Dipeptidyl peptidase IV (DPPIV) is a serine protease, a member of the prolyl oligopeptidase (POP) family, and has been implicated in several diseases. Therefore, it seems important to develop selective inhibitors for human DPPIV (hDPPIV) that are able to control the biological function of hDPPIV. In order to elucidate the binding mode and substrate specificity, we determined the crystal structure complex of hDPPIV and diprotin A (IIe-Pro-IIe), a slowly hydrolyzed substrate of hDPPIV, at 2.2 A resolution. In this paper, we discuss the molecular interaction mechanism of diprotin A with hDPPIV based on the X-ray crystal structure.

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BackgroundWe evaluated the utility of L-3,4-dihydroxy-6-[18F]fluoro-phenylalanine ([18F]FDOPA) positron emission tomography (PET) as a method for assessing the severity of dopaminergic dysfunction in unilaterally 6-hydroxydopamine (6-OHDA)-lesioned rats by comparing it with quantitative biochemical, immunohistochemical, and behavioral measurements.MethodsDifferent doses of 6-OHDA (0, 7, 14, and 28 μg) were unilaterally injected into the right striatum of male Sprague-Dawley rats. Dopaminergic functional activity in the striatum was assessed by [18F]FDOPA-PET, measurement of striatal dopamine (DA) and DA metabolite levels, tyrosine hydroxylase (TH) immunostaining, and methamphetamine-induced rotational testing.ResultsAccumulation of [18F]FDOPA in the bilateral striatum was observed in rats pretreated with both aromatic L-amino acid decarboxylase and catechol-O-methyltransferase (COMT) inhibitors. Unilateral intrastriatal injection of 6-OHDA produced a significant site-specific reduction in [18F]FDOPA accumulation. The topological distribution pattern of [18F]FDOPA accumulation in the ipsilateral striatum agreed well with the pattern in TH-stained corresponding sections. A significant positive relationship was found between Patlak plot Ki values and striatal levels of DA and its metabolites (r = 0.958). A significant negative correlation was found between both Ki values (r = -0.639) and levels of DA and its metabolites (r = -0.719) and the number of methamphetamine-induced rotations.ConclusionsKi values determined using [18F]FDOPA-PET correlated significantly with the severity of dopaminergic dysfunction. [18F]FDOPA-PET makes it possible to perform longitudinal evaluation of dopaminergic function in 6-OHDA-lesioned rats, which is useful in the development of new drugs and therapies for Parkinson's disease (PD).

Glycoprotein M6A (GPM6A) is known as a transmembrane protein and an abundant cell surface protein on neurons in the central nervous system (CNS). However, the function of GPM6A in the differentiation of neurons derived from human embryonic stem (ES) cells is unknown. To investigate the function of GPM6A in neural differentiation, we generated human ES cell lines with overexpressed (B2h-oeM6A) or suppressed (B2h-shM6A) human GPM6A. Real-time polymerase chain reaction (PCR) showed that overexpression of GPM6A markedly increased the expression of neuroectodermal-associated genes (OTX1, Lmx1b, En1, Pax2, Sox2, and Wnt1), and the number of neural stem cells (NSCs) derived from B2h-oeM6A cells compared to control vector transfected human ES cells (B2h-Mock1). Our results show an increase in the number of differentiated neuronal cells (cholinergic, catecholaminergic, and GABAergic neurons) from NSCs derived from B2h-oeM6A cells. On the other hand, suppression of human GPM6A expression using a short hairpin RNA (shRNA) in human ES cells led to a decrease in both the expression of neuroectodermal-associated genes and the number of NSCs derived from B2h-shM6A cells. In addition, our results show a decrease in the number of differentiated neuronal cells from NSCs in B2h-shM6A cells compared to control vector transfected human ES cells (B2h-shNSP1). Moreover, overexpression or suppression of human GPM6A in human ES cells led to an increase or decrease, respectively, of neuronal migration. Hence, our findings suggest that expression level of GPM6A is, directly or indirectly, associated with the differentiation and neuronal migration of neurons derived from undifferentiated human ES cells.

Glycoprotein M6A (GPM6A) is known as a transmembrane protein and an abundant cell surface protein on neurons in the central nervous system (CNS). However, the function of GPM6A is still unknown in the differentiation of neurons derived from embryonic stem (ES) cells. To investigate the function of GPM6A, we generated knockdown mouse ES cell lines (D3m-shM6A) using a short hairpin RNA (shRNA) expression vector driven by the U6 small nuclear RNA promoter, which can significantly suppress the expression of mouse GPM6A mRNA. Real-time polymerase chain reaction (real-time PCR) and immunocytochemical analysis showed that expression of shRNA against GPM6A markedly reduced the expression of neuroectodermal-associated genes (OTX1, Lmx1b, En1, Pax2, Pax5, Sox1, Sox2, and Wnt1), and also the number of neural stem cells (NSC) derived from D3mshM6A cells compared to control vector-transfected mouse ES cells (D3m-Mock). Moreover, our results show a decrease in both the number of neuronal markers and the number of differentiating neuronal cells (cholinergic, catecholaminergic, and GABAergic neurons) from NSC in D3m-shM6A cells. Hence, our findings suggest that expression level of GPM6A is directly or indirectly associated with the differentiation of neurons derived from undifferentiated ES cells.