolic distribution of fluorescent signals of the protein was observed for up to ~ 90% cells treated with the LNPs, indicating the facilitated endocytic uptake and endosomal escape of the cargo attained using the LNP system.

Uso de la fórmula extensamente hidrolizada de proteína con 30% o más de triglicéridos de cadena media en el manejo nutricional de niños menores de cinco años, con cirrosis hepática colestásica, candidatos a trasplante hepático y postrasplantados hasta los seis meses / Use of extensively hydrolyzed protein formula with 30% or more of medium chain triglycerides in the nutritional management of children under five years of age, with cholestatic liver cirrhosis, liver transplant candidates and post-transplant patients up to six months

INTRODUCCIÓN: El presente dictamen expone la evaluación de la eficacia y seguridad de la fórmula extensamente hidrolizada de proteína con 30 % o más de triglicéridos de cadena media (TCM), en comparación con la mejor terapia de soporte nutricional, para preservar y/o mejorar el estado nutricional de niños menores de cinco años, con cirrosis hepática, con colestasis y candidatos a trasplante y postrasplantados hasta los seis meses. La cirrosis hepática es una condición producida por daño hepático crónico que conduce a falla hepática y reemplazo del parénquima hepático por tejido cicatricial. En recién nacidos, se describe una incidencia de aproximadamente 0.8 casos por cada 10,000 y en el Perú no se dispone de estudios epidemiológicos sobre esta patología. El trasplante hepático incrementa la sobrevida y calidad de vida de los pacientes pediátricos con enfermedad hepática crónica. La colestasis, una condición que suele presentarse en estos pacientes, limita la absorción de grasas a nivel intestinal; por tal

PLGA/PLA-Based Long-Acting Injectable Depot Microspheres in Clinical Use: Production and Characterization Overview for Protein/Peptide Delivery.

Over the past few decades, long acting injectable (LAI) depots of polylactide-co-glycolide (PLGA) or polylactic acid (PLA) based microspheres have been developed for controlled drug delivery to reduce dosing frequency and to improve the therapeutic effects. Biopharmaceuticals such as proteins and peptides are encapsulated in the microspheres to increase their bioavailability and provide a long release period (days or months) with constant drug plasma concentration. The biodegradable and biocompatible properties of PLGA/PLA polymers, including but not limited to molecular weight, end group, lactide to glycolide ratio, and minor manufacturing changes, could greatly affect the quality attributes of microsphere formulations such as release profile, size, encapsulation efficiency, and bioactivity of biopharmaceuticals. Besides, the encapsulated proteins/peptides are susceptible to harsh processing conditions associated with microsphere fabrication methods, including exposure to organic solvent, shear stress, and temperature fluctuations. The protein/peptide containing LAI microspheres in clinical use is typically prepared by double emulsion, coacervation, and spray drying techniques. The purpose of this review is to provide an overview of the formulation attributes and conventional manufacturing techniques of LAI microspheres that are currently in clinical use for protein/peptides. Furthermore, the physicochemical characteristics of the microsphere formulations are deliberated.

Antioxidant Activity Derived from Marine Green-Lipped Mussel Perna canaliculus Extracts in Mice.

This study investigates the antioxidant activities of lipid, protein, and carbohydrate extracts from the marine mollusk Perna canaliculus. Lipids were extracted using acetone, which was followed by protein extraction using the broad-spectrum enzyme Alcalase and then carbohydrate extraction using cetylpyridinium chloride. Eighty white BALB/c mice were divided into eight groups according to the administered extracts. Groups 1 and 5 were the control and toxin control groups, respectively. Groups 2, 3, and 4 were administered lipid, protein, and carbohydrate extracts, respectively. The other groups were administered P. canaliculus extracts as well as gentamicin and acetaminophen, known as ethanolic extracts, derived from Nerium oleander to induce oxidation stress. All groups showed significant improvements in body weight (p < 0.05). The lipid extract group showed a significant decrease in low-density lipoprotein cholesterol (p < 0.05) and a significant increase in high-density lipoprotein cholesterol (p < 0.05). After the toxin injection, all groups treated with P. canaliculus extracts showed increased antioxidant effects on hepatocytes (p < 0.05). The lipid extracts induced antioxidant effects to protect the kidney by increasing lipid peroxidation (p < 0.05) and catalase activities (p < 0.05). Also, protein extracts showed antioxidant effects by increasing glutathione and catalase levels significantly (p < 0.005). In conclusion, P. canaliculus extracts, especially lipids and proteins, have potent antioxidant activities that protect vital organs from oxidation stress.

