A Study on Vitamin D and Cathelicidin Status in Patients with Rosacea: Serum Level and Tissue Expression.

BACKGROUND: Rosacea is a chronic inflammatory disease characterized by centrofacial erythema. Excess cathelicidin is suggested to be important to the pathophysiology of the disease. Recently, presence of a vitamin D response element was revealed in the cathelicidin gene promoter. OBJECTIVE: The aim of this study was to determine whether vitamin D and cathelicidin are associated with rosacea, both serologically and histopathologically. METHODS: Subjects with rosacea and without chronic skin disorders were enrolled in the patient and control groups, respectively. Serum 25-hydroxy-vitamin D and cathelicidin levels were measured. Tissue expression of cathelicidin and vitamin D receptor were measured with immunostaining-intensity-distribution index. RESULTS: The mean serum 25-hydroxyvitamin D level of patients with rosacea was 12.18±5.65 ng/ml, which is lower than that of the controls (17.41±6.75 ng/ml). Mean serum cathelicidin levels in patients with rosacea and the controls were 85.0±26.1 ng/ml and 55.0±23.3 ng/ml, respectively. Cathelicidin expression in rosacea tissue was significantly higher than that in control tissue (5.21 vs. 4.03). No significant difference was observed in vitamin D receptor expression. CONCLUSION: Higher cathelicidin expression in rosacea supports the hypothesis that an abnormal inflammatory response of the innate immune system is important in pathogenesis of rosacea, but the role of high cathelicidin serum levels is complicated. Serum vitamin D was lower in patients with rosacea, although serum cathelicidin was higher than that of the controls. This suggests that the role of vitamin D level in the pathogenesis of rosacea merits further investigation.

Phosphatidylinositol-glycan-phospholipase D is involved in neurodegeneration in prion disease.

PrPSc is formed from a normal glycosylphosphatidylinositol (GPI)-anchored prion protein (PrPC) by a posttranslational modification. Most GPI-anchored proteins have been shown to be cleaved by GPI phospholipases. Recently, GPI-phospholipase D (GPI-PLD) was shown to be a strictly specific enzyme for GPI anchors. To investigate the involvement of GPI-PLD in the processes of neurodegeneration in prion diseases, we examined the mRNA and protein expression levels of GPI-PLD in the brains of a prion animal model (scrapie), and in both the brains and cerebrospinal fluids (CSF) of sporadic and familial Creutzfeldt-Jakob disease (CJD) patients. We found that compared with controls, the expression of GPI-PLD was dramatically down-regulated in the brains of scrapie-infected mice, especially in the caveolin-enriched membrane fractions. Interestingly, the observed decrease in GPI-PLD expression levels began at the same time that PrPSc began to accumulate in the infected brains and this decrease was also observed in both the brain and CSF of CJD patients; however, no differences in expression were observed in either the brains or CSF specimens from Alzheimer's disease patients. Taken together, these results suggest that the down-regulation of GPI-PLD protein may be involved in prion propagation in the brains of prion diseases.

Immunohistochemical study of arginase-1 in the spinal cords of Lewis rats with experimental autoimmune encephalomyelitis.

Arginase-1, a marker for M2 phenotype alternatively activated macrophages, inhibits inflammation and is associated with phagocytosis of cell debris and apoptotic cells. We analyzed the expression of arginase-1, a competitive enzyme of inducible nitric oxide synthase (iNOS), in the spinal cords of Lewis rats with experimental autoimmune encephalomyelitis (EAE). Western blot analysis showed that both arginase-1 and iNOS significantly increased in the spinal cords of rats at the peak stage of EAE compared with the expression level in control animals (p<0.05) and declined thereafter. Immunofluorescent staining demonstrated that increased expression of arginase-1 in EAE spinal cords was confirmed in macrophages as well as in some neurons and astrocytes that were constitutively positive for arginase-1 in normal spinal cords. A semiquantitative analysis by immunofluorescence showed that in EAE lesions, an increased level of arginase-1 immunoreactivity was matched with ED1-positive macrophages, which were also positive for activin A, a marker for the M2 phenotype. Taking all of these findings into consideration, we postulate that the increased level of arginase-1, which is partly from M2 macrophages, contributes to the modulation of neuroinflammation in EAE lesions, possibly through the reduction of nitric oxide in the lesion via competition with iNOS for the use of L-arginine.

