Rapid and Noninvasive Quantification of Capsid Gene Filling Level Using Water Proton Nuclear Magnetic Resonance.

The present work reports an enabling novel technology for quantifying the gene content in adeno-associated viral capsids. The method is based on the water proton nuclear magnetic resonance (wNMR) technique. Instead of analyzing the capsid directly, it utilizes water molecules to distinguish empty and full capsids, as water interacts with them differently. The transverse relaxation rate of water protons, R2(1H2O), readily distinguishes empty and full capsids and is capable of quantifying the fraction of full capsids in a mixture of full and empty ones. It involves no sample preparation and no reagents. Measurement is rapid (data collection takes 1-2 min), noninvasive (the capsid sample can stay inside the original sealed and labeled container to be used in other studies or administered to a patient), and performed using a wide-bore benchtop NMR instrument. The method can be readily implemented at a production plant for product release as part of product quality control.

Evaluation of the Physicochemical Properties of the Iron Nanoparticle Drug Products: Brand and Generic Sodium Ferric Gluconate.

Complex iron nanoparticle-based drugs are one of the oldest and most frequently administered classes of nanomedicines. In the US, there are seven FDA-approved iron nanoparticle reference drug products, of which one also has an approved generic drug product (i.e., sodium ferric gluconate (SFG)). These products are indicated for the treatment of iron deficiency anemia and are administered intravenously. On the molecular level, iron nanomedicines are colloids composed of an iron oxide core with a carbohydrate coating. This formulation makes nanomedicines more complex than conventional small molecule drugs. As such, these products are often referred to as nonbiological complex drugs (e.g., by the nonbiological complex drugs (NBCD) working group) or complex drug products (e.g., by the FDA). Herein, we report a comprehensive study of the physiochemical properties of the iron nanoparticle product SFG. SFG is the single drug for which both an innovator (Ferrlecit) and generic product are available in the US, allowing for comparative studies to be performed. Measurements focused on the iron core of SFG included optical spectroscopy, inductively coupled plasma mass spectrometry (ICP-MS), X-ray powder diffraction (XRPD), 57Fe Mössbauer spectroscopy, and X-ray absorbance spectroscopy (XAS). The analysis revealed similar ferric-iron-oxide structures. Measurements focused on the carbohydrate shell comprised of the gluconate ligands included forced acid degradation, dynamic light scattering (DLS), analytical ultracentrifugation (AUC), and gel permeation chromatography (GPC). Such analysis revealed differences in composition for the innovator versus the generic SFG. These studies have the potential to contribute to future quality assessment of iron complex products and will inform on a pharmacokinetic study of two therapeutically equivalent iron gluconate products.

A Comparison of Clinical Characteristics and Outcomes in Elderly and Younger Patients with COVID-19.

BACKGROUND The aim of this study was to describe the clinical characteristics and outcomes of patients with coronavirus disease 2019 (COVID-19) and compare these parameters in an elderly group with those in a younger group. MATERIAL AND METHODS This retrospective, single-center observational study included 69 hospitalized patients with laboratory-confirmed COVID-19 from a tertiary hospital in Wuhan, China, between January 14, 2020, and February 26, 2020. Epidemiological, demographic, clinical, and laboratory data, as well as treatments, complications, and outcomes were extracted from electronic medical records and compared between elderly patients (aged ≥60 years) and younger patients (aged <60 years). Patients were followed until March 19, 2020. RESULTS Elderly patients had more complications than younger patients, including acute respiratory distress syndrome (ARDS; 9/25, 36% vs. 5/44, 11.4%) and cardiac injury (7/25, 28% vs. 1/44, 2.3%), and they were more likely to be admitted to the intensive care unit (6/25, 24% vs. 2/44, 4.5%). As of March 19, 2020, 60/69 (87%) of the patients had been discharged, 6/69 (8.7%) had died, and 3/69 (4.3%) remained in the hospital. Of those who were discharged or died, the median duration of hospitalization was 13.5 days (interquartile range, 10-18 days). CONCLUSIONS Elderly patients with confirmed COVID-19 were more likely to develop ARDS and cardiac injury than younger patients and were more likely to be admitted to the intensive care unit. In addition to routine monitoring and respiratory support, cardiac monitoring and supportive care should be a focus in elderly patients with COVID-19.

