Ultrastructural localization of adhalin in normal murine skeletal myofiber

The ultrastructural localization of adhalin and its relations to dystrophin, β-dystroglycan, and β-spectrin were studied in normal murine skeletal myofibers. The C-terminal peptides of adhalin and β-dystroglycan were synthesized based on their cDNAs, and the affinity-purified antibodies against these peptides were produced. Single-immunolabeling electron microscopy showed that the adhalin was located just inside the muscle plasma membrane or inside the myofiber a short distance from the plasma membrane. The adhalin signal was also noted at the sarcoplasmic side of plasmalemmd invaginations or at vesicular structures in subsarcolemmal areas. Double-immunogold-labeling electron microscopy disclosed a similar localization of dystrophin, β-dystroglycan, and β-spectrin. The close association of adhalin with dystrophin or β-dystroglycan was demonstrated by formation of doublets by signals of antibodies of adhalin with those of dystrophin or β-dystroglycan and was confirmed by statistical analyses. This study demonstrated that the location of adhalin is close to that of dystrophin and β-dystroglycan at the muscle plasma membrane.     

Functional morphology and physiological properties of bronchopulmonary C-fiber afferents

Nonmyelinated (C-) fibers represent the majority of vagal afferents innervating the airways and lung, and play an important role in regulating the respiratory and cardiovascular functions under both normal and abnormal physiologic conditions. Studies of the relationship between the conduction velocities of the vagal afferents and their sensitivities to capsaicin and other chemical irritants reveal that C-fibers are the primary type of chemosensitive afferents in the rat lung. Furthermore, a distinct sensitivity to capsaicin and a weak response to lung inflation are the defining characteristics of these afferents. In cultured rat nodose and jugular ganglion neurons, capsaicin-sensitive cells were identified by measurement of the capsaicin-evoked calcium transients using the Fura-2-based ratiometric imaging technique. The percentage of capsaicin-sensitive neurons gradually decreases as the cell diameter increases. However, the capsaicin-sensitive neurons cannot be precisely identified solely on the basis of the cell size. Anandamide, an endogenous cannabinoid released from leukocytes and epithelial cells, consistently evokes a stimulatory effect on pulmonary C-fiber endings by activating vanilloid receptor type 1 (VR1). The discharge pattern of pulmonary C-fibers evoked by anandamide closely resembles that produced by a much lower ( approximately 1/600) dose of capsaicin in the same fibers. Whether anandamide acts as a potential endogenous ligand to VR1 at the C-fiber terminals is unclear, and the physiological role of VR1 in modulating the transduction properties of these afferents also remains to be determined.     

Proliferation of intrahepatic bile-duct epithelium in biliary atresia

Proliferating cell nuclear antigen (PCNA) and transforming growth factor (TGF) are considered as markers of cell proliferation. The expression of PCNA and TGF was evaluated immunohistochemically using anti-PCNA antibody and TGF in 31 patients with biliary atresia (BA) (15 jaundice-free and 16 with persistent jaundice) and 6 control infants. The labeling indices (LI) for PCNA- and TGF-positive bile-duct epithelium in BA were 14.1±14.0% and 51.4±33.7%, respectively, which was significantly higher than in the controls (P <0.01). In BA, the number of PCNA-immunoreactive cells was higher in the peripheral bile ductules than in the central bile ducts of the portal tract (P <0.01). LI was not related to patient age at the time of hepatic portoenterostomy in two groups divided at the age of 60 days. Patients in the persistent jaundice group had greater expression of PCNA and TGF (21.7±16.0% and 76.9±20.7%, respectively) compared to those in the jaundice-free group (6.0±2.7% and 24.3±20.9%, P <0.001). PCNA and TGF expression in the bile-duct epithelium of the portal tract was closely related to prognosis in BA patients, and thus could be useful as a prognostic marker.     

Possible Cerebral Vasculitis in a Case with Rheumatoid Arthritis

Cerebral rheumatoid vasculitis (CRV) is a rare, fatal, and diagnostically challenging disorder. We herein report an 81-year-old woman with a 4-year history of rheumatoid arthritis who presented with a fever, progressive disturbance of consciousness, high level of rheumatoid factor, and hypocomplementemia. The enhancement of the perforating branches in the left middle cerebral artery led us to suspect CRV. A brain biopsy could not be performed. After we intensified steroid therapy, the size of the cerebral lesions temporarily decreased. However, recurrence in the left frontal lobe occurred one month later, and the patient subsequently died. Early intensive treatments may be needed for CRV.     

APP knockout attenuates microglial activation and enhances neuron survival in substantia nigra compacta after axotomy

Focal microglial activation and progressive dopaminergic neurodegeneration in substantia nigra compacta (SNc) have characterized Parkinson's disease (PD). We have hypothesized that the microglial response may be provoked by molecular signals from chronically stressed SNc neurons. To test whether amyloid precursor protein (APP) could serve as such a signal, we evaluated microglial activation in SN after unilateral transection of the medial forebrain bundle (MFB) in mice either wild-type (WT) or null (KO) for APP. WT and KO mice displayed comparable microglial response at the MFB transection site. In WT mice microglial activation was first apparent in the ipsilateral SN at 3 days postlesion (dpl), marked by morphological change and increased isolectin immunoreactivity. The microglial response intensified at 7 dpl and persisted in the medial nigra through 14 dpl. In contrast, in KO mice activated microglia appeared predominantly at 7 dpl, with little activation at 3 dpl and none at 14 dpl. Neuron number in affected WT SNc at 14 dpl was significantly reduced compared with loss in affected KO SNc. The delayed and limited local microglial activation and increased neuron survival in response to distal axotomy of SNc neurons in APP KO mice are consistent with the important role APP in neuronal stress responses in vivo.     

Observations of muscle plasma membrane undercoats in Duchenne and fukuyama muscula dystrophies

Muscle plasma membrane undercoats were investigated by conventional electron microscopy in both Duchenne muscular dystrophy (DMD) and Fukuyama congenital muscular dystrophy (FCMD). The densities of the plasma membrane undercoats were rarefied in the parts of the plasma membranes overlying the degenerating focus in both DMD and FCMD myofibers. The degree of rarefaction tended to be parallel to the degree of degeneration in the myofibers. It was hard to distinguish the undercoat densities of normal-looking myofibers of DMD and FCMD muscles from those of control myofibers from histochemically-normal muscles. On the other hand, the undercoats of regenerating myofibers in DMD and FCMD muscles were denser than normal.     

Ultrastructural localization of aquaporin 4 and alpha1-syntrophin in the vascular feet of brain astrocytes

Aquaporin 4 (AQP4) is a recently discovered membrane bound water-selective channel and has been described at the light microscopic level to be predominantly expressed in the astrocytes of the brain, especially at the perivascular astrocyte endfoot processes. Alpha1-syntrophin, a member of dystrophin-associated protein, has also been reported at the light microscopic to be expressed level in the same site of astrocytes as AQP4 and interacts with other molecules through its PDZ domain. AQP4 expression has been reported to be absent at the sarcolemma and the perivascular astrocyte endfoot processes of alpha1-syntrophin knockout mice. Based on these observations, the molecular association between AQP4 and alpha1-syntrophin could be speculated. To test this hypothesis, we investigated the ultrasturctural localization of AQP4 and alpha1-syntrophin in the brain astrocytes by using double immunogold labeled electron microscopy. The results showed that AQP4 and alpha1-syntrophin colocalized frequently at the astrocyte membrane, especially at the perivascular astrocyte endfoot processes and suggested the presence of linkage between AQP4 and alpha1-syntrophin at the astrocyte plasma membrane.