Control of neurite outgrowth by RhoA inactivation.

cAMP induces neurite outgrowth in the rat pheochromocytoma cell line 12 (PC12). In particular, di-butyric cAMP (db-cAMP) induces a greater number of primary processes with shorter length than the number induced by nerve growth factor (NGF). db-cAMP up- and down-regulates GTP-RhoA levels in PC12 cells in a time-dependent manner. Tat-C3 toxin stimulates neurite outgrowth, whereas lysophosphatidic acid (LPA) and constitutively active (CA)-RhoA reduce neurite outgrowth, suggesting that RhoA inactivation is essential for the neurite outgrowth from PC12 cells stimulated by cAMP. In this study, the mechanism by which RhoA is inactivated in response to cAMP was examined. db-cAMP induces phosphorylation of RhoA and augments the binding of RhoA with Rho guanine nucleotide dissociation inhibitor (GDI). Moreover, RhoA (S188D) mimicking phosphorylated RhoA induces greater neurite outgrowth than RhoA (S188A) mimicking dephosphorylated form does. Additionally, db-cAMP increases GTP-Rap1 levels, and dominant negative (DN)-Rap1 and DN-Rap-dependent RhoGAP (ARAP3) block neurite outgrowth induced by db-cAMP. DN-p190RhoGAP and the Src inhibitor PP2 suppress neurite outgrowth, whereas transfection of c-Src and p190RhoGAP cDNAs synergistically stimulate neurite outgrowth. Taken together, RhoA is inactivated by phosphorylation of itself, by p190RhoGAP which is activated by Src, and by ARAP3 which is activated by Rap1 during neurite outgrowth from PC12 cells in response to db-cAMP.

Association of endothelial nitric oxide synthase and mitochondrial dysfunction in the hippocampus of scrapie-infected mice.

The elevation of nitric oxide (NO) within the central nervous system (CNS) is known to be associated with the pathogenesis of neurodegenerative diseases such as HIV-associated dementia (HAD), brain ischemia, Parkinson's disease, and Alzheimer's disease. NO is enzymatically formed by the enzyme nitric oxide synthase (NOS). There are two forms of NOS, the constitutive and the inducible form. The constitutive form is present in endothelial cells (eNOS) and neurons (nNOS). The inducible form (iNOS) is expressed in various cell types including astroglia and microglia of the CNS. Using an animal model, we investigated the involvement of eNOS in the pathology of prion disease. We showed dramatic upregulation of eNOS immunoreactivity in reactive astroglial cells in the hippocampus in the prion disease animal model, scrapie in mice. Expression of eNOS was upregulated in cytosolic and mitochondrial fractions of whole brain. In the hippocampal region, eNOS was widely overexpressed in various components of the cell. We found that eNOS dramatically accumulated in hippocampal mitochondria and was particularly prevalent in structurally dysfunctional mitochondria. In association with the accumulation of eNOS in mitochondria, we showed that mitochondrial superoxide dismutase (Mn-SOD or SOD2), cytochrome c, and ATP activity were downregulated both in whole brain and in the hippocampal region. These results indicate that eNOS plays a role in the development of dysfunctional mitochondria and this, in turn, could induce some of the histopathological changes seen in prion diseases.

Neurites from PC12 cells are connected to each other by synapse-like structures.