Hepatic portal venous gas with pneumatosis intestinalis secondary to mesenteric ischemia in elderly patients: Two case reports.

INTRODUCTION: Hepatic portal venous gas (HPVG) is a rare imaging finding. When HPVG is accompanied with pneumatosis intestinalis (PI), the underlying cause is usually mesenteric ischemia with consequent intestinal necrosis. This combination of clinical conditions is associated with a poor prognosis. In this study, we present the cases of 2 elderly patients with HPVG and PI secondary to mesenteric ischemia. PATIENT CONCERNS: In case 1, a 89-year-old male patient was admitted to intensive care unit with respiratory failure, On the fifth day of admission, he developed a high fever (39.5°C) and abdominal distension. In case 2, a 92-year-old male patient admitted to our intensive care unit and received mechanical ventilation due to acute respiratory failure. During the treatment, the patient developed gastrointestinal bleeding. On physical examination, abdominal bulging and tense abdominal walls were detected. Both patients underwent abdominal contrast-enhanced computed tomography, showed abundant HPVG with PI. DIAGNOSES: The patients were diagnosed as acute mesenteric ischemia, bowel necrosis, septic shock, multiple organ dysfunction syndrome based on computed tomography scan, abdominal signs, and laboratory tests. INTERVENTIONS: Fluid resuscitation, high-dose vasopressors, and intravenous antibiotic therapy were given. OUTCOMES: Despite prompt treatment, the condition of both patients rapidly deteriorated, and the patients died shortly thereafter. CONCLUSION: Mesenteric ischemia is a clinical emergency. In patients with risk factors and abdominal signs, the clinical suspicion for this condition should be high. Although rare, both HPVG and PI are important radiological clues that usually indicate the presence of mesenteric ischemia with consequent intestinal necrosis.

Age and cigarette smoking modulate the relationship between pulmonary function and arterial stiffness in heart failure patients.

The aim of this study was to assess the relationship between arterial stiffness and pulmonary function in chronic heart failure (CHF).Outpatients previously diagnosed as CHF were enrolled between April 2008 and March 2010, and submitted to arterial stiffness measurement and lung function assessment. Spirometry was performed by measuring forced vital capacity (FVC), the fraction of predicted FVC, forced expiratory volume in 1 second (FEV1), the percentage of predicted FEV1 in 1 second, FEV1 to FVC ratio, and the percentage of predicted FEV1/FVC. Cardio-ankle vascular index (CAVI) was considered for the estimation of arterial stiffness.The 354 patients assessed included 315 nonsmokers, and were 68.2 ±â€Š7.2 years' old. Unadjusted correlation analyses demonstrated CAVI was positively related to age (r = 0.3664, P < 0.0001), and negatively related to body mass index (BMI, r = -0.2040, P = 0.0001), E/A ratio (r = -0.1759, P = 0.0010), and FEV1 (r = -0.2987, P < 0.0001). Stepwise multivariate regression analyses showed age (r = 0.2391, P < 0.0001), BMI (r = -0.2139, P < 0.0001), smoking (r = 0.1211, P = 0.0130), E/A ratio (r = -0.1082, P = 0.0386), and FEV1 (r = -0.2550, P < 0.0001) were independent determinants of CAVI. In addition, there is a significant interaction between CAVI and forced expiratory volume in 1 second (FEV1) in relation to age (Pint < 0.0001) and smoking (Pint = 0.0001). Meanwhile, pulmonary function was not associated with BMI or E/A ratio.These findings demonstrated that reduced pulmonary function is associated with the increased CAVI, and had an interactive effect with age and smoking on CAVI in patients with CHF.

ß3-integrin inhibits lipopolysaccharide-induced autophagy in cardiomyocytes via the Akt signaling pathway.

