Targeted protein delivery: carbodiimide crosslinking influences protein release from microparticles incorporated within collagen scaffolds

open access articleTissue engineering response may be tailored via controlled, sustained release of active agents from protein-loaded degradable microparticles incorporated directly within three-dimensional (3D) ice-templated collagen scaffolds. However, the effects of covalent crosslinking during scaffold preparation on the availability and release of protein from the incorporated microparticles have not been explored. Here, we load 3D ice-templated collagen scaffolds with controlled additions of poly-(DL-lactide-co-glycolide) microparticles. We probe the effects of subsequent N-(3-dimethylaminopropyl)- N0-ethylcarbodiimide hydrochloride crosslinking on protein release, using microparticles with different internal protein distributions. Fluorescein isothiocyanate labelled bovine serum albumin is used as a model protein drug. The scaffolds display a homogeneous microparticle distribution, and a reduction in pore size and percolation diameter with increased microparticle addition, although these values did not fall below those reported as necessary for cell invasion. The protein distribution within the microparticles, near the surface or more deeply located within the microparticles, was important in determining the release profile and effect of crosslinking, as the surface was affected by the carbodiimide crosslinking reaction applied to the scaffold. Crosslinking of microparticles with a high proportion of protein at the surface caused both a reduction and delay in protein release. Protein located within the bulk of the microparticles, was protected from the crosslinking reaction and no delay in the overall release profile was seen

Simultaneous and accurate measurement of the dielectric constant at many frequencies spanning a wide range

We present an innovative technique which allows the simultaneous measurement of the dielectric constant of a material at many frequencies, spanning a four orders of magnitude range chosen between 10 --2 Hz and 10 4 Hz. The sensitivity and accuracy are comparable to those obtained using standard single frequency techniques. The technique is based on three new and simple features: a) the precise real time correction of the amplication of a current amplier; b) the specic shape of the excitation signal and its frequency spectrum; and c) the precise synchronization between the generation of the excitation signal and the acquisition of the dielectric response signal. This technique is useful in the case of relatively fast dynamical measurements when the knowledge of the time evolution of the dielectric constant is needed

Lignin: A Biopolymer from Forestry Biomass for Biocomposites and 3D Printing

Biopolymers from forestry biomass are promising for the sustainable development of new biobased materials. As such, lignin and fiber-based biocomposites are plausible renewable alternatives to petrochemical-based products. In this study, we have obtained lignin from Spruce biomass through a soda pulping process. The lignin was used for manufacturing biocomposite filaments containing 20% and 40% lignin and using polylactic acid (PLA) as matrix material. Dogbones for mechanical testing were 3D printed by fused deposition modelling. The lignin and the corresponding biocomposites were characterized in detail, including thermo-gravimetric analysis (TGA), differential scanning calorimetry (DSC), Fourier transform infrared (FTIR) spectroscopy, X-ray diffraction analysis (XRD), antioxidant capacity, mechanical properties, and scanning electron microscopy (SEM). Although lignin led to a reduction of the tensile strength and modulus, the reduction could be counteracted to some extent by adjusting the 3D printing temperature. The results showed that lignin acted as a nucleating agent and thus led to further crystallization of PLA. The radical scavenging activity of the biocomposites increased to roughly 50% antioxidant potential/cm2, for the biocomposite containing 40 wt % lignin. The results demonstrate the potential of lignin as a component in biocomposite materials, which we show are adequate for 3D printing operations

Diagrammar In Classical Scalar Field Theory

In this paper we analyze perturbatively a g phi^4 classical field theory with and without temperature. In order to do that, we make use of a path-integral approach developed some time ago for classical theories. It turns out that the diagrams appearing at the classical level are many more than at the quantum level due to the presence of extra auxiliary fields in the classical formalism. We shall show that several of those diagrams cancel against each other due to a universal supersymmetry present in the classical path integral mentioned above. The same supersymmetry allows the introduction of super-fields and super-diagrams which considerably simplify the calculations and make the classical perturbative calculations almost "identical" formally to the quantum ones. Using the super-diagrams technique we develop the classical perturbation theory up to third order. We conclude the paper with a perturbative check of the fluctuation-dissipation theorem.Comment: 67 pages. Improvements inserted in the third order calculation

Targeted protein delivery: carbodiimide crosslinking influences protein release from microparticles incorporated within collagen scaffolds.