PC12 cells have been used as a model of sympathetic neurons. Nerve growth factor (NGF), basic fibroblast growth factor (bFGF), and cAMP induce neurite outgrowth from PC12 cells. cAMP induced a greater number of neurites than did NGF. In particular, we attempted to elucidate whether PC12 cell neurites, induced by several factors including NGF, bFGF, and cAMP, form synapses, and whether each neurite has presynaptic and postsynaptic properties. Using scanning electron microscopy (SEM) and transmission electron microscopy (TEM), we observed that neurites are connected to each other. The connected regions presented dense core vesicles and a clathrin-coated membrane invagination. In addition, typical maker proteins for axon and dendrite were identified by an immuno-staining method. Tau-1, an axonal marker in neurons, was localized at a high concentration in the terminal tips of neurites from PC12 cells, which were connected to neurite processes containing MAP-2, a dendritic marker in neurons. Furthermore, neurites containing SV2 and synaptotagmin, markers of synaptic vesicles, were in contact with neurites harboring drebrin, a marker of the postsynaptic membrane, suggesting that neurites from PC12 cells induced by NGF, bFGF, and cAMP may form synapse-like structures. Tat-C3 toxin, a Rho inhibitor, augmented neurite outgrowth induced by NGF, bFGF, and cAMP. Tat-C3 toxin together with neurotrophins also exhibited synapse-like structures between neurites. However, it remains to be studied whether RhoA inhibition plays a role in the formation of synapse-like structures in PC12 cells.

Glial cell line-derived neurotrophic factor is expressed by inflammatory cells in the sciatic nerves of Lewis rats with experimental autoimmune neuritis.

Neurotrophic factors, including glial cell line-derived neurotrophic factor (GDNF), have been known to play a role in neuroprotection in the injured peripheral nervous system (PNS). To evaluate the involvement of GDNF in experimental autoimmune neuritis (EAN) pathogenesis, the expression of GDNF in rat sciatic nerves with EAN was studied. Western blot analysis showed that the level of GDNF protein significantly increased 1.8-fold at the paralytic stage of EAN at day 12 post-immunization (PI) (p < 0.01), and its level further increased approximately 2.5-fold at day 21 PI (p < 0.001) in the sciatic nerves of EAN-affected rats compared with those of control rats, and then declined thereafter at day 28 PI. Immunohistochemical analysis showed that axons and Schwann cells constitutively contained GDNF in normal controls. In sciatic nerves with EAN at day 12 PI, GDNF was immunostained in infiltrating inflammatory cells including macrophages and T cells. Collectively, we postulate that GDNF plays a regulatory role in EAN paralysis. A paradoxical role of inflammatory cells to ameliorate PNS inflammation remains to be further studied in EAN, an animal model of human demyelinating disease.

Involvement of peptidylarginine deiminase-mediated post-translational citrullination in pathogenesis of sporadic Creutzfeldt-Jakob disease.

Peptidylarginine deiminases (PADs)-mediated post-translational citrullination processes play key roles in protein functions and structural stability through the conversion of arginine to citrulline in the presence of excessive calcium concentrations. In brain, PAD2 is abundantly expressed and can be involved in citrullination in disease. Recently, we have reported pathological characterization of PAD2 and citrullinated proteins in scrapie-infected mice, but the implication of protein citrullination in the pathophysiology in human prion disease is not clear. In the present study, we explored the molecular and biological involvement of PAD2 and the pathogenesis of citrullinated proteins in frontal cortex of patients with sporadic Creutzfeldt-Jakob disease (sCJD). We found increased expression of PAD2 in reactive astrocytes that also contained increased levels of citrullinated proteins. In addition, PAD activity was significantly elevated in patients with sCJD compared to controls. From two-dimensional gel electrophoresis and MALDI-TOF mass analysis, we found various citrullinated candidates, including cytoskeletal and energy metabolism-associated proteins such as vimentin, glial fibrillary acidic protein, enolase, and phosphoglycerate kinase. Based on these findings, our investigations suggest that PAD2 activation and aberrant citrullinated proteins could play a role in pathogenesis and have value as a marker for the postmortem classification of neurodegenerative diseases.