OBJECTIVE: To investigate the role of ß 3 -integrin in lipopolysaccharide (LPS)-induced autophagy in cardiomyocytes and its underlying mechanism. METHODS: ß 3 -Integrin expression in cardiomyocytes was up- or downregulated by adenovirus transfection or cyclic arginine-glycine-aspartic acid (cRGD) peptide treatment before LPS stimulation. The expression of autophagy-associated proteins (LC3-II, Beclin-1 and Bcl-2) and the activation of Akt were determined using Western blotting. Autophagosomes and autophagic vacuoles were observed using monodansylcadaverine (MDC) dye and transmission electron microscopy, respectively. RESULTS: Downregulation of ß 3 -integrin with cRGD peptide resulted in enhanced LC3-II and Beclin-1 and decreased Bcl-2 expression. Low Beclin-1 levels were detected after LPS stimulation in adenovirus ß 3 -integrin-transfected cardiomyocytes. There was no significant difference in LC3-II levels between control and adenovirus ß 3 -integrin-transfected cardiomyocytes. Enhanced accumulation of MDC dye and autophagosomes, which were inhibited by ß 3 -integrin overexpression, were detected after LPS treatment. The increased phosphorylation of Akt after LPS stimulation was inhibited by cRGD and enhanced by ß 3 -integrin overexpression. Furthermore, the Akt inhibitor triciribine inhibited the negative effect of ß 3 - integrin on autophagy, as shown by LC3-II and Beclin-1 upregulation. CONCLUSIONS: ß 3 -Integrin inhibits LPS-induced autophagy in cardiomyocytes. The inhibition of Akt signaling might be an important mechanism in this process.

Metabonomic studies of pancreatic cancer response to radiotherapy in a mouse xenograft model using magnetic resonance spectroscopy and principal components analysis.

AIM: To investigate the metabolic profiles of xenograft pancreatic cancer before and after radiotherapy by high-resolution magic angle spinning proton magnetic resonance spectroscopy (HRMAS (1)H NMR) combined with principal components analysis (PCA) and evaluate the radiotherapeutic effect. METHODS: The nude mouse xenograft model of human pancreatic cancer was established by injecting human pancreatic cancer cell SW1990 subcutaneously into the nude mice. When the tumors volume reached 800 mm(3), the mice received various radiation doses. Two weeks later, tumor tissue sections were prepared for running the NMR measurements. (1)H NMR and PCA were used to determine the changes in the metabolic profiles of tumor tissues after radiotherapy. Metabolic profiles of normal pancreas, pancreatic tumor tissues, and radiation- treated pancreatic tumor tissues were compared. RESULTS: Compared with (1)H NMR spectra of the normal nude mouse pancreas, the levels of choline, taurine, alanine, isoleucine, leucine, valine, lactate, and glutamic acid of the pancreatic cancer group were increased, whereas an opposite trend for phosphocholine, glycerophosphocholine, and betaine was observed. The ratio of phosphocholine to creatine, and glycerophosphocholine to creatine showed noticeable decrease in the pancreatic cancer group. After further evaluation of the tissue metabolic profile after treatment with three different radiation doses, no significant change in metabolites was observed in the (1)H NMR spectra, while the inhibition of tumor growth was in proportion to the radiation doses. However, PCA results showed that the levels of choline and betaine were decreased with the increased radiation dose, and conversely, the level of acetic acid was dramatically increased. CONCLUSION: The combined methods were demonstrated to have the potential for allowing early diagnosis and assessment of pancreatic cancer response to radiotherapy.

Reactivity of a bis(amidinato)iron(II) complex [Fe(MesC(NPr(i))2)2] toward some oxidizing reagents.

A diversified reactivity of the mononuclear bis(amidinato)iron(II) complex [Fe(MesC(NPr(i))2)2] (1) toward oxidizing reagents has been disclosed. The bis(amidinato)iron(II) complex was synthesized from the reaction of [Fe(Mes)2]2 with 4 equiv of diisopropyl carbodiimide in good yield. Treatment of 1 with 1 equiv of benzyl chloride gives the high-spin ferric complex [FeCl(MesC(NPr(i))2)2] (2), with 0.25 equiv of S8 affords the sulfur-insertion product [Fe(MesC(NPr(i))(NPr(i)S))2] (3), with 1 equiv of 3,5-dimethylphenyl azide or phenyl azide yields nitrene-insertion product [Fe(MesC(NPr(i))2)(Pr(i)NC(Mes)N(Pr(i))NAr)] (Ar = 3,5-dimethylphenyl, 4a; phenyl, 4b), and with 1 equiv of oxo-transfer reagent, trimethylamine oxide or 2,6-dichloropyridine oxide, generates the oxo-bridged diferric complex [(MesC(NPr(i))2)2FeOFe(MesC(NPr(i))2)2] (5). Complexes 1-3, 4a, and 5 have been characterized by (1)H NMR, UV-vis, IR, elemental analysis, and single-crystal X-ray diffraction studies. The formations of these unusual sulfur- and nitrene-insertion products 3, 4a, and 4b, can be explained by the sequential redox reaction between 1 and the oxidants, followed by migratory insertion steps.

pH-responsive supramolecular polymerization in aqueous media driven by electrostatic attraction-enhanced crown ether-based molecular recognition.