Combined Application of Prototype Ultrasound and BSA-Loaded PLGA Particles for Protein Delivery.

PURPOSE: To develop an in vitro culture system for tissue engineering to mimic the in vivo environment and evaluate the applicability of ultrasound and PLGA particle system. METHODS: For tissue engineering, large molecules such as growth factors for cell differentiation should be supplied in a controlled manner into the culture system, and the in vivo microenvironment need to be reproduced in the system for the regulation of cellular function. In this study, portable prototype ultrasound with low intensity was devised and tested for protein release from bovine serum albumin (BSA)-loaded poly(lactic-co-glycolic acid) (PLGA) particles. RESULTS: BSA-loaded PLGA particles were prepared using various types of PLGA reagents and their physicochemical properties were characterized including particle size, shape, or aqueous wetting profiles. The BSA-loaded formulation showed nano-ranged size distribution with optimal physical stability during storage period, and protein release behaviors in a controlled manner. Notably, the application of prototype ultrasound with low intensity influenced protein release patterns in the culture system containing the BSA-loaded PLGA formulation. The results revealed that the portable ultrasound set controlled by the computer could contribute for the protein delivery in the culture medium. CONCLUSIONS: This study suggests that combined application with ultrasound and protein-loaded PLGA encapsulation system could be utilized to improve culture system for tissue engineering or cell regeneration therapy.

Rapid protein delivery to living cells for biomolecular investigation.

The lack of a simple, fast and efficient method for protein delivery is limiting the widespread application of in-cell experiments, which are crucial for understanding the cellular function. We present here an innovative strategy to deliver proteins into both prokaryotic and eukaryotic cells, exploiting thermal vesiculation. This method allows to internalize substantial amounts of proteins, with different molecular weight and conformation, without compromising the structural properties and cell viability. Characterizing proteins in a physiological environment is essential as the environment can dramatically affect the conformation and dynamics of biomolecules as shown by in-cell EPR spectra vs those acquired in buffer solution. Considering its versatility, this method opens the possibility to scientists to study proteins directly in living cells through a wide range of techniques.

Nanotechnologies for the delivery of biologicals: Historical perspective and current landscape.

Biological macromolecule-based therapeutics irrupted in the pharmaceutical scene generating a great hope due to their outstanding specificity and potency. However, given their susceptibility to degradation and limited capacity to overcome biological barriers new delivery technologies had to be developed for them to reach their targets. This review aims at analyzing the historical seminal advances that shaped the development of the protein/peptide delivery field, along with the emerging technologies on the lead of the current landscape. Particularly, focus is made on technologies with a potential for transmucosal systemic delivery of protein/peptide drugs, followed by approaches for the delivery of antigens as new vaccination strategies, and formulations of biological drugs in oncology, with special emphasis on mAbs. Finally, a discussion of the key challenges the field is facing, along with an overview of prospective advances are provided.

Intracellular Delivery of Proteins into Living Cells by Low-Molecular-Weight Polyethyleneimine.

INTRODUCTION: Intracellular protein delivery is emerging as a potential strategy to revolutionize therapeutics in the field of biomedicine, aiming at treating a wide range of diseases including cancer, inflammatory diseases and other oxidative stress-related disorders with high specificity. However, the current challenges and limitations are addressed to either synthetically or biologically through multipotency of engineering, such as protein modification, insufficient delivery of large-size proteins, deficiency or mutation of proteins, and high cytotoxicity. METHODS: We prepared the nanocomposites by mixing protein with PEI1200 at a certain molar ratio and demonstrated that it can deliver proteins into living cells in high efficiency and safety through the following experiments, such as dynamic light scattering, fluorescent detection, agarose gel electrophoresis, ß-Galactosidase activity detection, immunofluorescence staining, digital fluorescent detection, cell viability assay and flow cytometry. RESULTS: The self-assembly of PEI1200/protein nanocomposites with appropriate molar ratio (4:1 and 8:1) could provide efficiently delivery of active proteins to a variety of cell types in the presence of serum. The nanocomposites could continuously release protein up to 96 h in their desired intracellular locations. In addition, these nanocomposites were able to preserve protein activity while maintain low cytotoxicity (when final concentration <1 µg/mL). CONCLUSION: Collectively, PEI1200-based delivery system provided an alternative strategy to direct protein delivery in high efficiency and safety, offering increased potential applications in clinical biomedicine.