Immunohistochemical localization of galectin-3 in the pig retina during postnatal development.

PURPOSE: The differential level and localization of galectin-3 protein were examined in the retinas of two-day-old pigs and six-month-old pigs. METHODS: The retinas sampled from two-day-old and six-month-old pigs were analyzed by western blot and immunohistochemistry. RESULTS: western blot analysis detected galectin-3 in both age groups, although the levels were significantly higher in six-month-old pigs. Immunohistochemical staining showed that galectin-3 was localized in the retinas of both two-day-old pigs and six-month-old pigs; the galectin-3 immunostaining was more intense in the six-month-old pig retina, as shown in the western blot analysis. Galectin-3 was expressed in glial cells, particularly in glutamine synthetase-positive Müller cells and their processes, across all retina layers in both age groups; however, it was not found in ganglion cells of the ganglion cell layer or neuronal cells of the inner and outer nuclear cell layers in either age group. CONCLUSIONS: This is the first demonstration that galectin-3 is detected in the retinas of two-day-old pigs and that the expression in Müller cells increases with postnatal development.

Reduction of prion infectivity and levels of scrapie prion protein by lithium aluminum hydride: implications for RNA in prion diseases.

Previous studies indicate that RNA may be required for proteinase-resistant prion protein (PrP) amplification and for infectious prion formation in vitro, suggesting that RNA molecules may function as cellular cofactors for abnormal PrP (PrPSc) formation and become part of the structure of the infectious agent. To address this question, we used chemicals that can cleave phosphodiester bonds of RNA and assessed their effects on the infectious agent. Lithium aluminum hydride, a reducing agent that can induce reductive cleavage of oxidized molecules such as carbonyls, carboxyl acids, esters, and phosphodiester bonds, did not affect cellular PrP degradation; however, it destroyed PrPSc, extended the scrapie incubation period, and markedly reduced total RNA concentrations. These results prompted us to investigate whether RNA molecules are cofactors for PrPSc propagation. RNase A treatment of partially purified PrP and of 263K scrapie brain homogenates was sufficient to increase the sensitivity of PrPSc to proteinase K degradation. This is the first evidence that suggests that RNA molecules are a component of PrPSc. Treatment with RNase A alone and PrP degradation by RNase A plus proteinase K in vitro, however, did not result in loss of scrapie infectivity compared with the effects of lithium aluminum hydride. Together, these data suggest that RNA molecules may be important for maintaining the structure of PrPSc and that oxidized molecules can be important in scrapie agent replication and prion infectivity.

Immunohistochemical localization of protein kinase C-alpha in the retina of pigs during postnatal development.

The cellular localization and protein expression level of protein kinase C (PKC)-alpha was examined in pig retina at different ages. Western blot analysis detected PKC-alpha in the retinas of 3-day-old piglets and indicated significantly increased expression in 6-month-old young adult and 2-year-old adult pigs. Immunohistochemistry of 3-day-old retinas revealed intense PKC-alpha reactivity in the inner plexiform and inner nuclear cell layers, weak reactivity in the ganglion cell layer, and few positive cells in the outer nuclear cell layer. The cellular localization of PKC-alpha in the adult retina was similar, with staining more intense than that in neonates. PKC-alpha was co-localized in some glial fibrillary acidic protein-positive cells and glutamine synthetase-positive cells in the retina. This study demonstrates that the protein level of retinal PKC-alpha is increased with maturation and suggests that PKC-alpha plays a role in signal transduction pathways for postnatal development in porcine retina.

Increased neurogenesis in brains of scrapie-infected mice.

