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Aberrant expression of tight junction proteins has recently been focused on in the cancer research field. We previously showed that claudin-1 is aberrantly expressed from an early stage of uterine cervical adenocarcinoma and contributes to malignant potentials. To elucidate the molecular mechanisms underlying tumor-promoting roles of claudin-1, we established and analyzed claudin-1 knockout cells. Knockout of claudin-1 suppressed conventional tight junctional functions, barrier and fence functions, and expression of cell adhesion-associated proteins including E-cadherin. Comparative proteome analysis revealed that expression of claudin-1 affected expression of a wide range of proteins, especially proteins that are associated with cell adhesion and actin cytoskeleton remodeling. Interactome analysis of the identified proteins revealed that E-cadherin and focal adhesion kinase play central roles in the claudin-1-dependently affected protein network. Moreover, knockout of claudin-1 significantly suppressed microvilli formation and activity of Ezrin/Radixin/Moesin. Taken together, the results indicate that expression of claudin-1 affects not only conventional tight junction function but also expression and activity of a wide range of proteins, especially proteins that are associated with cell adhesion and actin cytoskeleton remodeling, to contribute to malignant potentials and microvilli formation in cervical adenocarcinoma cells.

In mammalian male germ-line cells, low-voltage-activated (LVA) Ca(2+) current has been identified and its electrophysiological properties have been studied. To investigate whether alpha(1)2.3 (alpha(1E)) subunit of the voltage-dependent Ca(2+) channel codes for the LVA current, whole-cell patch clamp and following reverse transcription-polymerase chain reaction (RT-PCR) experiments were performed in pachytene spermatocytes from Ca(v)2.3+/+ and Ca(v)2.3-/- mice. Whole-cell current in acutely dissociated pachytene spermatocytes from Ca(v)2.3+/+ and Ca(v)2.3-/- mice displayed a typical profile of LVA Ca(2+) currents and kinetics with no significant differences. Single-cell RT-PCR revealed the expression of Cacna1g in the pachytene spermatocytes from Ca(v)2.3+/+ and Ca(v)2.3-/- mice in which LVA Ca(2+) currents were actually recorded. These results suggest that the Ca(v)2.3 channel makes no detectable contribution to the LVA Ca(2+) current in the pachytene spermatocyte. Instead, Ca(v)3 family such as Ca(v)3.1 may be the likely candidates responsible for the LVA currents in pachytene spermatocytes.

Voltage-dependent Ca(2+) channels play important roles in cerebellar functions including motor coordination and learning. Since abundant expression of Ca(V)2.3 Ca(2+) channel gene in the cerebellum was detected, we searched for possible deficits in the cerebellar functions in the Ca(V)2.3 mutant mice. Behavioral analysis detected in delayed motor learning in rotarod tests in mice heterozygous and homozygous for the Ca(V)2.3 gene disruption (Ca(V)2.3+/- and Ca(V)2.3-/-, respectively). Electrophysiological analysis of mutant mice revealed perplexing results: deficit in long-term depression (LTD) at the parallel fiber Purkinje cell synapse in Ca(V)2.3+/- mice but apparently normal LTD in Ca(V)2.3-/- mice. On the other hand, the number of spikes evoked by current injection in Purkinje cells under the current-clamp mode decreased in Ca(V)2.3 mutant mice in a gene dosage-dependent manner, suggesting that Ca(V)2.3 channel contributed to spike generation in Purkinje cells. Thus, Ca(V)2.3 channel seems to play some roles in cerebellar functions.

The blood-brain barrier (BBB) is a biological unit composed of capillary endothelial cells and astrocytes. Here we examined the effects of various types of advanced glycation end-products (AGEs) on astrocytes and BBB-forming endothelial cells. While no type of AGE we examined changed the permeability of endothelial sheets, glyceraldehyde-derived AGE induced VEGF expression most significantly in astrocytes. The expression of glial cell line-derived neurotrophic factor (GDNF), which reduces the vascular permeability, was decreased in the astrocytes by treatment with glyceraldehyde-derived AGE. These results indicate that glyceraldehyde-derived AGE is the biologically active substance for astrocytes by regulating the VEGF and GDNF expression, which is causally contributing to an increase in the permeability of the BBB.