All the previously reported supramolecular polymers based on crown ether-based molecular recognition have been prepared in anhydrous organic solvents. This is mainly due to the weakness of crown ether-based molecular recognition in the presence of water. Here we report a linear supramolecular polymer constructed from a heteroditopic monomer in an aqueous medium driven by crown ether-based molecular recognition through the introduction of electrostatic attraction. In addition, the reversible transition between the linear supramolecular polymer and oligomers is achieved by adding acid and base. This study realizes the breakthrough of the solvent for supramolecular polymerization driven by crown ether-based molecular recognition from anhydrous organic solvents to aqueous media. It is helpful for achieving supramolecular polymerization driven by crown ether-based molecular recognition in a completely aqueous medium.

Effects of gadolinium chelate on the evolution of the nanoscale structure in peptide hydrogels.

Small-angle X-ray scattering (SAXS) was used to monitor peptide hydrogelation with and without the MRI contrast agent gadolinium chelate (Gd(III)-chelate). The gelators are a pair of decapeptides that are self-repulsive but mutually attractive. Gd(III)-chelate was either free or covalently bound to one of the decapeptides. Free and conjugated Gd(III)-chelate have the opposite effects on peptide gelation: free Gd(III)-chelate slows down gelation while having little effect on the cross-sectional area of peptide fibers; covalently conjugated Gd(III)-chelate speeds up gelation and results in peptide fibers with significantly greater cross-sectional area. After 24 h of gelation, the cross-sectional areas of hydrogels with no Gd(III)-chelate, with free Gd(III)-chelate and with conjugated Gd(III) chelate are 3700, 3800, and 6300 Å(2), respectively. Free Gd(III)-chelate is not incorporated into peptide fibers and remains in solution with little effect on MRI intensity upon gelation. In contrast, covalently conjugated Gd(III)-chelate is not only incorporated into peptide fibers, but further aggregates toward the center of the peptide fibers. In conclusion, to visualize hydrogelation using (1)H MRI, it is necessary to conjugate Gd(III)-chelate to the material covalently and use T(2)-weighted imaging.

Understanding the binding interactions between dendrimer and 18 common amino acids by NMR techniques.

In the present study, we focus on the interactions between poly(propylene imine) (PPI) dendrimer and 18 of the 20 common amino acids by several NMR techniques, including NMR titrations and NOESY analysis. Surface ionic interactions and interior encapsulations were observed, and the binding behavior of amino acids with PPI dendrimer depends much on the side-chain properties of the amino acid, such as charge and hydrophobic/hydrophilic properties. The (1)H NMR titration results show that the formation of PPI dendrimer-amino acid complexes are driven mainly by ionic interactions for all the amino acids except tryptophan, which is involved in strong hydrophobic interactions with the interior pockets of PPI. The hydrophobic encapsulation of tryptophan in PPI pockets is confirmed by NOESY analysis. Amino acids with negatively charged residues much more easily saturate the surface charges on PPI than amino acids with uncharged residues, whereas amino acids with positively charged residues are the most difficult to bind with the surface amine groups on the PPI dendrimer. A simultaneous occurrence of interior encapsulation (hydrophobic, hydrogen bond, or ionic interactions) and surface binding (ionic interactions) was observed for tryptophan, phenylalanine, arginine, lysine, histidine, cysteine, and asparagine, and a preferential surface ionic binding on the PPI surface rather than encapsulations in the interior was obtained for the other amino acids.

Formation of polypseudorotaxane networks by cross-linking the quadruple hydrogen bonded linear supramolecular polymers via bisparaquat molecules.