Tissue engineering response may be tailored via controlled, sustained release of active agents from protein-loaded degradable microparticles incorporated directly within three-dimensional (3D) ice-templated collagen scaffolds. However, the effects of covalent crosslinking during scaffold preparation on the availability and release of protein from the incorporated microparticles have not been explored. Here, we load 3D ice-templated collagen scaffolds with controlled additions of poly-(DL-lactide-co-glycolide) microparticles. We probe the effects of subsequent N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride crosslinking on protein release, using microparticles with different internal protein distributions. Fluorescein isothiocyanate labelled bovine serum albumin is used as a model protein drug. The scaffolds display a homogeneous microparticle distribution, and a reduction in pore size and percolation diameter with increased microparticle addition, although these values did not fall below those reported as necessary for cell invasion. The protein distribution within the microparticles, near the surface or more deeply located within the microparticles, was important in determining the release profile and effect of crosslinking, as the surface was affected by the carbodiimide crosslinking reaction applied to the scaffold. Crosslinking of microparticles with a high proportion of protein at the surface caused both a reduction and delay in protein release. Protein located within the bulk of the microparticles, was protected from the crosslinking reaction and no delay in the overall release profile was seen.This work was supported by the European Research Council [ERC Advanced Grant 320598 3D-E] and was also funded by a grant from the Medical Research Council, Arthritis Research UK, Reumafonds and the UKRM

Electric single-molecule hybridization detector for short DNA fragments

By combining DNA nanotechnology and high-bandwidth single-molecule detection in nanopipets, we demonstrate an electric, label-free hybridization sensor for short DNA sequences (<100 nucleotides). Such short fragments are known to occur as circulating cell-free DNA in various bodily fluids, such as blood plasma and saliva, and have been identified as disease markers for cancer and infectious diseases. To this end, we use as a model system an 88-mer target from the RV1910c gene in Mycobacterium tuberculosis, which is associated with antibiotic (isoniazid) resistance in TB. Upon binding to short probes attached to long carrier DNA, we show that resistive-pulse sensing in nanopipets is capable of identifying rather subtle structural differences, such as the hybridization state of the probes, in a statistically robust manner. With significant potential toward multiplexing and high-throughput analysis, our study points toward a new, single-molecule DNA-assay technology that is fast, easy to use, and compatible with point-of-care environments

Molybdenum-99 (99Mo) Adsorption Profile of Zirconia-Based Materials for 99Mo/99mTc Generator Application

Technetium-99m (99mTc) plays a major role in diagnostic nuclear medicine and has not yet been replaced with any other radionuclides. It is available through the 99Mo/99mTc generator. The use of low-specific-activity 99Mo for 99Mo/99mTc generator application requires high adsorptive capacity sorbents. This study focused on the determination of 99Mo adsorption capacity of several zirconia materials, namely monoclinic nanozirconia, orthorhombic nanozirconia, sulfated zirconia,   and phosphated zirconia. These materials were synthesized by using the sol-gel method and characterized using FT-IR spectroscopy, X-ray diffraction (XRD), and scanning electron microscopy/energy-dispersive X-ray spectroscopy (SEM-EDS). The determination of 99Mo adsorption capacity of these materials was carried out by soaking the materials in a Na299MoO4 solution with pH of 3 and 7, at temperatures ranging from room temperature to 90 °C, for 1 and 3 hours. The results indicated that monoclinic nanozirconia has a 99Mo adsorption capacity of 76.9 mg Mo/g, whereas orthorhombic nanozirconia, sulfated zirconia, and phosphated zirconia have 99Mo adsorption capacities of 150.1 mg Mo/g, 15.58 mg Mo/g, and 12.74 mg Mo/g, respectively. It appears that orthorhombic nanozirconia has the highest 99Mo adsorption capacity among the synthesized materials and can be applied as a candidate material for the 99Mo/99mTc generator