Persistent neurogenesis occurs in the adult brain throughout the life of all mammals. Recent studies have shown that neurogenesis was increased in adult gerbil and rat brains after ischemia. Neurogenesis has not been examined during neurodegenerative diseases such as scrapie. To investigate the regeneration of neurons after scrapie-infection, we infused 5-bromo-2'-deoxyuridine (BrdU), a DNA replication indicator, into both control and scrapie-infected mice. Mice were sacrificed at 150 days post-infection, i.e., at the start of clinical disease and a time when PrP(Sc) was readily detected in brain by both immunostaining and Western blot. We investigated expression of BrdU in each region of brain and observed cellular localization of BrdU using various cell markers such as neuronal nuclear (NeuN), microtubule-associated protein 2 (MAP2) and glial fibrillary acidic protein (GFAP). Immunohistochemically, BrdU-labeled cells were observed in the striatum, hippocampus, and brain stem of scrapie-infected brains. BrdU-labeled cells were much more prevalent in the hippocampus of scrapie-infected mice compared to hippocampus of control brains. In scrapie mice, there was more staining in hippocampus than in other brain regions. We also found that BrdU-positive cells colocalized with the neuronal markers NeuN and MAP2, whereas BrdU staining was not merged with GFAP, an astrocytic marker. Taken together, our results suggest that scrapie-infection induces region-specific increases in neuron regeneration.

Accumulation of citrullinated proteins by up-regulated peptidylarginine deiminase 2 in brains of scrapie-infected mice: a possible role in pathogenesis.

Peptidylarginine deiminases (PADs), which are a group of posttranslational modification enzymes, are involved in protein citrullination (deimination) by the conversion of peptidylarginine to peptidylcitrulline in a calcium concentration-dependent manner. Among the PADs, PAD2 is widely distributed in various tissues and is the only type that is expressed in brain. To elucidate the involvement of protein citrullination by PAD2 in the pathogenesis of brain-specific prion diseases, we examined the profiles of citrullinated proteins using the brains of scrapie-infected mice as a prion disease model. We found that, compared with controls, increased levels of citrullinated proteins of various molecular weights were detected in different brain sections of scrapie-infected mice. In support of this data, expression levels of PAD2 protein as well as its enzyme activity were significantly increased in brain sections of scrapie-infected mice, including hippocampus, brain stem, and striatum. Additionally, the expression levels of PAD2 mRNA were increased during scrapie infection. Moreover, PAD2 immunoreactivity was increased in scrapie-infected brains, with staining detected primarily in reactive astrocytes. Using two-dimensional electrophoresis and matrix-assisted laser desorption/ionization-time of flight mass spectrometry, various citrullinated proteins were identified in the brains of scrapie-infected mice, including glial fibrillary acidic protein, myelin basic protein, enolases, and aldolases. This study suggests that accumulated citrullinated proteins and abnormal activation of PAD2 may function in the pathogenesis of prion diseases and serve as potential therapeutic targets.

The effect of Fenton reaction on protease-resistant prion protein (PrPSc) degradation and scrapie infectivity.

In prion diseases, metal imbalances in brain and/or metal substitutions for copper in prion protein suggest that metal-catalyzed oxidation (MCO) and oxidative stress may affect cellular function and accumulation of protease-resistant prion protein (PrP(Sc)). We examined the effect of metal-induced oxidative stress by Fenton reaction on prion protein with regard to its degradation, insolubility, and infectivity. Precipitation and insolubility of prion protein were induced by Fenton reaction in scrapie-infected brain homogenate. Results showed an increase in hydroxylation products (thiobarbituric acid reactive substances; TBARS) and a decrease of ferrous ion (Fe(2+)) levels after Fenton reaction. Efficiency of metal-induced oxidation was higher for Fe(2+) than Mn(2+). Compared to untreated samples, there was increased susceptibility to proteolytic degradation of PrP(Sc) after treatment with 3.12-12.5 mM Fe(2+)-Mn(2+)/H(2)O(2). Interestingly, we observed that Fenton reaction could extend incubation periods, indicating a decrease in scrapie infectivity. These results suggest that PrP(Sc) hydroxylation and degradation may affect PrP conversion and the pathogenesis of prion diseases.

