AP160 Sigma-AldrichRabbit Anti-Mouse IgG Antibody

The relationship between dopaminergic neuronal structures and dopaminoceptive structures in the amphibian brain and spinal cord are assessed by means of single and double immunohistochemical techniques with antibodies directed against DARPP-32 (a phosphoprotein related to the dopamine D(1)-receptor) and tyrosine hydroxylase (TH) applied to the brain of the anurans Rana perezi and Xenopus laevis. The DARPP-32 antibody yielded a well-differentiated pattern of staining in the brain of these anurans. In general, areas that are densely innervated by TH-immunoreactive fibers such as the nucleus accumbens, striatum, amygdaloid complex, thalamus, optic tectum, torus semicircularis and spinal cord display a remarkable immunoreactivity for DARPP-32 in cell bodies and neuropil. Distinct cellular DARPP-32 immunoreactivity was also found in the septum, preoptic area, suprachiasmatic nucleus, tuberal hypothalamic region, habenula, retina, midbrain tegmentum, rhombencephalic reticular formation and solitary tract nucleus. Hodological data supported that striatal projection neurons were DARPP-32 immunoreactive. Double immunohistofluorescence staining revealed that catecholaminergic cells generally do not stain for DARPP-32, except for some cells in the ventral mesencephalic tegmentum of Xenopus and cells in the nucleus of the solitary tract of Rana. Several interspecies differences were noted for the DARPP-32 distribution in the brain of the two anurans, namely in the habenula, the thalamus and prethalamus, the cerebellum and octavolateral area and the structures with DARPP-32/TH colocalization. However, in general, the distribution of DARPP-32 in the brain of the anuran amphibians resembles in many aspects the pattern observed in amniotes, especially in reptiles.

Hepatocyte transplantation (Tx) holds promise for curing genetic liver diseases. However, a limited number of donor hepatocytes can be transplanted into the host liver. Recipient preconditioning and donor cell engineering are under investigation to improve cell engraftment. In theory, genetically engineered cells secreting therapeutic proteins with superior function could compensate for poor engraftment efficiency. We have generated a bioengineered human coagulation factor IX (FIX) with augmented specific activity (named FIX-Triple). The aim of this study was to evaluate therapeutic efficacy of cell therapy using hemophilia B (HB) as a disease model by transplanting FIX-Triple-secreting hepatocytes. The donor hepatocytes were isolated from FIX-Triple knock-in (KI) or FIX-WT (wild-type) KI mice and transplanted intrasplenically into FIX knock-out (KO) mice. FIX-Triple KI recipients exhibited fourfold higher plasma FIX clotting activity than FIX-WT KI recipients. By repeated Txs, the clotting activity of FIX-Triple KI recipients even increased to more than 10% of normal mouse plasma. The engraftment and FIX production efficiencies of transplanted cells were equivalent between the FIX-WT KI and FIX-Triple KI donors. A hemostatic function assay showed that FIX-Triple KI recipients with repeated Txs had more enhanced clot kinetics and a greater maximum rate of thrombus generation than those with a single Tx. Moreover, FIX inhibitors in these recipients rarely developed. In conclusion, hepatocyte Tx with genetically engineered donor cells is an effective therapeutic strategy for HB.

The primary goal of this study was to examine whether the locus coeruleus (LC) provides collateral projections to whisker-related, sensorimotor brain regions. After injections of retrograde tracers into the primary sensory (S1) barrel field/primary whisker motor (M1) cortices, ventroposteromedial (VPM)/ventrolateral (VL) thalamic nuclei, or principal sensory trigeminal (Pr5)/facial motor (Mo7) nuclei, the distribution of double-labeled neurons within the LC was examined. Our observations indicated that a large number of individual LC cells provided axon collaterals to S1-M1 or VPM-VL regions, whereas only a few projected to Pr5-Mo7 nuclei. The laterality and the distribution of dual-projecting LC neurons were as follows. 1) The neurons projecting to the S1-M1 cortices were predominantly ipsilateral (96% +/- 0.7%). Labeled neurons were located ventrally at the rostral pole but were evenly distributed along the dorsoventral aspect of the principal LC. 2) The cells projecting to the VPM-VL nuclei were bilateral, with ipsilateral (68% +/- 2.3%) dominance. Neurons were observed at the rostrocaudal extent of the LC, where the labeling was most pronounced at the ventral, principal LC. 3) The neurons projecting to the Pr5-Mo7 regions exhibited slightly contralateral (56% +/- 2.9%) dominance, where labeled cells were confined within the ventral margin of the principal subdivision. Taken together, the present observations demonstrate that each subdivision of the LC possesses a differential functional organization with respect to its collateral projection to whisker-related sensorimotor targets, suggesting that the nucleus might play a modulatory role in vibrissal sensorimotor integration that allows the guidance of behavioral action essential for the survival of the animal.

Inactivating mutations of the tumor-suppressor kinase gene LKB1 underlie Peutz-Jeghers syndrome (PJS), which is characterized by gastrointestinal hamartomatous polyps with a prominent smooth-muscle and stromal component. Recently, it was noted that PJS-type polyps develop in mice in which Lkb1 deletion is restricted to SM22-expressing mesenchymal cells. Here, we investigated the stromal functions of Lkb1, which possibly underlie tumor suppression. Ablation of Lkb1 in primary mouse embryo fibroblasts (MEFs) leads to attenuated Smad activation and TGFbeta-dependent transcription. Also, myofibroblast differentiation of Lkb1(-/-) MEFs is defective, resulting in a markedly decreased formation of alpha-smooth muscle actin (SMA)-positive stress fibers and reduced contractility. The myofibroblast differentiation defect was not associated with altered serum response factor (SRF) activity and was rescued by exogenous TGFbeta, indicating that inactivation of Lkb1 leads to defects in myofibroblast differentiation through attenuated TGFbeta signaling. These results suggest that tumorigenesis by Lkb1-deficient SM22-positive cells involves defective myogenic differentiation.