The 14-3-3 protein belongs to a family of 30-kD proteins originally identified by two-dimensional analysis of brain protein extracts. Recently, the detection of the 14-3-3 protein in the cerebrospinal fluid (CSF) is utilized as a highly reliable test for the premortem diagnosis of prion diseases such as Creutzfeldt-Jakob disease. For the initial step, to clarify the biological implication of the CSF 14-3-3 protein in these diseases, its expression was investigated in neural tissues and cultures and CSF samples from patients with a variety of neurological diseases by Western blot analysis and immunocytochemistry. The constitutive expression of the 14-3-3 protein was identified in all neural and nonneural tissues examined. It was expressed in all neurons, astrocytes, oligodendrocytes, and microglia in culture with its location in both cytoplasmic and nuclear regions. The 14-3-3 protein was detected in the CSF of 8 out of 71 patients, including 1 Gerstmann-Sträussler-Scheinker disease patient and 7 patients with prion-unrelated neurological diseases, such as meningoencephalitis of viral, bacterial, or tuberculous origin, multiple sclerosis, and mitochondrial myopathy, encephalopathy, lactic acidosis, and strokelike episodes. These results suggest that the 14-3-3 protein expressed constitutively at substantial levels in both neurons and glial cells might be released into the CSF as a disease-nonspecific consequence of the extensive brain damage and indicate that the analysis of the 14-3-3 protein in the CSF is not useful as a screening test for prion diseases.
Prion diseases are a group of neurodegenerative disorders characterized by intracerebral accumulation of a protease-resistant prion protein (PrP(Sc)) that causes extensive neuronal degeneration and astrogliosis. The regulation of prion protein (PrP) gene expression by a panel of glial and neuronal cytokines (TNF-alpha, IFN-gamma, IL-1beta, IL-10, and TGF-beta1) was investigated in human neural cell lines by reverse transcription-polymerase chain reaction and Northern blot analysis. The constitutive expression of PrP mRNA was identified in all human neural cell lines and tissues examined including Y79 retinoblastoma, IMR-32 neuroblastoma, SK-N-SH neuroblastoma, U-373MG astrocytoma, KG-1-C glioma, NTera2 teratocarcinoma, NTera2-derived differentiated neurons (NTera2-N), peripheral nerve, and cerebral and cerebellar tissues. In SK-N-SH cells, a 48 hour (h) treatment with 100 ng/ml IL-1beta, 100 ng/ml TNF-alpha, or 100 nM phorbol 12-myristate 13-acetate induced a 2.7- to 4.2-fold increase in the level of PrP mRNA, while the exposure to 100 ng/ml IFN-gamma resulted in a 50% decrease. By contrast, none of these cytokines significantly altered the levels of PrP mRNA in IMR-32, NTera2-N, or U-373MG cells. These results indicate that the PrP gene expression is constitutive in a wide range of human neural cell lines and tissues where it is controlled by cell type-specific regulatory mechanisms.
The expression of mRNAs for neurotrophins and neurotrophin receptors was examined in NTera2kl.Dl (NT2) human embryonal carcinoma cells at various stages of retinoic acid (RA)-induced neuronal differentiation using reverse transcription-polymerase chain reaction and southern blot analysis in order to investigate the biological role of neurotrophins in the human neuronal differentiation. The expression of high steady levels of BDNF, NT-3, NT-4, and low-affinity NGF receptor mRNAs was observed in both untreated NT2 cells and NT2-derived differentiated neurons induced by RA treatment. The lower levels of expression of NGF, trkA, trkB, and trkC mRNAs were identified in untreated NT2 cells and their expression was elevated markedly following 4 week treatment with RA. The increased expression of NGF, trkB, and trkC mRNAs were stable in NT2-derived differentiated neurons, while the expression of trkA mRNA was reduced substantially in these cells. These results indicate that mRNAs for a battery of neurotrophins and their receptors are coexpressed in NT2 cells and their expression is regulated by RA-induced neuronal differentiation.
A 51-year-old woman developed a slowly progressive spastic paraparesis and diminished vibration sense beginning at age 38. Intellectual capacity was normal. Krabbe disease was confirmed by markedly reduced leukocyte galactocerebrosidase (GALC) activity, typical inclusions in Schwann cell cytoplasm, and an identification of the homozygous point mutation T1835C (Leu618Ser) in the GALC gene. T2-weighted MRI of the brain showed symmetric high-signal-intensity lesions in the bilateral frontoparietal white matter, the centrum semiovale, and the posterior limb of the internal capsule with sparing of the periventricular white matter. This case is unusual because of the late onset, protracted clinical course, and MRI findings of demyelination confined to the corticospinal tracts.
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