Biochemical profile of the proteolytic processes in blood serum and in the prostatic lodge fluid at the patients with transvesical adenomectomy with the usage of endourethral drainage

IP USMF "Nicolae Testemiţanu", IMSP Spitalul Clinic Municipal "Sf. Treime", Secţia Urologie, Al VI-lea Congres de Urologie, Dializă şi Transplant Renal din Republica Moldova cu participare internaţională (21-23 octombrie 2015)Studiului constă în evaluarea proceselor proteolitice în serul sanguin în perioada pre - şi postoperatorie la pacienţii cu AP şi în eliminările din loja după adenomectomie întru determinarea rolului lorpredictiv în evoluţia proceselor lezionare concomitente şi monitorizării hemostazeiîn loja prostatică utilizând drenarea endouretrală a lo je i. Cercetările actualului studiu sunt de tip prospectiv fiind bazate pe un eşantion de 65 bărbaţi, conlocuitori ai R. Moldova, cu vârsta între 50-75 ani. Eşantionul general de studiu a fost divizat în două loturi specifice Lf - lotul de cercetare cu AP (n=45)şi LM - lotul m artor(n=20). în lotul de cercetare, sa inclus pacienţi cu AP rezolvat chirurgical prin metoda transvezicală în modificarea elaborată şi propusă de noi, [3] (fig.1). Sa constatat că în cadrul evoluţiei AP are loc creşterea pronunţată a funcţionalităţii majorităţii enzimelor proteolitice atât în serul sanguin cât şi în eliminările din lojă. Sa stabilit unii parametrii aşa ca SN şi MMM în serul sanguin şi la nivel de lojă, servind drept marcheri ale prezenţei intoxicaţiei endogene şi toxicităţii din serul sanguin ca rezultat a proceselor proteolitice derulate în cadrul AP şi postoperatoriu. Este relevantă importanţa rezultatelor obţinute privind sensibilitatea diferită a proteazelor acide în AP, fapt ce confirmă că utilizarea drenării lojei deschide noi posibilităţi a unei tactici medico-chirurgicale şi terapeutice alternative în optimizarea unei hemostaze la nivel de lojă.Summary The aim of study is to appreciate proteolytic processes both in blood serum and the lodge drainage fluid in the pre-and post-operative period in patients with BPH after adenomectomy in order to identify their predictive role in the development of concomitant injurious processes and monitoring of the hemostasis in the prostatic lodge using endourethral drainage. Current research is a prospective study based on a sample of 65 men from the Republic of Moldova, aged 50-75 years. General study sample was divided into two specific lots: Rg - research group with PA, (n = 45) and Cg - control group (n = 20). The research group included patients with PA surgically solved by the changed transvesical method developed and proposed by us. It was found that in the evolution of PA most proteolytic enzymes functionality is increasing both in the blood serum and in the fluid drained from the lodge. Were determined some parameters such as SN and MMM in blood serum and in the lodge, serving as markers of endogenous intoxication presence and toxicity of blood serum as a result of proteolytic processes developed within BPH and postoperatively. The results revealed the importance of the different sensitivity of the acidic proteases in BPH, which confirms that the use of lodge drainage opens new possibilities of alternative surgical and therapeutic tactics to optimize the hemostasis in the prostatic lodge

