Preparation of nano-hydroxyapatite/chitosan aqueous dispersions: From lab scale to continuous production using an innovative static mixer.

Chitosan is widely used in the preparation of organic-inorganic composite materials, such as n-HAp/CS composites, which find application for bone regeneration. The methods for their preparation are various, and usually based on the preparation of intermediate n-HAp/CS dispersions, which can greatly influence the final properties of the resulting composites since it is expected that homogenous and stable dispersions lead to composite materials with improved final properties. This work hypothesizes that, additionally to process parameters such as pH, n-HAp/CS weight ratio, mixing conditions and the presence of salts, chitosan itself has a high impact on dispersions stability. Thus, the importance of properly control the preparation of the n-HAp/CS intermediate dispersions is highlighted by doing a systematic study where relevant processing parameters were studied at lab scale using ultrasonication, alone or in the presence of chitosan, namely on particle size and zeta potential. Furthermore, and based on the best laboratorial conditions, the production of n-HAp/CS nanocomposite dispersions in continuous mode was attempted through NETmix® technology, an innovative static mixer and reactor developed at the Associate Laboratory LSRE-LCM of the Faculty of Engineering of the University of Porto (FEUP).

CC chemokine receptor 10 cell surface presentation in melanocytes is regulated by the novel interaction partner S100A10.

The superfamily of G-protein-coupled receptors (GPCR) conveys signals in response to various endogenous and exogenous stimuli. Consequently, GPCRs are the most important drug targets. CCR10, the receptor for the chemokines CCL27/CTACK and CCL28/MEC, belongs to the chemokine receptor subfamily of GPCRs and is thought to function in immune responses and tumour progression. However, there is only limited information on the intracellular regulation of CCR10. We find that S100A10, a member of the S100 family of Ca(2+) binding proteins, binds directly to the C-terminal cytoplasmic tail of CCR10 and that this interaction regulates the CCR10 cell surface presentation. This identifies S100A10 as a novel interaction partner and regulator of CCR10 that might serve as a target for therapeutic intervention.

The Hippo pathway is controlled by Angiotensin II signaling and its reactivation induces apoptosis in podocytes.

The Hippo pathway fulfills a crucial function in controlling the balance between proliferation, differentiation and apoptosis in cells. Recent studies showed that G protein-coupled receptors (GPCRs) serve as upstream regulators of Hippo signaling, that either activate or inactivate the Hippo pathway via the large tumor suppressor kinase (LATS) and its substrate, the co-transcription factor Yes-associated protein (YAP). In this study, we focused on the Angiotensin II type 1 receptor (AT1R), which belongs to the GPCR family and has an essential role in the control of blood pressure and water homeostasis. We found that Angiotensin II (Ang II) inactivates the pathway by decreasing the activity of LATS kinase; therefore, leading to an enhanced nuclear shuttling of unphosphorylated YAP in HEK293T cells. This shuttling of YAP is actin-dependent as disruption of the actin cytoskeleton inhibited dephosphorylation of LATS and YAP. Interestingly, in contrast to HEK293T cells, podocytes, which are a crucial component of the glomerular filtration barrier, display a predominant nuclear YAP localization in vivo and in vitro. Moreover, stimulation with Ang II did not alter Hippo pathway activity in podocytes, which show a deactivated pathway. Reactivation of the LATS kinase activity in podocytes resulted in an increased cytoplasmic YAP localization accompanied by a strong induction of apoptosis. Thus, our work indicates that the control of LATS activation and subsequent YAP localization is important for podocyte homeostasis and survival.

Common exonic missense variants in the C2 domain of the human KIBRA protein modify lipid binding and cognitive performance.

The human KIBRA gene has been linked to human cognition through a lead intronic single-nucleotide polymorphism (SNP; rs17070145) that is associated with episodic memory performance and the risk to develop Alzheimer's disease. However, it remains unknown how this relates to the function of the KIBRA protein. Here, we identified two common missense SNPs (rs3822660G/T [M734I], rs3822659T/G [S735A]) in exon 15 of the human KIBRA gene to affect cognitive performance, and to be in almost complete linkage disequilibrium with rs17070145. The identified SNPs encode variants of the KIBRA C2 domain with distinct Ca(2+) dependent binding preferences for monophosphorylated phosphatidylinositols likely due to differences in the dynamics and folding of the lipid-binding pocket. Our results further implicate the KIBRA protein in higher brain function and provide direction to the cellular pathways involved.

Lung function before and after oxygen diving: a randomized crossover study.

RATIONALE: Breathing oxygen with a partial pressure of > 50 kPa can cause pulmonary oxygen toxicity (POT). Diffusing capacity for carbon monoxide (DL(CO)) is thought to be a more sensitive indicator of POT than vital capacity (VC). Because diffusing capacity can be measured more specifically using nitric oxide (DL(NO)), we hypothesized that DL(NO) is better able to monitor and exclude POT than DL(CO). OBJECTIVE: To compare changes in lung function after oxygen and air dives which include measurement of DL(NO) and DL(CO). METHOD: Eleven healthy male divers (mean age 27.5 +/- 3.1 years) made two immersed dives to 150 kPa for three hours on two separate days, during which they randomly breathed 100% oxygen or air. VC, DL(NO) and DL(CO) were measured six times during a 26-hour period on both days and on a third non-diving day. RESULTS: There were no significant changes in DL(CO), DL(NO) or other diffusing capacity or spirometric parameters after either type of dive. CONCLUSION: Lung function after a single three-hour oxygen dive at a pO2 of approximately 150 kPa is comparable to that after an air dive at the same depth and duration. This suggests that such an oxygen dive does not induce detectable signs of POT. Our hypothesis that DL(NO) is more sensitive than DL(CO) for detection of POT could not be tested because the oxygen exposure did not affect either parameter.

The Golgi matrix protein GM130: a specific interacting partner of the small GTPase rab1b.

To detect specific partners of the small Golgi-localized GTPase rab1b we generated rab1b mutants and used them as bait proteins in yeast two-hybrid screens. We isolated several specifically interacting clones. Two of them encode large protein fragments highly homologous to rat GM130 and to human Golgin95. The full-length human GM130 cDNA was cloned and its interaction with rab1b was characterized in detail by yeast two-hybrid and in vitro binding assays. Here we report for the first time that the rab1b protein interacts specifically with GM130 in a GTP-dependent manner and therefore needs the hypervariable regions of the N- and C-termini. We mapped the rab1b binding site of GM130 and provide evidence that it is different to the previously described p115 and Grasp65 binding sites of the GM130 protein.

Inactive and active mutants of rab1b are not tightly integrated into target membranes.

The rab1b GTPase is localized on the ER and cis-Golgi membranes and involved in early exocytotic membrane traffic processes. To analyze the influence of rab1b conformation on specific membrane targeting processes three point mutants simulating permanently inactive or active rab1b proteins were constructed and expressed in BHK cells. To increase the expression of the growth inhibiting rab1b S22N and rab1b N121I mutants we co-expressed the Mss4 protein and observed an elevated expression of the inactive rab1b mutants. Surprisingly, only the rab1b wild-type protein shows the correct intracellular localization, and a tight membrane association. We conclude that the targeting process of rab1b depends predominantly on GDP/GTP exchange.