Differential regulation of apoptosis by caspase-mediated cleavage of phospholipase D isozymes.

Phospholipase D (PLD) has been implicated in survival and anti-apoptosis, but the molecular mechanism by which it responds to apoptotic stimuli is poorly unknown. Here, we demonstrate that cleavage of PLD isozymes as specific substrates of caspase differentially regulates apoptosis. PLD1 is cleaved at one internal site (DDVD(545)S) and PLD2 is cleaved at two or three sites (PTGD(13)ELD(16)S and DEVD(28)T) in the front of N-terminus. Cleavage of PLD was endogenously detected in post-mortem Alzheimer brain together with activated caspase-3, suggesting the physiological relevance. The cleavage of PLD1 but not PLD2 might act as an inactivating process since PLD1 but not PLD2 activity is significantly decreased during apoptosis, suggesting that differential cleavage of PLD isozymes could affect its enzymatic activity. Moreover, caspase-resistant mutant of PLD1 showed more potent anti-apoptotic capacity than that of wild type PLD1, whereas PLD2 maintained anti-apoptotic potency in spite of its cleavage during apoptosis. Moreover, PLD2 showed more potent anti-apoptotic effect than that of PLD1 in overexpression and knockdown experiments, suggesting that difference in anti-apoptotic potency between PLD1 and PLD2 might be due to its intrinsic protein property. Taken together, our results demonstrate that differential cleavage pattern of PLD isozymes by caspase might affect its enzymatic activity and anti-apoptotic function.

Transient impairment of hippocampus-dependent learning and memory in relatively low-dose of acute radiation syndrome is associated with inhibition of hippocampal neurogenesis.

Neurogenesis in the adult hippocampus, which occurs constitutively, is vulnerable to ionizing radiation. In the relatively low-dose exposure of acute radiation syndrome (ARS), the change in the adult hippocampal function is poorly understood. This study analyzed the changes in apoptotic cell death and neurogenesis in the DGs of hippocampi from adult ICR mice with single whole-body gamma-irradiation using the TUNEL method and immunohistochemical markers of neurogenesis, Ki-67 and doublecortin (DCX). In addition, the hippocampus-dependent learning and memory tasks after single whole-body gamma-irradiation were examined in order to evaluate the hippocampus-related behavioral dysfunction in the relatively low-dose exposure of ARS. The number of TUNEL-positive apoptotic nuclei in the dentate gyrus (DG) was increased 6-12 h after acute gamma-irradiation (a single dose of 0.5 to 4 Gy). In contrast, the number of Ki-67- and DCX-positive cells began to decrease significantly 6 h postirradiation, reaching its lowest level 24 h after irradiation. The level of Ki-67 and DCX immunoreactivity decreased in a dose-dependent manner within the range of irradiation applied (0-4 Gy). In passive avoidance and object recognition memory test, the mice trained 1 day after acute irradiation (2 Gy) showed significant memory deficits, compared with the sham controls. In conclusion, the pattern of the hippocampus-dependent memory dysfunction is consistent with the change in neurogenesis after acute irradiation. It is suggested that a relatively low dose of ARS in adult ICR mice is sufficiently detrimental to interrupt the functioning of the hippocampus, including learning and memory, possibly through the inhibition of neurogenesis.

JAK-STAT signaling pathway mediates astrogliosis in brains of scrapie-infected mice.