Tratamentul litiazei vezicii urinare prin litotriţie endoscopica mecanică

Summary Lithiasis of the bladder is known from antiquity, beeing one of the first diseases wich profits by surgical treatment. The discovery of lithotripsy had spreaded the indication of endoscopic treatment. In the period of 1998-2006 we practiced the mecanical endoscopy in 30% of patients with lithiasis of the bladder. The endoscopic lithotripsy is the initial method of therary in the treatment of lithiasis of the bladder. The endoscopic acces alow to solve also another diseases of the lower urinar system as TURP, TURB, TUIP and other. Introducere. Litiaza vezicii urinare este cunoscută din antichitate, fiind una dintre primele maladii care au beneficiat de tratament chirurgical, iar operaţia de cistolitotomie, fiind pâna la finele secolului XIX unica metodă de ablaţie a calcului. Până în anii 60 ai secolului trecut litotriţia endoscopică era efectuată prin metoda oarbă, fiind grevată de numeroase incidente şi complicaţii intraoperatorii. Apariţia litoriptoarelor controlabile optic Hendrixon şi Punch, iar apoi a celor electorhidraulice şi ultrasonice au largit cu mult indicaţiile tratamentului endoscopic. Scopul lucrării. Analiza experienţei personale a tratamentului litiazei vezicii urinare prin metode endoscopice mecanice. Material şi metodă. În perioada 1998-2006 am practicat litotriţia endoscopică mecanică la 81 pacienţi, 68 bărbaţi şi 13 femei, cea ce reprezintă 30% din numărul pacienţilor cu litiază a vezicii urinare, internaţi în Clinica Urologie a SCR. Dimensiunile calculului au variat de la 0,6 la 3,5 cm, cu o perioadă simptomatică de la 1 lună la 3 ani. Am efectuat litotriţia mecanică cu respectarea celor 3 faze ale litotirţiei vezicale: fragmentarea primară a calculului, prelucrarea şi evacurarea fragmentelor. În situaţia când calculul a avut dimensiuni sub 2,5 cm am efectuat toate cele 3 etape cu ajutorul litoriptorului Punch. Dacă calculul a avut dimensiuni mai mari, am practicat iniţial fragmentarea primară cu ajutorul litotriptorului mecanic Hendrixon, care are o vizibilitate mai redusă şi o manevriabilitate mai greoaie, din cauza branşelor mai mari ale instrumentului. Evacuarea fragmentelor cu ajutorul litoriptorului Punch a fost facilă, datorită construcţiei sale speciale, cu un orificiu central al extermităţii active, fapt ce permite aspirarea totală şi orientată şi a fragmentelor impregnate în mucoasă, evită leziunile peretelui vezicii urinare, apelând destul de rar la instrumentar suplimentar, precum seringa Guyon sau pompa de aspiraţie. Nu am abordat deliberat calculi cu dimensiuni mai mari de 3,5 cm, supuşi cistolitotomiei. Rezultate. Dintre 68 cazuri de litiază vezicală la bărbaţi, 60 (88%) au avut o litiază secundară. La 23 pacienţi (39%) în aceeaşi şedinţă operatorie am practicat şi alte intervenţii endoscopice (TUR P, ITUP, TURV, uretrotomie optică internă). Intervenţia chirurgicală a fost efectuată sub anestezie rahidiană (50 pacienti, 62%), intravenoasă (29 pacienţi, 35%), locală (2 pacienţi, 3%). Durata intervenţiei a variat de la 10 minute la 1 ora, cu o medie de 25 minute. În 21 cazuri (26%) litotriţia propriu zisa a fost precedată de meatotomia meatului uretral extern, pentru a facilita introducerea tecii de 26 Ch. În perioada postoperatorie în 43 (53%) cazuri nu am aplicat sonda uretro-vezicală permanentă, pacientul fiind externat la un interval de 24-48 ore de la intervenţie. În cazul intervenţiilor dificlie, prelungite sau ale celor asociate cu alte manevre endoscopice, am menţinut drenajul uretro-vezical permanent pe o durată de 3-4 zile, pacientul fiind externat la un interval maxim de 5 zile de la intervenţie. Durata medie de spitalizare a pacienţilor a fost de 3,5 zile. Concluzii. Litoriţia endoscopică este metoda terapeutică de primă intenţie în tratamentul litiazei vezicii urinare. Abordul endoscopic permite rezolvarea concomitenă şi a altor maladii ale aparatului urinar inferior (adenom de prostată, scleroza coluli vezicii urinare, strictura uretrei, etc.). În situaţiile când calculul are dimensiuni sub 2,5 cm, şi/sau nu este complet neted, preferăm efectuarea litotiţiei Punch. Cistolitotomia îşi pastrează locul în arsenalul terapeutic al litiazei vezicale şi este rezervată calculilor cu dimensiuni mari, peste 3,5 cm, excesiv de duri, cu o suprafaţă netedă sau în situaţii particulare, precum adenomul de prostată voluminos