Injection of Porcine Adipose Tissue-Derived Stromal Cells by a Novel Waterjet Technology

Previously, we developed a novel, needle-free waterjet (WJ) technology capable of injecting viable cells by visual guided cystoscopy in the urethral sphincter. In the present study, we aimed to investigate the effect of WJ technology on cell viability, surface markers, differentiation and attachment capabilities, and biomechanical features. Porcine adipose tissue-derived stromal cells (pADSCs) were isolated, expanded, and injected by WJ technology. Cell attachment assays were employed to investigate cell–matrix interactions. Cell surface molecules were analyzed by flow cytometry. Cells injected by Williams Needle (WN), normal cannula, or not injected cells served as controls. Biomechanical properties were assessed by atomic force microscopy (AFM). pADSCs injected by the WJ were viable (85.9%), proliferated well, and maintained their in vitro adipogenic and osteogenic differentiation capacities. The attachment of pADSCs was not affected by WJ injection and no major changes were noted for cell surface markers. AFM measurements yielded a significant reduction of cellular stiffness after WJ injections (p < 0.001). WJ cell delivery satisfies several key considerations required in a clinical context, including the fast, simple, and reproducible delivery of viable cells. However, the optimization of the WJ device may be necessary to further reduce the effects on the biomechanical properties of cells.     

High‐yield secretion of recombinant gelatins by Pichia pastoris

Recombinant non‐hydroxylated gelatins based on mouse type I and rat type III collagen sequences were secreted from the methylotrophic yeast Pichia pastoris, using the Saccharomyces cerevisiae α‐mating factor prepro signal. Proteolytic degradation could be minimized to a large extent by performing fermentations at pH 3·0 and by adding casamino acids to the medium, even though gelatin is extremely susceptible to proteolysis due to its open, unfolded structure. Proteolytic cleavage at specific mono‐arginylic sites, by a putative Kex2‐like protease, could be successfully abolished by site‐directed mutagenesis of these sites. Production levels as high as 14·8 g/l clarified both were obtained, using multicopy tranformants. To our knowledge, this represents the highest level of heterologous protein secretion reported to date for P. pastoris. Copyright © 1999 John Wiley & Sons, Ltd.     

Affinity and Specificity for Binding to Glycosaminoglycans Can Be Tuned by Adapting Peptide Length and Sequence

Although glycosaminoglycan (GAG)–protein interactions are important in many physiological and pathological processes, the structural requirements for binding are poorly defined. Starting with GAG-binding peptide CXCL9(74-103), peptides were designed to elucidate the contribution to the GAG-binding affinity of different: (1) GAG-binding motifs (i.e., BBXB and BBBXXB); (2) amino acids in GAG-binding motifs and linker sequences; and (3) numbers of GAG-binding motifs. The affinity of eight chemically synthesized peptides for various GAGs was determined by isothermal fluorescence titration (IFT). Moreover, the binding of peptides to cellular GAGs on Chinese hamster ovary (CHO) cells was assessed using flow cytometry with and without soluble GAGs. The repetition of GAG-binding motifs in the peptides contributed to a higher affinity for heparan sulfate (HS) in the IFT measurements. Furthermore, the presence of Gln residues in both GAG-binding motifs and linker sequences increased the affinity of trimer peptides for low-molecular-weight heparin (LMWH), partially desulfated (ds)LMWH and HS, but not for hyaluronic acid. In addition, the peptides bound to cellular GAGs with differential affinity, and the addition of soluble HS or heparin reduced the binding of CXCL9(74-103) to cellular GAGs. These results indicate that the affinity and specificity of peptides for GAGs can be tuned by adapting their amino acid sequence and their number of GAG-binding motifs.     

The effects of microwave heating on the kinetics of isothermal dehydration of equilibrium swollen poly(acrylic‐co‐methacrylic acid) hydrogel

The isothermal dehydration of poly(acrylic‐co‐methacrylic acid) (PAM) hydrogel under microwave heating (MWH) was investigated. The isothermal kinetics curves of the PAM hydrogel dehydration at temperature range from 293 K to 333 K were recorded. Based on the differential isoconversion method it was concluded that the microwave dehydration of poly(acrylic‐co‐methacrylic acid) hydrogel is an elementary kinetics process. Applying the model‐fitting method it was established that the kinetics of microwave isothermal hydrogel dehydration can be described by the kinetics model of the phase‐boundary controlled process (contracting area). The values of the kinetics parameters (activation energy (E_a) and preexponential factor (lnA)) of the dehydration process under microwave heating are lower than the values for conventional heating (CH). The established influence of MWH on the kinetics of hydrogel dehydration is explained with a specific activation mechanism of water molecules for dehydration and with the increase in the value of the energy of the ground level of the resonant oscillator of water molecule (v = 837 cm^?1) due to the absorption of microwave energy. POLYM. ENG. SCI., 56:87–96, 2016. © 2015 Society of Plastics Engineers     

