Propolis induces AQP3 expression: A possible way of action in wound healing

Propolis is the generic name of a complex of resinous compound collected by honeybees and it has been utilized for many years in folk medicine. As other products generated by honeybees (such as royal jelly, pollen, honey), propolis has great therapeutic properties, but very little scientific information is available. Therefore, this study was aimed at exploring the potential wound healing properties of propolis. To that end, we utilized an in vitro scratch wound healing model consisting of human immortalized keratinocytes. Our scratch wound data clearly demonstrated that propolis induced a pronounced increase in the wound repair abilities of keratinocytes. A cell migration assay showed that propolis stimulated keratinocytes to close the wound. We revealed the role of H2O2 as the main mediator of propolis regenerative properties. We showed that this extracellularly released H2O2 could pass across the plasma membrane through a specific aquaporin (i.e., AQP3) modulating intracellular responses. The data offer a biological characterization of propolis positive effects suggesting that propolis could also be utilized in wound treatment within clinical settings

Human sperm functioning is related to the aquaporin-mediated water and hydrogen peroxide transport regulation

Aquaporins (AQPs) are transmembrane water channels and some of them are permeable in addition to water to other small solutes including hydrogen peroxide. The sperm cells of mammals and fishes express different AQPs, although there is no agreement in the literature on their localization. In humans, AQP3 and AQP11 are expressed mainly in the tail, AQP7 in the head and AQP8 in the midpiece. Thanks to the results of experiments with KO mice and to data obtained by comparing sub-fertile patients with normospermic subjects, the importance of AQPs for the normal functioning of sperms to ensure normal fertility emerged. AQP3, AQP7 and AQP11 appeared involved in the sperm volume regulation, a key role for fertility because osmoadaptation protect the sperm against a swelling and tail bending that could affect sperm motility. AQP8 seems to have a fundamental role in regulating the elimination of hydrogen peroxide, the most abundant reactive oxygen species (ROS), and therefore in the response to oxidative stress. In this review, the human AQPs expression, their localization and functions, as well as their relevance in normal fertility are discussed. To understand better the AQPs role in human sperm functionality, the results of studies obtained in other animal species were also considered

In vitro systems for studying thiamin transport in mammals.

This chapter discusses in vitro systems employed for studying thiamin transport in mammals. In mammals the transport of low (physiological) concentrations of thiamin is a carrier-mediated, mainly energy-dependent process that, in vitro, can be studied in preparations with different levels of anatomical complexity (intact tissues, isolated cells, and membrane vesicles). Each of these allows different aspects of the transport mechanism to be investigated. Thus intact tissue preparations (everted jejunal sacs or rings, human small intestinal biopsies, and brain slices) can be used to study biotransformations of thiamin related to transport. Isolated cell preparations both from normal (hepatocytes, enterocytes, and erythrocytes) and pathological tissues (Ehrlich ascites tumor and neuroblastoma cells) can also be used, according to the particular problem to be assessed. Choroid plexus and placenta preparations, as well as blood–brain barrier, can give insight into the modalities of thiamin transfer from the plasma through complex biological structures. By contrast, membrane preparations (small intestinal brush border or basolateral membrane vesicles, brain microsacs, and erythrocyte ghosts) are well suited for characterizing pure membrane thiamin transport mechanism without the interference of intracellular thiamin metabolism

Distribution of thiamine, thiamine phosphates, and thiamine metabolizing enzymes in neuronal and glial cell enriched fractions of rat brain.

The distribution of thiamine, thiamine phosphoesters, and the thiamine pyrophosphate synthetizing [thiamine-pyrophosphokinase (TPKase)] as well as hydrolyzing [thiamine pyrophosphatase (TPPase) and thiamine monophosphatase (TMPase)] enzymes was determined in neuronal and glial enriched fractions prepared from rat brain. Nucleoside diphosphatases [inosine diphosphatase (IDPase) and uridine diphosphatase (UDPase)] and nucleoside monophosphatases [uridine monophosphatase (UMPase) and inosine monophosphatase (IMPase)] were also determined. Thiamine and thiamine mono- and pyrophosphate were present in neuronal enriched fractions at concentrations 2.8, 3.6, and 4.6 times higher than in glial fractions. TMPase was found only in glial enriched fractions, whereas the levels of TPKase, UMPase, IMPase, IDPase, UDPase, and TPPase were 2.0-, 2.2-, 1.3-, 2.8-, 3.7-, and 20.8-fold higher in neuronal than in glial fractions

Thiamine outflow from the enterocyte: a study using basolateral membrane vesicles from rat small intestine.

