Permeation enhancer effect of chitosan and chitosan derivatives: comparison of formulations as soluble polymers and nanoparticulate systems on insulin absorption in Caco-2 cells.

In this study four quaternized derivatives of chitosan: trimethyl chitosan (TMC), dimethylethyl chitosan (DMEC), diethylmethyl chitosan (DEMC) and triethyl chitosan (TEC) with degree of substitution of approximately 50+/-5% were synthesized and their effect on the permeability of insulin across intestinal Caco-2 monolayers was studied and compared with chitosan both in free-soluble form and in nanoparticulate systems. Transepithelial electrical resistance (TEER) studies revealed that all four chitosan derivatives in free-soluble forms were able to decrease the TEER value in the following order TMC>DMEC>DEMC=TEC>chitosan, indicating their abilities to open the tight junctions. Recovery studies on the TEER showed that the effect of the polymers on Caco-2 cell monolayer is reversible and proves the viability of cells after incubation with all polymers. A similar rank order was also observed when measuring the zeta-potentials of the various polymers in solution form. Transport studies of insulin together with the soluble polymers across Caco-2 cell layers showed the following ranking: TMC>DMEC>DEMC>TEC>chitosan which is in agreement with the strength of the cationic charge of the polymer. In comparison to the free-soluble polymers, the nanoparticles prepared by ionic gelation of the chitosan and its quaternized derivatives had much lower effect on decreasing the TEER by opening of the tight junctions. This can be explained by the reduced available amount of positive charge at the surface of the nanoparticles. In accordance with these results, the insulin loaded nanoparticles showed much less permeation across the Caco-2 cell monolayer in comparison to the free-soluble polymers. Mass balance transport studies revealed that a substantial amount of the nanoparticles has been entrapped into the Caco-2 monolayer or attached to the cell surface. It can thus be stated that while free-soluble polymers can reversibly open the tight junctions and increase the permeation of insulin, the nanoparticles had basically only a low effect on the opening of the tight junction and the paracellular transport of insulin across the Caco-2 cell monolayer. These data convincingly show that nanoparticles consisting of chitosan and its quaternary ammonium derivatives loaded with insulin are less effective in facilitating paracellular transport across Caco-2 cell monolayers than the corresponding free polymers.

Accuracy of non-invasive ICP assessment can be increased by an initial individual calibration.

OBJECTIVE: In a formerly introduced mathematical model, intracranial pressure (ICP) could be non-invasively assessed using cerebral blood flow velocity (FV) and arterial blood pressure (ABP). The current study attempts to check whether the accuracy of the non-invasive ICP assessment (nICP) improves after an initial individual calibration by implanted ICP probes. METHODS: Thirteen patients with brain lesions (35-77 years, mean: 58 +/- 13 years) were studied. FV, ABP and ICP signals were recorded at days 1, 2, 4 and 7. nICP was calculated and compared to ICP. In the first recording of each patient the (invasively assessed) ICP signal was used to calibrate the nICP calculation procedure, while the follow-up recordings were used for its validation. FINDINGS: In 11 patients 22 follow-up recordings were performed. The mean deviation between ICP and the original nICP (+/- SD) was 8.3 +/- 7.9 mmHg. Using the calibrated method this deviation was reduced to 6.7 +/- 6.7 mmHg (P < 0.005). CONCLUSIONS: Initial individual calibration of nICP assessment method significantly improves the accuracy of nICP estimation on subsequent days. This hybrid method of ICP assessment may be used in intensive care units in patients with initially implanted ICP probes. After removal of the probes, ICP monitoring can be continued using the calibrated nICP assessment procedure.

Structure and bonding of the multifunctional amino acid L-DOPA on Au(110).

In investigations of the proteins which are responsible for the surface adhesion of the blue mussel Mytilus edulis, an unusually frequent appearance of the otherwise rare amino acid 3-(3,4-dihydroxyphenyl)-L-alanine (L-DOPA) has been observed. This amino acid is thought to play a major role in the mechanism of mussel adhesion. Here we report a detailed structural and spectroscopic investigation of the interface between L-DOPA and a single-crystalline Au(110) model surface, with the aim of understanding fundamentals about the surface bonding of this amino acid and its role in mussel adhesion. Molecular layers are deposited by organic molecular beam deposition (OMBD) in an ultrahigh-vacuum environment. The following experimental techniques have been applied: ex situ Fourier transform infrared spectroscopy (FTIR), X-ray photoelectron spectroscopy (XPS), low-energy electron diffraction (LEED), high-resolution electron energy loss spectroscopy (HREELS), and scanning tunneling microscopy (STM). Vibrational spectra of isolated L-DOPA molecules and the zwitterionic bulk have been calculated using density functional theory (DFT). The predicted modes are assigned to observed spectra, allowing conclusions regarding the molecule-substrate and molecule-molecule interactions at the L-DOPA/Au(110) interface. We find that zwitterionic L-DOPA forms a monochiral, one-domain commensurate monolayer on Au(110), with the catechol rings on top of [110] gold rows, oriented parallel to the surface. The (2 x 1)-Au(110) surface reconstruction is not lifted. The carboxylate group is found in a bidentate or bridging configuration, the amino group is tilted out of the surface plane, and the hydroxyl groups do not dehydrogenate on Au(110). Similar to the case for the bulk, molecules form dimers on Au(110). However, the number of hydrogen bridge bonds between L-DOPA molecules is reduced as compared to the bulk. Thicker layers which are deposited onto the commensurate interface do not order in the bulk structure. In conclusion, our study shows that the aromatic ring system of L-DOPA functions as a surface anchor. Since it is also known that the hydroxyl groups support cross-link reactions between L-DOPA residues in the mussel glue protein, we can conclude that the catechol ring supports surface adhesion of mussel proteins via two independent functions.

Cloning and expression of P60, a conserved surface-localized protein of Mycoplasma hominis, in Escherichia coli.

The clp60 gene encoding P60, a conserved lipoprotein of Mycoplasma hominis, was cloned and sequenced from both the type strain PG21 and the isolate FBG. Both open reading frames were identical in length, comprising 1746 nucleotides. The deduced amino acid sequences differed in 16 out of 582 amino acids. As expected, none of these divergences mapped within the epitope that was recognized by mAb CG4 in all of the 198 isolates of M. hominis analyzed so far. This conserved epitope was narrowed down to amino acids 454 through 464 within the C terminus of P60. For the expression of the recombinant homolog P60, P60rec, in E. coli the TGA codons of clp60 were substituted for TGG codons prior to cloning of clp60 into the expression plasmid pQE41. The expression of P60rec as a fusion protein with dihydrofolate reductase carrying an N-terminal His-tag enabled the purification of large amounts of P60rec in a soluble form.

[Psychosocial factors and cancer. A study of 80 women by means of a psychosocial questionnaire (author's transl)]. / Psychosoziale Faktoren und Krebs. Untersuchung von 80 Frauen mit einem psychosozialen Fragebogen.

The psychosocial questionnaire compiled by C. B. Bahnson and M. B. Bahnson and adapted for the German-speaking area was used to question 40 female cancer patients (30 mammary, 6 gastric and 4 pulmonary carcinomata) aged between 36 and 64 years and 40 control subjects selected by the matched pairs method. The cross section examination showed significant differences (p less than or equal to 0.05) between cancer patients and control subjects on single variate examination in 10 individual variables. Discriminant analysis (multivariate examination) revealed a discriminant function of 12 variables which enabled the classification of the entire collective of subjects with 95 percent accuracy in the carcinoma and control groups. Our results confirm largely the hypotheses developed in the USA.