Screening of protease inhibitors as antiplasmodial agents. Part I: Aziridines and epoxides

A broad protease-based and cell-based screening of protease inhibitors yielded the aziridine-2-carboxylic acid derivative 2 a and the N-acylated aziridine-2,3-dicarboxylic acid derivatives 32 a and 34 b as the most potent inhibitors of falcipain-2 and falcipain-3 (IC(50) falcipain-2: 0.079-5.4 microM, falcipain-3: 0.25-39.8 microM). As the compounds also display in vitro activity against the P. falciparum parasite in the submicromolar and low micromolar range, these compound classes are leads for new antiplasmodial falcipain inhibitors.     

Sperm Physiological Response to Female Serum—Potential New Insights into the Reproductive Incompatibility Diagnostics

Infertility is assumed to arise exclusively from male- and female-dependent pathological factors. However, recent studies have indicated that reproductive failure may also result from the reproductive incompatibility of the partners. Selection against such incompatibilities likely occurs via female-derived reproductive secretions, including follicular fluid (FF), that mediate gamete-level mate choice towards the sperm of specific males. To facilitate potential development of diagnostic tests for human reproductive incompatibility, we examined whether sperm physiological response to female serum indicate male–female compatibility in the presence of FF. We performed a full-factorial experiment, in which the sperm of 10 males were treated with the FF and serum of 6 healthy females. We found that sperm motility and viability in both biofluids were highly similar and that in 70% of the males, sperm serum treatment predicted male–female compatibility. We also identified male human leucocyte antigen (HLA) alleles and female (FF and serum) anti-HLA antibodies and tested whether the number of allele–antibody matches predict sperm physiological response to female fluids. However, no association was found between measured sperm traits and the number of allele–antibody matches. Overall, the present results may open novel possibilities for the future development of reproductive incompatibility tests and may pave the way towards more accurate infertility diagnostics and treatments.     

Computational calibration method for optical tomography

We propose a computational calibration method for optical tomography. The model of the calibration scheme is based on the rotation symmetry of source and detector positions in the measurement setup. The relative amplitude losses and phase shifts at the optic fibers are modeled by complex-valued coupling coefficients. The coupling coefficients can be estimated when optical tomography data from a homogeneous and isotropic object are given. Once these coupling coefficients have been estimated, any data measured with the same measurement setup can be corrected for the relative variation in the data due to source and detector losses. The final calibration of the data for the source and detector losses and the source calibration between the data and the forward model are obtained as part of the initial estimation for reconstruction. The calibration method was tested with simulations and measurements. The results show that the coupling coefficients of the sources and detectors can be estimated with good accuracy. Furthermore, the results show that the method can significantly improve the quality of reconstructed images.     

Predicting practical fitness to drive in drivers with visual field defects caused by ocular pathology

Vision, viewing efficiency, visual attention, and on-road driving performance were assessed in 100 participants with central and/or peripheral visual field defects caused by ocular pathology. Driving was evaluated by the Dutch driving license authority making use of the protocol for investigating practical fitness to drive. A smaller percentage of participants with central visual field defects passed the on-road driving test, in comparison with participants with peripheral or mild field defects. The predictive power of a model based on the current vision requirements for driving significantly increased when taking compensatory viewing efficiency into account. The results of the latter model were comparable to those of a model based on tests of visual attention and contrast sensitivity. Despite the increased explained variance of practical fitness to drive when taking higher-order visual functions into account, sensitivity and specificity remained quite low, limiting the use of these tests in identifying unfit drivers. Actual or potential applications of this research include the development of training programs to improve practical fitness to drive in drivers with visual field defects.     

First-principles quantum chemistry in the life sciences

The area of computational quantum chemistry, which applies the principles of quantum mechanics to molecular and condensed systems, has developed drastically over the last decades, due to both increased computer power and the efficient implementation of quantum chemical methods in readily available computer programs. Because of this, accurate computational techniques can now be applied to much larger systems than before, bringing the area of biochemistry within the scope of electronic-structure quantum chemical methods. The rapid pace of progress of quantum chemistry makes it a very exciting research field; calculations that are too computationally expensive today may be feasible in a few months' time! This article reviews the current application of 'first-principles' quantum chemistry in biochemical and life sciences research, and discusses its future potential. The current capability of first-principles quantum chemistry is illustrated in a brief examination of computational studies on neurotransmitters, helical peptides, and DNA complexes.