1. Rat small intestinal basolateral membrane vesicles (BLMVs) were prepared and found to be 31% non-vesiculated and 69% vesiculated, 4.9% right side out and 63.8% inside out. 2. Thiamine uptake by BLMVs followed a hyperbolic time course reaching equilibrium after 60-90 min incubation. Uptake was not affected by the transmembrane potential or by the presence or absence of Na+ or K+ in the incubation medium. 3. At concentrations below 1.25 microM, [3H]thiamine was taken up mainly by a saturable mechanism with an apparent Michaelis-Menten constant (Km) = 1.32 microM and maximal flux (Jmax) = 1.93 pmol (mg protein)-1 (4 s)-1. At higher concentrations, a non-saturable mechanism prevailed. 4. Only 29% of [3H]thiamine taken up by the vesicles was membrane bound, the remaining being translocated into the vesicular space. No thiamine phosphoesters could be detected inside the vesicles. 5. In the absence of ATP, the Na(+)-K(+)-ATPase inhibitors ouabain, frusemide and vanadate reduced thiamine uptake by 35, 30 and 15% respectively. 6. In experiments conducted with K+ inside the vesicles and Na+, Mg2+ and ATP outside, the time course of thiamine uptake by BLMVs displayed an overshoot (80-90% increment) at 30 s incubation as compared to controls. When ATP was replaced with phosphocreatine, or when NaCl was replaced with isosmotic amounts of KCl, the overshoot disappeared. 7. The thiamine analogues pyrithiamine, amprolium and 4'-oxythiamine decreased the ATPase-dependent transport of [3H]thiamine by 100, 86 and 31% respectively. 8. These results provide evidence that the transport of thiamine by BLMVs is coupled directly to the hydrolysis of ATP (primary active transport)

Aquaporin-10 Represents an Alternative Pathway for Glycerol Efflux from Human Adipocytes

BACKGROUND: Glycerol outflow from adipocytes has been considered for a decade to be mediated by aquaporin-7, an aquaglyceroporin highly expressed in the adipose tissue. Its involvement in glycerol metabolism has been widely studied also in humans. Recent studies in different aquaporin-7 KO mice models pose two different questions 1) the exact localization of aquaporin-7 in human white adipose tissue; 2) the existence of other aquaglyceroporins that work with aquaporin-7 to guarantee glycerol efflux and thus a normal adiposity in humans. To this purpose we investigated the expression, the localization and the functioning of aquaglyceroporin-10 in subcutaneous white adipose tissue, in isolated and cultured differentiated adipocytes. METHODOLOGY/PRINCIPAL FINDINGS: Aquaporin-7 and -10 were expressed in the white adipose tissue both at mRNA and at protein level. Immunofluorescence revealed aquaporin-7 and -10 labelling in the human adipose tissue both to the plasma membrane and to a thin rim of cytoplasm of adipocytes. Aquaporin-7, but not aquaporin-10, colocalized with the endothelial marker CD34. Human cultured differentiated adipocytes showed an aquaporin-7 and -10 labelling mainly in the cytoplasm and in the lipid droplets with insulin reinforcing the lipid droplets staining and isoproterenol inducing its translocation to the plasma membrane compartment. Water and glycerol permeability measurements using adipocytes and adipose membrane vesicles confirmed the presence of functioning aquaglyceroporins. Aquaporin-10 silencing in human differentiated adipocytes resulted in a 50% decrease of glycerol and osmotic water permeability. CONCLUSIONS/SIGNIFICANCE: The results indicate that aquaporin-7, differently from mice, is present in both adipocyte and capillary plasma membranes of human adipose tissue. Aquaporin-10, on the contrary, is expressed exclusively in the adipocytes. The expression of two aquaglyceroporins in human adipose tissue is particularly important for the maintenance of normal or low glycerol contents inside the adipocyte, thus protecting humans from obesity