Scrapie is characterized histologically, in part, by astrogliosis in brain and spinal cord. However, the mechanisms of astrogliosis in brain injury occurring during prion infection are not well understood. In this study, we investigated the expression levels and cellular localization of Janus kinase (JAK) -signal transducers and activators of transcription (STAT) signaling molecules and growth factors such as leukemia inhibitory factor (LIF) and ciliary neurotropic factor (CNTF) by western blot analysis and immunohistochemistry. We found that expression levels of LIF and CNTF were increased in scrapie-infected brains and phosphorylated (p)-JAK2, p-STAT1 (Ser727 and Tyr701), p-STAT3 (Tyr705), and glial fibrillary acidic protein were expressed strongly in scrapie-infected brains. Moreover, we found that p-STAT1 and p-STAT3 were found mainly in the nucleus in scrapie-infected brains. Immunohistochemically, p-STAT1 was colocalized with LIF and CNTF and p-JAK2 in many reactive astrocytes in scrapie-infected brains. In contrast, immunostaining for p-STAT3 was found in comparatively few astrocytes in limited regions; p-STAT3 staining merged with p-JAK2 in hippocampus sections of scrapie-infected brains. Taken together, our results suggest that activation of JAK2-STAT1 signaling pathway occurred in reactive astrocytes in hippocampus of scrapie-infected brains.

Alteration of iron regulatory proteins (IRP1 and IRP2) and ferritin in the brains of scrapie-infected mice.

Considerable evidence suggests that oxidative stress may be involved in the pathogenesis of Transmissible Spongiform Encephalopathies (TSEs). To investigate the involvement of iron metabolism in TSEs, we examined the expression levels of iron regulatory proteins (IRPs), ferritins, and binding activities of IRPs to iron-responsive element (IRE) in scrapie-infected mice. We found that the IRPs-IRE-binding activities and ferritins were increased in the astrocytes of hippocampus and cerebral cortex in the brains of scrapie-infected mice. These results suggest that alteration of iron metabolism contributes to development of neurodegeneration and that some protective mechanisms against iron-induced oxidative damage may occur during the pathogenesis of TSEs.

Increased expression and localization of cyclooxygenase-2 in astrocytes of scrapie-infected mice.

A number of aspects of the pathogenesis of scrapie, the archetype disease of the transmissible spongiform encephalopathies (prion disorders), remain to be elucidated. There is increasing evidence that there are cerebral based inflammatory processes that may contribute to the pathogenesis and to the progression of a number of neurodegenerative disorders, including prion diseases. In peripheral tissues, a key element that controls the generation of proinflammatory mediators is the highly inducible protein cyclooxygenase-2 (COX-2). In this study, in order to examine the possible association of COX-2 with the pathogenesis of scrapie, we analyzed the expression level and the cellular localization of COX-2 in the brains of control and scrapie-infected mice. The COX-2 mRNA and protein levels were increased significantly compared to the control group of mice. By immunohistological analysis, intense immunoreactivity of COX-2 was localized primarily in reactive astrocytes, with virtually no staining in sections from control mice. The staining for COX-2 was co-localized with the pathological form of the prion protein (PrP(Sc)) and with nuclear factor-kappa B (NF-kappaB). These results suggest that the upregulation of COX-2 expression in astrocytes may be related to the accumulation of PrP(Sc), and that COX-2 may then lead to the progression of scrapie, possibly by propagation of a cerebral inflammatory response.

Galectin-3 expression is correlated with abnormal prion protein accumulation in murine scrapie.

To investigate the involvement of galectin-3 in the process of neurodegeneration in prion diseases, the expression and cellular localization of galectin-3 in the brain were studied in scrapie, a mouse model of prion disease. Reverse transcription-polymerase chain reaction (RT-PCR) and Western blot analyses showed that the expression of galectin-3 protein and mRNA was induced in scrapie-affected brains, particularly at the time when the abnormal prion protein PrP(Sc) began to accumulate in the brains. Immunohistochemically, immunostaining for galectin-3 was found mainly in B4-isolectin-positive cells (presumably activated microglia/macrophages), but not in astrocytes. Galectin-3 immunoreactivity was localized mainly in areas of PrP(Sc) accumulation and neuronal death in scrapie-infected brains. These findings suggest that the expression of galectin-3 by activated microglia/macrophages in prion disease correlates with abnormal prion protein accumulation.