The creation of novel crown ether-paraquat polypseudorotaxane networks, constructed by bisparaquat monomers threading into the cavity of the crown ether units of linear supramolecular polymers that are formed based on the quadruple hydrogen bonded unit ureidopyrimidinone (Upy) in the concentrated solution, is described.

Sol and gel states in peptide hydrogels visualized by Gd(III)-enhanced magnetic resonance imaging.

The hydrogels assembled from a pair of self-repulsive but mutually attractive decapeptides are visualized by magnetic resonance imaging (MRI). It is found that in the absence of Gd(III)-chelate, gelation has little effect on MRI signal intensity. In the presence of Gd(III)-chelate, gelation leads to significant changes in water relaxation and MR signal intensity. The sol to gel transition is best visualized by T2-weighted imaging using large echo time with the sol producing a bright spot and the gel producing a dark spot. MRI studies point to high local Gd(III)-chelate concentration. Small-angle X-ray scattering study indicates that this local enrichment of Gd(III)-chelate has two contributing processes: first, the aggregation of peptides into fibers; second, within peptide fibers, Gd(III)-chelate further aggregate into clusters. This work demonstrates that the status of peptide-based hydrogels can be visualized by MRI with the aid of covalently linked Gd(III)-chelates. This result has implications for monitoring peptide scaffolds in vivo.

Discrimination of metabolic profiles of pancreatic cancer from chronic pancreatitis by high-resolution magic angle spinning 1H nuclear magnetic resonance and principal components analysis.

The metabolic profiles of Sprague-Dawley rat pancreases were investigated by high-resolution magic angle spinning proton magnetic resonance spectroscopy ((1)H NMR) combined with principal components analysis (PCA) to discriminate pancreatic cancer from chronic pancreatitis. Intact pancreatic tissue samples were obtained from Sprague-Dawley rats with histologically proven pancreatic cancer (n = 5), chronic pancreatitis (n = 5), and two matched controls (n = 5 per group). Two (1)H NMR experiments, single-pulse and Carr-Purcell-Meiboom-Gill, were carried out separately. Increases in phosphocholine and glycerophosphocholine levels and decreases in leucine, isoleucine, valine, lactate and alanine levels were observed in chronic pancreatitis, whereas the opposite trends were observed in pancreatic cancer. Increasing taurine and decreasing betaine were found both in chronic pancreatitis and in pancreatic cancer. Additionally, the lipid content in pancreatic cancer was higher than that in chronic pancreatitis. PCA was carried out for the single-pulse and Carr-Purcell-Meiboom-Gill (1)H NMR spectra, respectively, to visualize separation among the samples and to extract characteristic metabolites of pancreatic cancer and chronic pancreatitis. Decreased phosphocholine and glycerophosphocholine were suggested as unique metabolite indicators of pancreatic cancer. Furthermore, even with the disturbance of various quantities of lipid contents pancreatic cancer and chronic pancreatitis could be differentiated well by the combination of high-resolution magic angle spinning (1)H NMR and PCA. Thus this combination was demonstrated to have the potential to improve magnetic resonance spectroscopy for positive early diagnosis of pancreatic cancer in clinical settings.

Peptide-based viscoelastic matrices for drug delivery and tissue repair.

Molecular self-assembly has paved the way to create novel, supramolecular, functional biomaterials. Peptide-based biomaterials are gaining interest as a result of their programmability, biodegradability, and bioresorbability. Further, unlike polymeric materials, peptides can be made monodisperse with precise control over sequence, chain length, and stereochemistry. Peptide-based viscoelastic matrices have been designed and characterized for various biomedical applications, such as tissue engineering scaffolds or drug delivery vehicles. The 'holy grail' in designing an ideal tissue engineering scaffold lies in mimicking the cues of the tissue's natural extracellular matrix (ECM). Some of the key elements of ECM that are incorporated into these peptide scaffolds include cell-adhesive and protease-sensitive sequences for enhanced cell-cell and cell-biomaterial interactions. Peptide-based viscoelastic matrices can also be engineered with drug carrying protease-sensitive sequences for controlled and site-specific drug delivery. Molecular-level engineering of simple oligopeptide modules can be used to control the position and density of the bio-mimetic functionalities in the supramolecular structures, which demonstrates the power of the 'bottom-up' approach in self-assembly.